Mercurial > repos > iarc > mutspec
diff mutspecStat.pl @ 7:eda59b985b1c draft default tip
Uploaded
| author | iarc |
|---|---|
| date | Mon, 13 Mar 2017 08:21:19 -0400 |
| parents | 46a10309dfe2 |
| children |
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--- a/mutspecStat.pl Tue Jun 28 02:59:32 2016 -0400 +++ b/mutspecStat.pl Mon Mar 13 08:21:19 2017 -0400 @@ -1,3340 +1,2767 @@ -#!/usr/bin/env perl - -#-----------------------------------# -# Author: Maude # -# Script: mutspecStat.pl # -# Last update: 16/06/16 # -#-----------------------------------# - -use strict; -use warnings; -use Getopt::Long; -use Pod::Usage; -use File::Basename; # my ($filename, $directories, $suffix) = fileparse($file, qr/\.[^.]*/); -use File::Path; -use Statistics::R; -use Spreadsheet::WriteExcel; - -our ($verbose, $man, $help) = (0, 0, 0); # Parse options and print usage if there is a syntax error, or if usage was explicitly requested. -our ($refGenome, $output, $folder_temp, $path_R_Scripts, $path_SeqrefGenome) = ("empty", "empty", "empty", "empty", "empty"); # The reference genome to use; The path for saving the result; The path for saving the temporary files; The path to R scripts; The path to the fasta reference sequences -our ($poolData, $oneReportPerSample) = (2, 2); # If a folder is pass as input file pool all the data and generate the report on the pool and for each samples; # Generate one report for each samples - - -GetOptions('verbose|v'=>\$verbose, 'help|h'=>\$help, 'man|m'=>\$man, 'refGenome=s'=>\$refGenome, 'outfile|o=s' => \$output, 'pathTemporary|temp=s' => \$folder_temp, 'pathRscript=s' => \$path_R_Scripts, 'pathSeqRefGenome=s' => \$path_SeqrefGenome, 'poolData' => \$poolData, 'reportSample' => \$oneReportPerSample) or pod2usage(2); - -our ($input) = @ARGV; - -pod2usage(-verbose=>1, -exitval=>1, -output=>\*STDERR) if ($help); -pod2usage(-verbose=>2, -exitval=>1, -output=>\*STDERR) if ($man); -pod2usage(-verbose=>0, -exitval=>1, -output=>\*STDERR) if(@ARGV == 0); # No argument is pass to the command line print the usage of the script -pod2usage(-verbose=>0, -exitval=>1, -output=>\*STDERR) if(@ARGV == 2); # Only one argument is expected to be pass to @ARGV (the input) - - - -###################################################################################################################################################### -# GLOBAL VARIABLES # -###################################################################################################################################################### -# Recover the current path -our $pwd = `pwd`; -chomp($pwd); - -# Path to R scripts -our $pathRScriptTxnSB = "$path_R_Scripts/R/transciptionalStrandBias.r"; -our $pathRScriptMutSpectrum = "$path_R_Scripts/R/mutationSpectra_Galaxy.r"; - -our $folderMutAnalysis = ""; -our @pathInput = split("/", $input); - -# Hash table with the length of each chromosomes -our %chromosomes; - -###################################################################################################################################################### -# MAIN # -###################################################################################################################################################### -# Check the presence of the flags and create the output and temp directories -CheckFlags(); - -# Retrieve chromosomes length -checkChrDir(); - - -print "-----------------------------------------------------------------\n"; -print "-----------------Report Mutational Analysis----------------------\n"; -print"-----------------------------------------------------------------\n"; - -# First check if the file is annotated or not -CheckAnnotationFile($input); - -# Calculate the statistics and generate the report -my @colInfoAV = qw(Chr Start Ref Alt); -ReportMutDist($input, $folderMutAnalysis, $folder_temp, \@colInfoAV, $refGenome); - -# Remove the temporary directory -rmtree($folder_temp); - - -###################################################################################################################################################### -# FUNCTIONS # -###################################################################################################################################################### - -# Check the presence of the flags and create the output and temp directories -sub CheckFlags -{ - # Check the reference genome - if($refGenome eq "empty") { print STDERR "You forget to specify the name for the reference genome!!!\nPlease specify it with the flag --refGenome\n"; exit; } - - # If no output is specified write the result as the same place as the input file - if($output eq "empty") - { - my $folderRes = ""; - for(my $i=0; $i<$#pathInput; $i++) { $folderRes .= "$pathInput[$i]/"; } - - $folderMutAnalysis = "$folderRes/Mutational_Analysis"; - if(!-e $folderMutAnalysis) { mkdir($folderMutAnalysis) or die "$!: $folderMutAnalysis\n"; } - } - else - { - if(!-e $output) { mkdir($output) or die "$!: $output\n"; } - - $folderMutAnalysis = "$output/Mutational_Analysis"; - if(!-e $folderMutAnalysis) { mkdir($folderMutAnalysis) or die "$!: $folderMutAnalysis\n"; } - } - - # If no temp folder is specified write the result in the current path - if($folder_temp eq "empty") { $folder_temp = "$pwd/TEMP_MutationalAnalysis_$pathInput[$#pathInput]"; } - if(!-e $folder_temp) { mkdir($folder_temp) or die "$!: $folder_temp\n"; } - - # Check the path to the R scripts - if($path_R_Scripts eq "empty") { print STDERR "You forget to specify the path for the R scripts!!!\nPlease specify it with the flag --pathRscript\n"; exit; } - - - # The input is a folder - if(-d $input) { foreach my $file (`ls $input`) { CheckLengthFilename("$input/$file"); } } - # The input is one file - else { CheckLengthFilename($input); } -} -# Check the length of the file, must be < 32 characters for the Excel sheet -sub CheckLengthFilename -{ - my ($inputFile) = @_; - - ## Verify the name of file, must be <= 31 chars for the sheet name - my ($filename, $directories, $suffix) = fileparse($inputFile, qr/\.[^.]*/); - - if(length($filename) > 31) { print STDERR "The file: $inputFile must be <= 31 chars\nPlease modify it before running the script\n"; exit; } -} - -# Retrieve chromosomes length -sub checkChrDir -{ - my @files = `ls $path_SeqrefGenome/$refGenome"_seq"/*.fa`; - foreach my $file (@files) - { - if ($file !~ /chr(\d+|x|y)\.fa/i){next;} - open(FILE,$file); - <FILE>; - my $seq=""; - while (<FILE>){ chomp; $seq.=$_;} - $file =~ /chr(.*)\.fa/; - $chromosomes{"chr".$1}=length($seq); - } -} - -# Check if the file is annotated or not -sub CheckAnnotationFile -{ - my ($inputFile) = @_; - - # A folder is pass in argument - if(-d $inputFile) - { - foreach my $file (`ls $inputFile`) - { - chomp($file); - - open(F1, "$inputFile/$file") or die "$!: $inputFile/$file\n"; - my $search_header = <F1>; $search_header =~ s/[\r\n]+$//; my @tab_search_header = split("\t",$search_header); - close F1; - # The number of the column - my $value_of_column_NB = "toto"; - for(my $i=0; $i<=$#tab_search_header; $i++) - { - if($tab_search_header[$i] eq "Func.refGene") { $value_of_column_NB = $i; } - } - if($value_of_column_NB eq "toto") { print STDERR "Error the input file you specify is not annotated! $inputFile/$file !!!!\nPlease first annotate your file before trying to generate the report on mutation patterns\n"; exit; } - } - } - else - { - open(F1, $inputFile) or die "$!: $inputFile\n"; - my $search_header = <F1>; $search_header =~ s/[\r\n]+$//; my @tab_search_header = split("\t",$search_header); - close F1; - # The number of the column - my $value_of_column_NB = "toto"; - for(my $i=0; $i<=$#tab_search_header; $i++) - { - if($tab_search_header[$i] eq "Func.refGene") { $value_of_column_NB = $i; } - } - if($value_of_column_NB eq "toto") { print STDERR "Error the input file you specify is not annotated! $inputFile !!!!\nPlease first annotate your file before trying to generate the report on mutation patterns\n"; exit; } - } -} - -# Calculate the statistics and generate the report -sub ReportMutDist -{ - our ($input, $output, $folder_temp, $refTab_colInfo, $refGenome) = @_; - - my @column = @$refTab_colInfo; - - our ($chr_name, $start_name, $ref_name, $alt_name) = split(/,/, join(',', @column)); # Separe each element - - our $func_name = "Func.refGene"; - our $exonicFunc_name = "ExonicFunc.refGene"; - our $strand_name = "Strand"; - our $context_name = "context"; - - my $folderFigure = "$output/Figures"; - if(-e $folderFigure) { rmtree($folderFigure); mkdir($folderFigure) or die "Can't create the directory $folderFigure\n"; } - else { mkdir($folderFigure) or die "Can't create the directory $folderFigure\n"; } - my $folderChi2 = "$folderFigure/Chi2"; - if(!-e $folderChi2) { mkdir($folderChi2) or die "Can't create the directory $folderChi2\n"; } - my $folderWebLogo = "$folderFigure/WebLogo"; - if(!-e $folderWebLogo) { mkdir($folderWebLogo) or die "Can't create the directory $folderWebLogo\n"; } - my $folderNMF = "$folderFigure/Input_NMF"; - if(!-e $folderNMF) { mkdir($folderNMF) or die "Can't create the directory $folderNMF\n"; } - - ################################################################################################ - ### Calculates all the statistics ### - ################################################################################################ - ############ Recover Annovar annotations (for having the save number of functional regions for each samples) - my @tab_func = recoverAnnovarAnnotation($input, $func_name); - if(@tab_func == 0) { print STDERR "Error the table for the functional region is empty!!!!! check $input $func_name\n"; exit; } - - ############ Calculate the different statistics present in the report - my %h_file = (); - CalculateStatistics(\%h_file, \@tab_func); - - ############ Calculate the chi2 for the strand bias - CalculateChi2(\%h_file, $folderChi2); - - ############ Write the different statistics present in the report - WriteStatistics(\%h_file, $#tab_func, $folderFigure, $folderChi2, $folderNMF); - - ############ Create logo for studying the 10 flanking bases of the mutation - CreateLogo(\%h_file, $folderWebLogo); - - - ################### Subroutines for generating the report for the mutational analysis - sub recoverAnnovarAnnotation - { - my ($input, $AV_annotation) = @_; - - my %hash = (); - - # The input is a folder - if(-d $input) - { - foreach my $file (`ls $input`) - { - $file =~ s/[\r\n]+$//; - my $AV_annotation_value = recoverNumCol("$input/$file", $AV_annotation); - - open(F1, "$input/$file") or die "$!: $input/$file\n"; - my $header = <F1>; - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - # Some files can have an empty line at the end and WE DON'T WANT to consider it - if(! defined $tab[0]) { next; } - # Some func value are repeated and separated by ";" - my $funcSegment = ""; - if($tab[$AV_annotation_value] =~ /;/) { my @temp = split(";", $tab[$AV_annotation_value]); $funcSegment = $temp[0]; } - else { $funcSegment = $tab[$AV_annotation_value]; } - - $hash{$funcSegment} = ""; - } - close F1; - } - my @tab_AVAnnotation = (); - foreach my $k (sort keys %hash) { push(@tab_AVAnnotation, $k); } - return @tab_AVAnnotation; - } - # The input is a file - else - { - my $AV_annotation_value = recoverNumCol($input, $AV_annotation); - - open(F1, $input) or die "$!: $input\n"; - my $header = <F1>; - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - # Some func value are repeated and separated by ";" - my $funcSegment = ""; - if($tab[$AV_annotation_value] =~ /;/) { my @temp = split(";", $tab[$AV_annotation_value]); $funcSegment = $temp[0]; } - else { $funcSegment = $tab[$AV_annotation_value]; } - - $hash{$funcSegment} = ""; - } - close F1; - my @tab_AVAnnotation = (); - foreach my$k (sort keys %hash) { push(@tab_AVAnnotation, $k); } - return @tab_AVAnnotation; - } - } - # Calculate the different statistics present in the report - sub CalculateStatistics - { - my ($refH_file, $refT_func) = @_; - - my ($chr_value, $start_value, $ref_value, $alt_value, $func_value, $exonicFunc_value, $strand_value, $contextSeq_value) = ("", "", "", "", "", "", "", "", "", ""); - - # If the input is a folder - if(-d $input) - { - my $folderPool = "$folder_temp/Pool"; - if(!-e $folderPool) { mkdir($folderPool) or die "Can't create the directory $folderPool\n"; } - - # Copy each sample - foreach my $file (`ls $input`) { chomp($file); system("cp $input/$file $folderPool"); } - - # Generate the pool of all the data - if($poolData == 1) - { - my @listFile = `ls $input`; - - # For keeping the header only one time - chomp($listFile[0]); - system("cp $input/$listFile[0] $folderPool/Pool_Data.txt"); - - open(OUT, ">>", "$folderPool/Pool_Data.txt") or die "$!: $folderPool/Pool_Data.txt\n"; - - for(my $i=1; $i<=$#listFile; $i++) - { - chomp($listFile[$i]); - open(F1, "$input/$listFile[$i]") or die "$!: $input/$listFile[$i]\n"; - my $header = <F1>; - while(<F1>) { print OUT $_; } - close F1; - } - close OUT; - } - - foreach my $file (`ls $folderPool`) - { - chomp($file); - ############ Recover the number of the columns of interest - $chr_value = recoverNumCol("$folderPool/$file", $chr_name); - $start_value = recoverNumCol("$folderPool/$file", $start_name); - $ref_value = recoverNumCol("$folderPool/$file", $ref_name); - $alt_value = recoverNumCol("$folderPool/$file", $alt_name); - $func_value = recoverNumCol("$folderPool/$file", $func_name); - $exonicFunc_value = recoverNumCol("$folderPool/$file", $exonicFunc_name); - $strand_value = recoverNumCol("$folderPool/$file", $strand_name); - $contextSeq_value = recoverNumCol("$folderPool/$file", $context_name); - ############ Recover the number of the columns of interest - - ############ Control the annotated file pass in argument - ## Check if the files have variants - my $nbLines_originalFile = `wc -l $folderPool/$file`; $nbLines_originalFile =~ /(\d+) /; - if($1==1) { print STDERR "\n\nNo line in the file $folderPool/$file\n\n"; exit; } - ## Check if there is variant with strand information. If not the rest of the script generates errors - my $testFile = 0; - CheckVariantReport("$folderPool/$file", $strand_value, \$testFile); - if($testFile==0) { print STDERR "\n\nNo strand information for the file $folderPool/$file\n\n"; exit; } - ############ Control the annotated file pass in argument - - ############ Calculate the statistics - File2Hash("$folderPool/$file", $func_value, $exonicFunc_value, $chr_value, $ref_value, $alt_value, $strand_value, $contextSeq_value, $refH_file, $refT_func); - } - } - # If the input is a file - else - { - ############ Recover the number of the columns of interest - $chr_value = recoverNumCol($input, $chr_name); - $start_value = recoverNumCol($input, $start_name); - $ref_value = recoverNumCol($input, $ref_name); - $alt_value = recoverNumCol($input, $alt_name); - $func_value = recoverNumCol($input, $func_name); - $exonicFunc_value = recoverNumCol($input, $exonicFunc_name); - $strand_value = recoverNumCol($input, $strand_name); - $contextSeq_value = recoverNumCol($input, $context_name); - ############ Recover the number of the columns of interest - - ############ Control the annotated file pass in argument - ## Check if the files have variants - my $nbLines_originalFile = `wc -l $input`; $nbLines_originalFile =~ /(\d+) /; - if($1==1) { print STDERR "\n\nNo line in the file $input\n\n"; exit; } - ## Check if there is variant with strand information. If not the rest of the script generates errors - my $testFile = 0; - CheckVariantReport($input, $strand_value, \$testFile); - if($testFile==0) { print STDERR "\n\nNo strand information for the file $input\n\n"; exit; } - ############ Control the annotated file pass in argument - - ############ Calculate the statistics - File2Hash($input, $func_value, $exonicFunc_value, $chr_value, $ref_value, $alt_value, $strand_value, $contextSeq_value, $refH_file, $refT_func); - } - } - # Check if there is at least one variant with a strand information - sub CheckVariantReport - { - my ($file, $strand_value, $refS_testFile) = @_; - - open(F1, $file) or die "$!: $file\n"; - my $header = <F1>; - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - if( ($tab[$strand_value] eq "+") || ($tab[$strand_value] eq "-") ) { $$refS_testFile++; } - } - close F1; - } - # Convert the annotated VCF into a hash table - sub File2Hash - { - my ($inputFile, $func_value, $exonicFunc_value, $chr_value, $ref_value, $alt_value, $strand_value, $contextSeq_value, $refH_file, $refT_func) = @_; - my ($filename, $directories, $suffix) = fileparse($inputFile, qr/\.[^.]*/); - - # Initialisation of the hash - my @tab_mutation = qw(C:G>A:T C:G>G:C C:G>T:A T:A>A:T T:A>C:G T:A>G:C); - my @tab_aaChange = ("NonTr", "Tr", "TotalMutG"); - my @tabExoFunc = ("frameshift insertion", "frameshift deletion", "frameshift block substitution", "frameshift substitution", "stopgain", "stoploss", "nonframeshift insertion", "nonframeshift deletion", "nonframeshift substitution", "nonframeshift block substitution", "nonsynonymous SNV", "synonymous SNV", "unknown", "NA"); - - # Total number of SBS on the genomic strand - $refH_file->{$filename}{'TotalSBSGenomic'} = 0; - # Total number of Indel on the genomic strand - $refH_file->{$filename}{'TotalIndelGenomic'} = 0; - # Total number of SBS on the coding strand - $refH_file->{$filename}{'TotalSBSCoding'} = 0; - # Total number of SBS and Indel on the genomic strand - $refH_file->{$filename}{'TotalMutGenomic'} = 0; - - ##################################### - # SBS by segment (6 mutation types) # - ##################################### - foreach my $elt_tabFunc (@$refT_func) - { - foreach my $elt_tabMutation (@tab_mutation) - { - foreach my $elt_aaChange (@tab_aaChange) - { - $refH_file->{$filename}{'6mutType'}{$elt_tabFunc}{$elt_tabMutation}{$elt_aaChange} = 0; - } - } - } - - ####################### - # Pearson correlation # - ####################### - $refH_file->{$filename}{'SBSPerChr'}{'AllMutType'} = 0; - # Count of SBS per chromosome foreach mutation types - foreach my $elt_tabMutation (@tab_mutation) - { - foreach my $chromosome (sort keys %chromosomes){ $refH_file->{$filename}{'SBSPerChr'}{$elt_tabMutation}{'CHR'}{$chromosome}{'chr'} = 0;} - $refH_file->{$filename}{'SBSPerChr'}{$elt_tabMutation}{'Pearson'} = 0; - } - - foreach my $chromosome (sort keys %chromosomes){ - $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}=0; - } - - ############################ - # Impact of SBS on protein # - ############################ - foreach my $elt_exoFunc (@tabExoFunc) - { - $refH_file->{$filename}{'ImpactSBS'}{$elt_exoFunc} = 0; - } - - ##################################### - # Sequence context (genomic strand) # - ##################################### - my @tab_mutation2 = qw(C>A C>G C>T T>A T>C T>G); - my @tab_context = qw(A_A A_C A_G A_T C_A C_C C_G C_T G_A G_C G_G G_T T_A T_C T_G T_T); - foreach my $elt_context (@tab_context) - { - foreach my $elt_mutation3 (@tab_mutation2) - { - $refH_file->{$filename}{'SeqContextG'}{$elt_context}{$elt_mutation3} = 0; - } - } - - #################################### - # Sequence context (coding strand) # - #################################### - my @tab_TrNonTr = qw(NonTr Tr); - foreach my $elt_context (@tab_context) - { - foreach my $elt_mutation2 (@tab_mutation2) - { - foreach my $trNonTr (@tab_TrNonTr) - { - $refH_file->{$filename}{'SeqContextC'}{$elt_context}{$elt_mutation2}{$trNonTr} = 0; - } - } - } - - open(F1,$inputFile) or die "$!: $inputFile\n"; - my $header = <F1>; - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - # Random chromosome and chromosome M - if( ($tab[$chr_value] =~ /random/i) || ($tab[$chr_value] =~ /M/i) ) { next; } - - ############################################## Extract the base just before and after the mutation ############################################## - my $context = ""; - my $contextSequence = $tab[$contextSeq_value]; $contextSequence =~ tr/a-z/A-Z/; - my @tempContextSequence = split("", $contextSequence); - my $total_nbBaseContext = $#tempContextSequence; - my $midlle_totalNbBaseContext = $total_nbBaseContext/2; # For having the middle of the sequence - my $before = $midlle_totalNbBaseContext - 1; my $after = $midlle_totalNbBaseContext + 1; - $context = $tempContextSequence[$before]."_".$tempContextSequence[$after]; - ############################################## Extract the base just before and after the mutation ############################################## - - - ############################################################### Impact on protein ############################################################### - my $exoFunc = ""; - # Sometimes the annotation is repeated frameshift deletion;frameshift deletion - if($tab[$exonicFunc_value] =~ /\;/) - { - my @temp = split(";", $tab[$exonicFunc_value]); - if($temp[0] eq $temp[1]) { $exoFunc = $temp[0]; } - } - # The annotations have changed after MAJ Annovar 2014Jul22 (stopgain SNV => stopgain) - elsif($tab[$exonicFunc_value] eq "stopgain SNV") { $exoFunc = "stopgain"; } - elsif($tab[$exonicFunc_value] eq "stoploss SNV") { $exoFunc = "stoploss"; } - elsif($tab[$exonicFunc_value] eq "nonsynonymous_SNV") { $exoFunc = "nonsynonymous SNV"; } - elsif($tab[$exonicFunc_value] eq "stopgain_SNV") { $exoFunc = "stopgain SNV"; } - elsif($tab[$exonicFunc_value] eq "synonymous_SNV") { $exoFunc = "synonymous SNV"; } - else { $exoFunc = $tab[$exonicFunc_value]; } - - if(exists $refH_file->{$filename}{'ImpactSBS'}{$exoFunc}) - { - # If the sequence context if not recovered correctly don't considered the variants - if( ($context =~ /N/) || (length($context) != 3) ) { next; } - - $refH_file->{$filename}{'ImpactSBS'}{$exoFunc}++; - $refH_file->{$filename}{'TotalMutGenomic'}++; - } - else { print "WARNING: Exonic function not considered: $exoFunc\n"; } - ############################################################### Impact on protein ############################################################### - - ################################################### Only SBS are considered for the statistics ################################################## - if( ($tab[$ref_value] =~ /^[ACGT]$/i) && ($tab[$alt_value] =~ /^[ACGT]$/i) ) - { - # If the sequence context if not recovered correctly don't considered the variants - if( ($context =~ /N/) || (length($context) != 3) ) { next; } - - # Total number of SBS on the genomic strand - $refH_file->{$filename}{'TotalSBSGenomic'}++; - - # Total number of SBS on the coding strand with a sequence context - if( ($tab[$strand_value] eq "+") || ($tab[$strand_value] eq "-") ) - { - if( ($context ne "NA") && (($context =~ /N/) || (length($context) != 3)) ) { next; } - $refH_file->{$filename}{'TotalSBSCoding'}++; - } - } - else { $refH_file->{$filename}{'TotalIndelGenomic'}++; } - ################################################### Only SBS are considered for the statistics ################################################## - - # Number of SBS per chromosome: remove the "chr" - my $chrNameForH=$tab[$chr_value]; - if(exists $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chrNameForH}{'chr'}++; } - - - ################################################### Some func value are repeated and separated by ";" ################################################## - my $funcSegment = ""; - if($tab[$func_value] =~ /;/) { my @temp = split(";", $tab[$func_value]); $funcSegment = $temp[0]; } - else { $funcSegment = $tab[$func_value]; } - - - ############################################################### MUTATION C> ############################################# - ###################################### C:G>A:T - if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "A")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "T") ) ) - { - my $mutation = "C:G>A:T"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "C>A"; - # We want to express the mutation in C> - if( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "T") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis C>A on NonTr strand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "A"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "T"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # C>A With the sequence context (C>A strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "A")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>A'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>A'}{'NonTr'}++; } - } - # C>A With the sequence context (G>T strand = -) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>A'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>A'}{'NonTr'}++; } - } - } - # Strand analysis C>A on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "A"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "T"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # C>A With the sequence context (C>A strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "A")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>A'}{'Tr'}) { { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>A'}{'Tr'}++; } } - } - # C>A with the sequence context (G>T strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "T")) ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>A'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>A'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "A")) - { - # For the logo all the sequences must have the same length - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."C".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CA'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."C".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CA'}}, $context); - } - } - ###################################### C:G>G:C - if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "G")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "C") ) ) - { - my $mutation = "C:G>G:C"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "C>G"; - # We want to express the mutation in C> - if( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "C") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis C>G on NonTr strand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "G"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "C"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # C>G with the sequence context (C>G strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "G")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>G'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>G'}{'NonTr'}++; } - } - # C>G with the sequence context (G>C strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "C")) ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>G'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>G'}{'NonTr'}++; } - } - } - # Strand analysis C>G on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "G"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "C"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # C>G with the sequence context (C>G strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "G")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>G'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>G'}{'Tr'}++; } - } - # C>G with the sequence context (G>C strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "C")) ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>G'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>G'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "G")) - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."C".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CG'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."C".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CG'}}, $context); - } - } - ###################################### C:G>T:A - if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "T")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "A") ) ) - { - my $mutation = "C:G>T:A"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "C>T"; - # We want to express the mutation in C> - if( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "A") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis C>T on NonTr strand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "T"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "A"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # C>T with the sequence context (C>T strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "T")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>T'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>T'}{'NonTr'}++; } - } - # C>T with the sequence context (G>A strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "A")) ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>T'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>T'}{'NonTr'}++; } - } - } - # Strand analysis C>T on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "T"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "A"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # C>T with the sequence context (C>T strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "C")&&($tab[$alt_value] eq "T")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'C>T'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'C>T'}{'Tr'}++; } - } - # C>T with the sequence context (G>A strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "G")&&($tab[$alt_value] eq "A")) ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>T'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'C>T'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "T")) - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."C".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CT'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."C".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'CT'}}, $context); - } - } - - ############################################################### MUTATION T> ############################################# - ###################################### T:A>A:T - if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "A")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "T") ) ) - { - my $mutation = "T:A>A:T"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "T>A"; - # We want to express the mutation in T> - if( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "T") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis T>A on NonTr stand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "A"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "T"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # T>A with the sequence context (T>A strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "A")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>A'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>A'}{'NonTr'}++; } - } - # T>A with the sequence context (A>T strand = -) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>A'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>A'}{'NonTr'}++; } - } - } - # Strand analysis T>A on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "A"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "T"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # T>A <ith the sequence context (T>A strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "A")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>A'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>A'}{'Tr'}++; } - } - # T>A with the sequence context (A>T strand = +) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>A'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>A'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "A")) - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."T".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TA'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."T".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TA'}}, $context); - } - } - ###################################### T:A>C:G - if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "C")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "G")) ) - { - my $mutation = "T:A>C:G"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "T>C"; - # We want to express the mutation in T> - if( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "T") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis T>C on NonTr strand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "C"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "G"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # T>C (T>C strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "C")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>C'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>C'}{'NonTr'}++; } - } - # T>C (A>G strand = -) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>C'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>C'}{'NonTr'}++; } - } - } - # Strand analysis T>C on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "C"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "G"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # T>C (T>C strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "C")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>C'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>C'}{'Tr'}++; } - } - # T>C (A>G strand = +) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>C'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>C'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "C")) - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."T".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TC'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."T".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TC'}}, $context); - } - } - ###################################### T:A>G:C - if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "G")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "C")) ) - { - my $mutation = "T:A>G:C"; - $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; # Count the total number of mutations - - # Pearson correlation - if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) { $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; } - - # Sequence context - 6 mutation types - genomic strand - my $mutationSeqContext6mutType = "T>G"; - # We want to express the mutation in T> - if( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "T") ) - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; } - } - elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) { $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; } - - # Strand analysis T>G on NonTr strand - if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "G"))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "C"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; } - - # T>G (T>G strand = +) - if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "G")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>G'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>G'}{'NonTr'}++; } - } - # T>G (A>C strand = -) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>G'}{'NonTr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>G'}{'NonTr'}++; } - } - } - # Strand analysis T>G on Tr strand - if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "G"))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq "A")&&($tab[$alt_value] eq "C"))) ) - { - if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) { $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; } - - # T>G (T>G strand = -) - if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq "T")&&($tab[$alt_value] eq "G")) ) - { - if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{'T>G'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context}{'T>G'}{'Tr'}++; } - } - # T>G (A>C strand = +) - else - { - my $base3 = complement($tempContextSequence[$before]); my $base5 = complement($tempContextSequence[$after]); - my $context_reverse = $base5."_".$base3; - if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>G'}{'Tr'}) { $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{'T>G'}{'Tr'}++; } - } - } - # WebLogo-3 - if(($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "G")) - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } - my $context = $contextTemp1."T".$contextTemp2; $context = reverse $context; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TG'}}, $context); - } - else - { - if(scalar(@tempContextSequence) == 2) { next; } - my ($contextTemp1, $contextTemp2) = ("", ""); - for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } - for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } - my $context = $contextTemp1."T".$contextTemp2; - push(@{$refH_file->{$filename}{'WebLogo3'}{'TG'}}, $context); - } - } - } - close F1; - } - # Write the different statistics in the report - sub WriteStatistics - { - my ($refH_file, $nb_func, $folderFigure, $folderChi2, $folderNMF) = @_; - - # Save the different graphs in specific folder instead of in a general one. - if(!-e "$folderFigure/Overall_mutation_distribution") { mkdir("$folderFigure/Overall_mutation_distribution") or die "Can't create the directory $folderFigure/Overall_mutation_distribution\n"; } - if(!-e "$folderFigure/Impact_protein_sequence") { mkdir("$folderFigure/Impact_protein_sequence") or die "Can't create the directory $folderFigure/Impact_protein_sequence\n"; } - if(!-e "$folderFigure/SBS_distribution") { mkdir("$folderFigure/SBS_distribution") or die "Can't create the directory $folderFigure/SBS_distribution\n"; } - if(!-e "$folderFigure/Stranded_Analysis") { mkdir("$folderFigure/Stranded_Analysis") or die "Can't create the directory $folderFigure/Stranded_Analysis\n"; } - if(!-e "$folderFigure/Trinucleotide_Sequence_Context") { mkdir("$folderFigure/Trinucleotide_Sequence_Context") or die "Can't create the directory $folderFigure/Trinucleotide_Sequence_Context\n"; } - if(!-e "$folderFigure/Distribution_SBS_Per_Chromosomes") { mkdir("$folderFigure/Distribution_SBS_Per_Chromosomes") or die "Can't create the directory $folderFigure/Distribution_SBS_Per_Chromosomes\n"; } - - - # Create a workbook with all the samples - my $wb = ""; my $ws_sum = ""; my %h_chi2 = (); - my ($ws_inputNMF_count, $ws_inputNMF_percent) = ("", ""); - ############### Define the format - my ($format_A10, $format_A10Boldleft, $format_A10ItalicRed) = ("", "", ""); - my ($formatT_left, $formatT_right, $formatT_bottomRight, $formatT_bottomLeft, $formatT_bottom, $formatT_bottomHeader, $formatT_bottomRightHeader, $formatT_bottomHeader2, $formatT_rightHeader); - my ($formatT_graphTitle); - my ($table_topleft, $table_topRight, $table_bottomleft, $table_bottomRight, $table_top, $table_right, $table_bottom, $table_bottomItalicRed, $table_left, $table_bottomrightHeader, $table_left2, $table_middleHeader, $table_middleHeader2); - - if($oneReportPerSample == 2) - { - $wb = Spreadsheet::WriteExcel->new("$output/Report_Mutation_Spectra.xls"); - - ############### Define the format - Format_A10($wb, \$format_A10); # Text center in Arial 10 - Format_A10BoldLeft($wb, \$format_A10Boldleft); # Text on the left in Arial 10 bold - Format_TextSection($wb, \$formatT_left, \$formatT_right, \$formatT_bottomRight, \$formatT_bottomLeft, \$formatT_bottom, \$formatT_bottomHeader, \$formatT_bottomRightHeader, \$formatT_bottomHeader2, \$formatT_rightHeader); - Format_GraphTitle($wb, \$formatT_graphTitle); - Format_Table($wb, \$table_topleft, \$table_topRight, \$table_bottomleft, \$table_bottomRight, \$table_top, \$table_right, \$table_bottom, \$table_bottomItalicRed, \$table_left, \$table_bottomrightHeader, \$table_left2, \$table_middleHeader, \$table_middleHeader2); - Format_A10ItalicRed($wb, \$format_A10ItalicRed); - - - ############### Worksheet with a summary of the samples - $ws_sum = $wb->add_worksheet("Sample_List"); - $ws_sum->write(0, 0, "Samples", $format_A10); $ws_sum->write(0, 1, "Total number SBS", $format_A10); $ws_sum->write(0, 2, "Total number of Indel", $format_A10); $ws_sum->write(0, 3, "Total number of mutations", $format_A10); - $ws_sum->set_column(0,0, 50); $ws_sum->set_column(1,1, 20); $ws_sum->set_column(2,2, 20); $ws_sum->set_column(3,3, 22); - - ############### Save the chi2 values into a hash table - if(-e "$folderChi2/Output_chi2_strandBias.txt") - { - open(F1, "$folderChi2/Output_chi2_strandBias.txt") or die "$!: $folderChi2/Output_chi2_strandBias.txt\n"; - my $header = <F1>; # Strand_Bias($tab[0]) NonTr-Tr($tab[1]) Proportion($tab[2]) P-val-Chi2($tab[3]) FDR($tab[4]) Confidence Interval($tab[5]) Mutation_Type($tab[6]) SampleName($tab[7]) - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - $h_chi2{$tab[7]}{$tab[6]}{'p-value'} = $tab[3]; $h_chi2{$tab[7]}{$tab[6]}{'ConfInt'} = $tab[5]; - - # For the pool data the FDR isn't calculated so replace the NA (=Missing values) by "-" - if($tab[7] eq "Pool_Data") { $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = "-"; } - else { $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = $tab[4]; } - } - close F1; - } - ############### Write the input matrix for NMF for the count and the un-normalized frequency - $ws_inputNMF_count = $wb->add_worksheet("Input_NMF_Count"); - $ws_inputNMF_percent = $wb->add_worksheet("Input_NMF_Percent"); - } - - - ################################################ Set the Rows and columns of the different part of the report ################################################ - my $row_SumSheet = 1; # First row for the summary sheet of the report - my $rowStart_SBSdistrBySeg = 48; my $colStart_SBSdistrBySeg = 0; # For the table SBS distribution by segment - my $colStart_matrixSeqContext = 19; # Sequence context - my $col_inputNMF = 0; # Write the names of the samples with at least 33 SBS - - # For NMF input - my %h_inputNMF = (); - - ## For each file - foreach my $k_file (sort keys $refH_file) - { - print "File in process: $k_file\n"; - if($k_file ne "Pool_Data") { $col_inputNMF++; } - - # Create one workbook for each sample - if($oneReportPerSample == 1) - { - $wb = Spreadsheet::WriteExcel->new("$output/Report_Mutation_Spectra-$k_file.xls"); - - ############### Define the format - Format_A10($wb, \$format_A10); # Text center in Arial 10 - Format_A10BoldLeft($wb, \$format_A10Boldleft); # Text on the left in Arial 10 bold - Format_TextSection($wb, \$formatT_left, \$formatT_right, \$formatT_bottomRight, \$formatT_bottomLeft, \$formatT_bottom, \$formatT_bottomHeader, \$formatT_bottomRightHeader, \$formatT_bottomHeader2, \$formatT_rightHeader); - Format_GraphTitle($wb, \$formatT_graphTitle); - Format_Table($wb, \$table_topleft, \$table_topRight, \$table_bottomleft, \$table_bottomRight, \$table_top, \$table_right, \$table_bottom, \$table_bottomItalicRed, \$table_left, \$table_bottomrightHeader, \$table_left2, \$table_middleHeader, \$table_middleHeader2); - Format_A10ItalicRed($wb, \$format_A10ItalicRed); - - - ############### Worksheet with a summary of the samples - $ws_sum = $wb->add_worksheet("Sample_List"); - $ws_sum->write(0, 0, "Samples", $format_A10); $ws_sum->write(0, 1, "Total number SBS", $format_A10); $ws_sum->write(0, 2, "Total number of Indel", $format_A10); $ws_sum->write(0, 3, "Total number of mutations", $format_A10); - $ws_sum->set_column(0,0, 50); $ws_sum->set_column(1,1, 20); $ws_sum->set_column(2,2, 20); $ws_sum->set_column(3,3, 22); - # Write in the Samples sheet the name and the total number of SBS - $ws_sum->write(1, 0, "$k_file", $format_A10); - $ws_sum->write(1, 1, $refH_file->{$k_file}{'TotalSBSGenomic'}, $format_A10); $ws_sum->write(1, 2, $refH_file->{$k_file}{'TotalIndelGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 3, $refH_file->{$k_file}{'TotalMutGenomic'}, $format_A10); - - ############### Save the chi2 values into a hash table - if(-e "$folderChi2/Output_chi2_strandBias.txt") - { - open(F1, "$folderChi2/Output_chi2_strandBias.txt") or die "$!: $folderChi2/Output_chi2_strandBias.txt\n"; - my $header = <F1>; # Strand_Bias($tab[0]) NonTr-Tr($tab[1]) Proportion($tab[2]) P-val-Chi2($tab[3]) FDR($tab[4]) Confidence Interval($tab[5]) Mutation_Type($tab[6]) SampleName($tab[7]) - while(<F1>) - { - $_ =~ s/[\r\n]+$//; - my @tab = split("\t", $_); - - if($tab[7] eq $k_file) - { - $h_chi2{$tab[7]}{$tab[6]}{'p-value'} = $tab[3]; $h_chi2{$tab[7]}{$tab[6]}{'ConfInt'} = $tab[5]; - - # For the pool data the FDR isn't calculated so replace the NA (=Missing values) by "-" - if($tab[7] eq "Pool_Data") { $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = "-"; } - else { $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = $tab[4]; } - } - } - close F1; - } - - ############### Write the input matrix for NMF - if($k_file ne "Pool_Data") - { - # For NMF don't consider the pool of the samples - $ws_inputNMF_count = $wb->add_worksheet("Input_NMF_Count"); - $ws_inputNMF_percent = $wb->add_worksheet("Input_NMF_Percent"); - # Write in the input NMF sheet the name of the samples - $ws_inputNMF_count->write(0, 1, $k_file); - $ws_inputNMF_percent->write(0, 1, $k_file); - } - } - # One workbook with all the samples - else - { - # Write in the Samples sheet the name and the total number of SBS - $ws_sum->write($row_SumSheet, 0, $k_file, $format_A10); - $ws_sum->write($row_SumSheet, 1, $refH_file->{$k_file}{'TotalSBSGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 2, $refH_file->{$k_file}{'TotalIndelGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 3, $refH_file->{$k_file}{'TotalMutGenomic'}, $format_A10); - - # For NMF don't consider the pool of the samples - if($k_file ne "Pool_Data") - { - # Write in the input NMF sheet the name of the samples - $ws_inputNMF_count->write(0, $col_inputNMF, $k_file); - $ws_inputNMF_percent->write(0, $col_inputNMF, $k_file); - } - } - - # Count of SBS per chromosome - PearsonCoefficient($refH_file, $k_file); - - # Add a worksheet to the workbook - my $ws = $wb->add_worksheet($k_file); - - # Write the titles of the different sections of the report - WriteBoderSection($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext); - - # Write the mutation types (6 types) - WriteHeaderSection($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext); - - - # Save the figures of each samples in a different folder - if(!-e "$folderFigure/Overall_mutation_distribution/$k_file") { mkdir("$folderFigure/Overall_mutation_distribution/$k_file") or die "Can't create the directory $folderFigure/Overall_mutation_distribution/$k_file\n"; } - if(!-e "$folderFigure/Impact_protein_sequence/$k_file") { mkdir("$folderFigure/Impact_protein_sequence/$k_file") or die "Can't create the directory $folderFigure/Impact_protein_sequence/$k_file\n"; } - if(!-e "$folderFigure/SBS_distribution/$k_file") { mkdir("$folderFigure/SBS_distribution/$k_file") or die "Can't create the directory $folderFigure/SBS_distribution\n"; } - if(!-e "$folderFigure/Stranded_Analysis/$k_file") { mkdir("$folderFigure/Stranded_Analysis/$k_file") or die "Can't create the directory $folderFigure/Stranded_Analysis/$k_file\n"; } - if(!-e "$folderFigure/Trinucleotide_Sequence_Context/$k_file") { mkdir("$folderFigure/Trinucleotide_Sequence_Context/$k_file") or die "Can't create the directory $folderFigure/Trinucleotide_Sequence_Context/$k_file\n"; } - - - - ########################################################################################################################################################### - ################################################################# Write the statistics ################################################################### - ########################################################################################################################################################### - my ($ca_genomique, $cg_genomique, $ct_genomique, $ta_genomique, $tc_genomique, $tg_genomique) = (0,0,0,0,0,0); - my ($ca_NonTr, $ca_Tr, $cg_NonTr, $cg_Tr, $ct_NonTr, $ct_Tr, $ta_NonTr, $ta_Tr, $tc_NonTr, $tc_Tr, $tg_NonTr, $tg_Tr) = (0,0,0,0,0,0, 0,0,0,0,0,0); - - my $row_SBSdistrBySeg = $rowStart_SBSdistrBySeg+4; - my $row_SBSDistrBySegAndFunc_CA = $rowStart_SBSdistrBySeg+$nb_func+12; - my $row_SBSDistrBySegAndFunc_CG = $rowStart_SBSdistrBySeg+($nb_func*2)+16; my $rowEndCG_SBSDistrBySegAndFunc_CG = $row_SBSDistrBySegAndFunc_CG+$nb_func; - my $row_SBSDistrBySegAndFunc_CT = $rowStart_SBSdistrBySeg+($nb_func*3)+20; - - ## 6 mutation types by segment - foreach my $k_func (sort keys $refH_file->{$k_file}{'6mutType'}) - { - my $totalSBS_bySegment = 0; - - # Write the functional region for the section SBS distribution by segment - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg, $k_func, $formatT_left); - - # Write the exonic func for the section strand bias by segment - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg, $k_func, $formatT_left); - - if($row_SBSDistrBySegAndFunc_CG == $rowEndCG_SBSDistrBySegAndFunc_CG) - { - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg, $k_func, $formatT_bottomLeft); - } - else - { - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg, $k_func, $formatT_left); - } - - foreach my $k_mutation (sort keys $refH_file->{$k_file}{'6mutType'}{$k_func}) - { - if($k_mutation eq "C:G>A:T") - { - # Write the ratio NonTr(CA)/Tr(GT) - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+1, $ratioSB, $format_A10); - - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $format_A10); - - # Calculate the total number of SBS per mut type (genomic strand) - $ca_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $ca_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $ca_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); - } - if($k_mutation eq "C:G>G:C") - { - # Write the ratio NonTr(CG)/Tr(GC) - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+5, $ratioSB, $format_A10); - - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+6, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $format_A10); - - # Calculate the total number of SBS per mut type (genomic strand) - $cg_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $cg_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $cg_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+5, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); - } - if($k_mutation eq "C:G>T:A") - { - # Write the ratio NonTr(CT)/Tr(GA) - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+9, $ratioSB, $format_A10); - - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+10, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+11, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_right); - - # Calculate the total number of SBS per mut type (genomic strand) - $ct_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $ct_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $ct_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); - } - if($k_mutation eq "T:A>A:T") - { - # Write the ratio NonTr(AT)/Tr(TA) - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - - - if($row_SBSDistrBySegAndFunc_CG == $rowEndCG_SBSDistrBySegAndFunc_CG) - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+1, $ratioSB, $formatT_bottom); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $formatT_bottom); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_bottom); - } - else - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+1, $ratioSB, $format_A10); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $format_A10); - } - - - # Calculate the total number of SBS per mut type (genomic strand) - $ta_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $ta_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $ta_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+9, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); - } - if($k_mutation eq "T:A>C:G") - { - # Write the ratio NonTr(AG)/Tr(TC) - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - - if($row_SBSDistrBySegAndFunc_CG == $rowEndCG_SBSDistrBySegAndFunc_CG) - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+5, $ratioSB, $formatT_bottom); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+6, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $formatT_bottom); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_bottom); - } - else - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+5, $ratioSB, $format_A10); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+6, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $format_A10); - } - - # Calculate the total number of SBS per mut type (genomic strand) - $tc_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $tc_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $tc_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+11, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); - } - if($k_mutation eq "T:A>G:C") - { - # Calculate the ratio for the strand bias - my $ratioSB = 0; - if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) { $ratioSB = 0; } - else { $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; } - $ratioSB = sprintf("%.2f", $ratioSB); - - if($row_SBSDistrBySegAndFunc_CG == $rowEndCG_SBSDistrBySegAndFunc_CG) - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+9, $ratioSB, $formatT_bottom); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+10, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $formatT_bottom); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+11, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_bottomRight); - } - else - { - # Write the ratio NonTr(AC)/Tr(TG) - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+9, $ratioSB, $format_A10); - # Write the count of SBS in the NonTr and Tr strand - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+10, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg+11, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_right); - } - - # Calculate the total number of SBS per mut type (genomic strand) - $tg_genomique += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - # Calculate the total number of SBS by NonTr / Tr strand - $tg_NonTr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; $tg_Tr += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - - # Write the count by exonic region - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+13, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $formatT_right); - } - - # Calculate the total number of SBS on the genomic strand for each mutation types by exonic region - $totalSBS_bySegment += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; - } # End $k_mutation - - $row_SBSDistrBySegAndFunc_CA++; $row_SBSDistrBySegAndFunc_CG++; $row_SBSDistrBySegAndFunc_CT++; - - # Write the percent by exonic region - my $percent_ca = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_ca = 0; } - else { $percent_ca = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_ca = sprintf("%.2f", $percent_ca); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+2, $percent_ca, $format_A10); - my $percent_cg = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_cg = 0; } - else { $percent_cg = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>G:C'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_cg = sprintf("%.2f", $percent_cg); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+4, $percent_cg, $format_A10); - my $percent_ct = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_ct = 0; } - else { $percent_ct = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>T:A'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_ct = sprintf("%.2f", $percent_ct); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+6, $percent_ct, $format_A10); - my $percent_ta = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_ta = 0; } - else { $percent_ta = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'T:A>A:T'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_ta = sprintf("%.2f", $percent_ta); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+8, $percent_ta, $format_A10); - my $percent_tc = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_tc = 0; } - else { $percent_tc = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'T:A>C:G'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_tc = sprintf("%.2f", $percent_tc); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+10, $percent_tc, $format_A10); - my $percent_tg = 0; - if($refH_file->{$k_file}{'6mutType'}{$k_func}{'C:G>A:T'}{'TotalMutG'} == 0) { $percent_tg = 0; } - else { $percent_tg = ($refH_file->{$k_file}{'6mutType'}{$k_func}{'T:A>G:C'}{'TotalMutG'} / $totalSBS_bySegment ) * 100; $percent_tg = sprintf("%.2f", $percent_tg); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+12, $percent_tg, $format_A10); - - # Write the count of SBS by segment - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+1, $totalSBS_bySegment, $format_A10); - - $row_SBSdistrBySeg++; - } # End $k_func - - # Write the total number of SBS on the genomic strand - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+1, $refH_file->{$k_file}{'TotalSBSGenomic'}, $formatT_bottomHeader); - - # Write the total and the percentage of SBS for each mutation types and save it to a text file - open(DISTRSBS, ">", "$folderFigure/SBS_distribution/$k_file/$k_file-SBS_distribution.txt") or die "$!: $folderFigure/SBS_distribution/$k_file/$k_file-SBS_distribution.txt\n"; - print DISTRSBS "Mutation_Type\tCount\tPercentage\tSample\n"; - my $percent_ca = 0; - if($ca_genomique == 0) { $percent_ca = 0; } - else { $percent_ca = ($ca_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ca = sprintf("%.2f", $percent_ca); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+2, $percent_ca, $formatT_bottom); print DISTRSBS "C:G>A:T\t$ca_genomique\t$percent_ca\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+3, $ca_genomique, $formatT_bottomHeader); - my $percent_cg = 0; - if($cg_genomique == 0) { $percent_cg = 0; } - else { $percent_cg = ($cg_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_cg = sprintf("%.2f", $percent_cg); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+4, $percent_cg, $formatT_bottom); print DISTRSBS "C:G>G:C\t$cg_genomique\t$percent_cg\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+5, $cg_genomique, $formatT_bottomHeader); - my $percent_ct = 0; - if($ct_genomique == 0) { $percent_ct = 0; } - else { $percent_ct = ($ct_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ct = sprintf("%.2f", $percent_ct); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+6, $percent_ct, $formatT_bottom); print DISTRSBS "C:G>T:A\t$ct_genomique\t$percent_ct\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+7, $ct_genomique, $formatT_bottomHeader); - my $percent_ta = 0; - if($ta_genomique == 0) { $percent_ta = 0; } - else { $percent_ta = ($ta_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ta = sprintf("%.2f", $percent_ta); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+8, $percent_ta, $formatT_bottom); print DISTRSBS "T:A>A:T\t$ta_genomique\t$percent_ta\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+9, $ta_genomique, $formatT_bottomHeader); - my $percent_tc = 0; - if($tc_genomique == 0) { $percent_tc = 0; } - else { $percent_tc = ($tc_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_tc = sprintf("%.2f", $percent_tc); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+10, $percent_tc, $formatT_bottom); print DISTRSBS "T:A>C:G\t$tc_genomique\t$percent_tc\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+11, $tc_genomique, $formatT_bottomHeader); - my $percent_tg = 0; - if($tg_genomique == 0) { $percent_tg = 0; } - else { $percent_tg = ($tg_genomique/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_tg = sprintf("%.2f", $percent_tg); } - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+12, $percent_tg, $formatT_bottom); print DISTRSBS "T:A>G:C\t$tg_genomique\t$percent_tg\t$k_file\n"; - $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+13, $tg_genomique, $formatT_bottomRightHeader); - close DISTRSBS; - - ########################################################################################################################################################### - ################################################################### Write Strand BIAS ##################################################################### - ########################################################################################################################################################### - # Write the SB for each mutation type (table 3) - $ws->write(28, 11, "Table 3. Significance of the strand biases", $format_A10Boldleft); - $ws->set_column(11, 11, 13); $ws->set_column(16, 16, 15); $ws->set_column(17, 17, 10); - $ws->write(29, 11, "Mutation Type", $table_topleft); $ws->write(29, 12, "Non-Tr/Tr", $table_top); $ws->write(29, 13, "Non-Tr", $table_top); $ws->write(29, 14, "Tr", $table_top); $ws->write(29, 15, "P-value", $table_top); $ws->write(29, 16, "FDR q value", $table_top); $ws->write(29, 17, "95% CI", $table_topRight); - - $ws->write(39, 11, "Table 3. Significance of the strand biases", $format_A10Boldleft); - $ws->write(40, 11, "Mutation Type", $table_topleft); $ws->write(40, 12, "Non-Tr/Tr", $table_top); $ws->write(40, 13, "Non-Tr", $table_top); $ws->write(40, 14, "Tr", $table_top); $ws->write(40, 15, "P-value", $table_top); $ws->write(40, 16, "FDR q value", $table_top); $ws->write(40, 17, "95% CI", $table_topRight); - - # For ggplot2 - open(SB, ">", "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandBias.txt") or die "$!: $folderFigure/Stranded_Analysis/$k_file/$k_file-StrandBias.txt\n"; - print SB "Alteration\tStrand\tCount\n"; - - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_ca, $ratio_gt, $percent_ca_NonTr, $percent_ca_Tr) = (0, 0, 0, 0, 0); - if( ($ca_NonTr==0) || ($ca_Tr==0) ) { $ratio_ca = 0; $ratio_gt = 0; $percent_ca_NonTr = 0; $percent_ca_Tr = 0; } - else - { - $ratio_ca = $ca_NonTr/$ca_Tr; $ratio_ca = sprintf("%.2f", $ratio_ca); - $ratio_gt = $ca_Tr/$ca_NonTr; $ratio_gt = sprintf("%.2f", $ratio_gt); - $percent_ca_NonTr = ($ca_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ca_Tr = ($ca_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "C>A\tNonTranscribed\t$ca_NonTr\n", "C>A\tTranscribed\t$ca_Tr\n"; - # C>A - $ws->write(30, 11, "C>A", $table_left); $ws->write(30, 12, $ratio_ca, $table_middleHeader); $ws->write(30, 13, $ca_NonTr, $format_A10); $ws->write(30, 14, $ca_Tr, $format_A10); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($ca_NonTr+$ca_Tr)< 10) - { - if($h_chi2{$k_file}{'C>A'}{'p-value'} eq "NA") { $ws->write_string(30, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10); } - else { $ws->write_string(30, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(30, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10); } - $ws->write(30, 16, $h_chi2{$k_file}{'C>A'}{'FDR'}, $format_A10); $ws->write(30, 17, $h_chi2{$k_file}{'C>A'}{'ConfInt'}, $table_right); - # G>T - $ws->write(41, 11, "G>T", $table_left); $ws->write(41, 12, $ratio_gt, $table_middleHeader); $ws->write(41, 13, $ca_Tr, $format_A10); $ws->write(41, 14, $ca_NonTr, $format_A10); - if(($ca_NonTr+$ca_Tr)< 10) - { - if($h_chi2{$k_file}{'C>A'}{'p-value'} eq "NA") { $ws->write_string(41, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10); } - else { $ws->write_string(41, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(41, 15, $h_chi2{$k_file}{'C>A'}{'p-value'}, $format_A10); } - $ws->write(41, 16, $h_chi2{$k_file}{'C>A'}{'FDR'}, $format_A10); $ws->write(41, 17, $h_chi2{$k_file}{'C>A'}{'ConfInt'}, $table_right); - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_cg, $ratio_gc, $percent_cg_NonTr, $percent_cg_Tr) = (0, 0, 0, 0, 0); - if( ($cg_NonTr==0) || ($cg_Tr==0) ) { $ratio_cg = 0; $ratio_gc = 0; $percent_cg_NonTr = 0; $percent_cg_Tr = 0; } - else - { - $ratio_cg = $cg_NonTr/$cg_Tr; $ratio_cg = sprintf("%.2f", $ratio_cg); - $ratio_gc = $cg_Tr/$cg_NonTr; $ratio_gc = sprintf("%.2f", $ratio_gc); - $percent_cg_NonTr = ($cg_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_cg_Tr = ($cg_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "C>G\tNonTranscribed\t$cg_NonTr\n", "C>G\tTranscribed\t$cg_Tr\n"; - # C>G - $ws->write(31, 11, "C>G", $table_left); $ws->write(31, 12, $ratio_cg, $table_middleHeader); $ws->write(31, 13, $cg_NonTr, $format_A10); $ws->write(31, 14, $cg_Tr, $format_A10); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($cg_NonTr+$cg_Tr)< 10) - { - if($h_chi2{$k_file}{'C>G'}{'p-value'} eq "NA") { $ws->write_string(31, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10); } - else { $ws->write_string(31, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(31, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10); } - $ws->write(31, 16, $h_chi2{$k_file}{'C>G'}{'FDR'}, $format_A10); $ws->write(31, 17, $h_chi2{$k_file}{'C>G'}{'ConfInt'}, $table_right); - # G>C - $ws->write(42, 11, "G>C", $table_left); $ws->write(42, 12, $ratio_gc, $table_middleHeader); $ws->write(42, 13, $cg_Tr, $format_A10); $ws->write(42, 14, $cg_NonTr, $format_A10); - if(($cg_NonTr+$cg_Tr)< 10) - { - if($h_chi2{$k_file}{'C>G'}{'p-value'} eq "NA") { $ws->write_string(42, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10); } - else { $ws->write_string(42, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(42, 15, $h_chi2{$k_file}{'C>G'}{'p-value'}, $format_A10); } - $ws->write(42, 16, $h_chi2{$k_file}{'C>G'}{'FDR'}, $format_A10); $ws->write(42, 17, $h_chi2{$k_file}{'C>G'}{'ConfInt'}, $table_right); - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_ct, $ratio_ga, $percent_ct_NonTr, $percent_ct_Tr) = (0, 0, 0, 0, 0); - if( ($ct_NonTr==0) || ($ct_Tr==0) ) { $ratio_ct = 0; $ratio_ga = 0; $percent_ct_NonTr = 0; $percent_ct_Tr = 0; } - else - { - $ratio_ct = $ct_NonTr/$ct_Tr; $ratio_ct = sprintf("%.2f", $ratio_ct); - $ratio_ga = $ct_Tr/$ct_NonTr; $ratio_ga = sprintf("%.2f", $ratio_ga); - $percent_ct_NonTr = ($ct_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ct_Tr = ($ct_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "C>T\tNonTranscribed\t$ct_NonTr\n", "C>T\tTranscribed\t$ct_Tr\n"; - # C>T - $ws->write(32, 11, "C>T", $table_left); $ws->write(32, 12, $ratio_ct, $table_middleHeader); $ws->write(32, 13, $ct_NonTr, $format_A10); $ws->write(32, 14, $ct_Tr, $format_A10); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($ct_NonTr+$ct_Tr)< 10) - { - if($h_chi2{$k_file}{'C>T'}{'p-value'} eq "NA") { $ws->write_string(32, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10); } - else { $ws->write_string(32, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(32, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10); } - $ws->write(32, 16, $h_chi2{$k_file}{'C>T'}{'FDR'}, $format_A10); $ws->write(32, 17, $h_chi2{$k_file}{'C>T'}{'ConfInt'}, $table_right); - # G>A - $ws->write(43, 11, "G>A", $table_left); $ws->write(43, 12, $ratio_ga, $table_middleHeader); $ws->write(43, 13, $ct_Tr, $format_A10); $ws->write(43, 14, $ct_NonTr, $format_A10); - if(($ct_NonTr+$ct_Tr)< 10) - { - if($h_chi2{$k_file}{'C>T'}{'p-value'} eq "NA") { $ws->write_string(43, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10); } - else { $ws->write_string(43, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(43, 15, $h_chi2{$k_file}{'C>T'}{'p-value'}, $format_A10); } - $ws->write(43, 16, $h_chi2{$k_file}{'C>T'}{'FDR'}, $format_A10); $ws->write(43, 17, $h_chi2{$k_file}{'C>T'}{'ConfInt'}, $table_right); - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_ta, $ratio_at, $percent_ta_NonTr, $percent_ta_Tr) = (0, 0, 0, 0, 0); - if( ($ta_NonTr==0) || ($ta_Tr==0) ) { $ratio_ta = 0; $ratio_at = 0; $percent_ta_NonTr = 0; $percent_ta_Tr = 0; } - else - { - $ratio_ta = $ta_NonTr/$ta_Tr; $ratio_ta = sprintf("%.2f", $ratio_ta); - $ratio_at = $ta_Tr/$ta_NonTr; $ratio_at = sprintf("%.2f", $ratio_at); - $percent_ta_NonTr = ($ta_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_ta_Tr = ($ta_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "T>A\tNonTranscribed\t$ta_NonTr\n", "T>A\tTranscribed\t$ta_Tr\n"; - # T>A - $ws->write(33, 11, "T>A", $table_left); $ws->write(33, 12, $ratio_ta, $table_middleHeader); $ws->write(33, 13, $ta_NonTr, $format_A10); $ws->write(33, 14, $ta_Tr, $format_A10); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($ta_NonTr+$ta_Tr)< 10) - { - if($h_chi2{$k_file}{'T>A'}{'p-value'} eq "NA") { $ws->write_string(33, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10); } - else { $ws->write_string(33, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(33, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10); } - $ws->write(33, 16, $h_chi2{$k_file}{'T>A'}{'FDR'}, $format_A10); $ws->write(33, 17, $h_chi2{$k_file}{'T>A'}{'ConfInt'}, $table_right); - # A>T - $ws->write(44, 11, "A>T", $table_left); $ws->write(44, 12, $ratio_at, $table_middleHeader); $ws->write(44, 13, $ta_Tr, $format_A10); $ws->write(44, 14, $ta_NonTr, $format_A10); - if(($ta_NonTr+$ta_Tr)< 10) - { - if($h_chi2{$k_file}{'T>A'}{'p-value'} eq "NA") { $ws->write_string(44, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10); } - else { $ws->write_string(44, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(44, 15, $h_chi2{$k_file}{'T>A'}{'p-value'}, $format_A10); } - $ws->write(44, 16, $h_chi2{$k_file}{'T>A'}{'FDR'}, $format_A10); $ws->write(44, 17, $h_chi2{$k_file}{'T>A'}{'ConfInt'}, $table_right); - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_tc, $ratio_ag, $percent_tc_NonTr, $percent_tc_Tr) = (0, 0, 0, 0, 0); - if( ($tc_NonTr==0) || ($tc_Tr==0) ) { $ratio_tc = 0; $ratio_ag = 0; $percent_tc_NonTr = 0; $percent_tc_Tr = 0; } - else - { - $ratio_tc = $tc_NonTr/$tc_Tr; $ratio_tc = sprintf("%.2f", $ratio_tc); - $ratio_ag = $tc_Tr/$tc_NonTr; $ratio_ag = sprintf("%.2f", $ratio_ag); - $percent_tc_NonTr = ($tc_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_tc_Tr = ($tc_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "T>C\tNonTranscribed\t$tc_NonTr\n", "T>C\tTranscribed\t$tc_Tr\n"; - # T>C - $ws->write(34, 11, "T>C", $table_left); $ws->write(34, 12, $ratio_tc, $table_middleHeader); $ws->write(34, 13, $tc_NonTr, $format_A10); $ws->write(34, 14, $tc_Tr, $format_A10); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($tc_NonTr+$tc_Tr)< 10) - { - if($h_chi2{$k_file}{'T>C'}{'p-value'} eq "NA") { $ws->write_string(34, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10); } - else { $ws->write_string(34, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(34, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10); } - $ws->write(34, 16, $h_chi2{$k_file}{'T>C'}{'FDR'}, $format_A10); $ws->write(34, 17, $h_chi2{$k_file}{'T>C'}{'ConfInt'}, $table_right); - # A>G - $ws->write(45, 11, "A>G", $table_left); $ws->write(45, 12, $ratio_ag, $table_middleHeader); $ws->write(45, 13, $tc_Tr, $format_A10); $ws->write(45, 14, $tc_NonTr, $format_A10); - if(($tc_NonTr+$tc_Tr)< 10) - { - if($h_chi2{$k_file}{'T>C'}{'p-value'} eq "NA") { $ws->write_string(45, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10); } - else { $ws->write_string(45, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10ItalicRed); } - } - else { $ws->write_string(45, 15, $h_chi2{$k_file}{'T>C'}{'p-value'}, $format_A10); } - $ws->write(45, 16, $h_chi2{$k_file}{'T>C'}{'FDR'}, $format_A10); $ws->write(45, 17, $h_chi2{$k_file}{'T>C'}{'ConfInt'}, $table_right); - - #-----------------------------------------------------------------------------------------------------# - my ($ratio_tg, $ratio_ac, $percent_tg_NonTr, $percent_tg_Tr) = (0, 0, 0, 0, 0); - if( ($tg_NonTr==0) || ($tg_Tr==0) ) { $ratio_tg = 0; $ratio_ac = 0; $percent_tg_NonTr = 0; $percent_tg_Tr = 0; } - else - { - $ratio_tg = $tg_NonTr/$tg_Tr; $ratio_tg = sprintf("%.2f", $ratio_tg); - $ratio_ac = $tg_Tr/$tg_NonTr; $ratio_ac = sprintf("%.2f", $ratio_ac); - $percent_tg_NonTr = ($tg_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; $percent_tg_Tr = ($tg_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; - } - print SB "T>G\tNonTranscribed\t$tg_NonTr\n", "T>G\tTranscribed\t$tg_Tr\n"; - # T>G - $ws->write(35, 11, "T>G", $table_bottomleft); $ws->write(35, 12, $ratio_tg, $table_middleHeader2); $ws->write(35, 13, $tg_NonTr, $table_bottom); $ws->write(35, 14, $tg_Tr, $table_bottom); - # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 - if(($tg_NonTr+$tg_Tr)< 10) - { - if($h_chi2{$k_file}{'T>G'}{'p-value'} eq "NA") { $ws->write_string(35, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottom); } - else { $ws->write_string(35, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottomItalicRed); } - } - else { $ws->write_string(35, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottom); } - $ws->write(35, 16, $h_chi2{$k_file}{'T>G'}{'FDR'}, $table_bottom); $ws->write(35, 17, $h_chi2{$k_file}{'T>G'}{'ConfInt'}, $table_bottomRight); - # A>C - $ws->write(46, 11, "A>C", $table_bottomleft); $ws->write(46, 12, $ratio_ac, $table_middleHeader2); $ws->write(46, 13, $tg_Tr, $table_bottom); $ws->write(46, 14, $tg_NonTr, $table_bottom); - if(($tg_NonTr+$tg_Tr)< 10) - { - if($h_chi2{$k_file}{'T>G'}{'p-value'} eq "NA") { $ws->write_string(46, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottom); } - else { $ws->write_string(46, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottomItalicRed); } - } - else { $ws->write_string(46, 15, $h_chi2{$k_file}{'T>G'}{'p-value'}, $table_bottom); } - $ws->write(46, 16, $h_chi2{$k_file}{'T>G'}{'FDR'}, $table_bottom); $ws->write(46, 17, $h_chi2{$k_file}{'T>G'}{'ConfInt'}, $table_bottomRight); - - ### Write a warning message when NonTr+Tr < 10 - my $format_italic_red = $wb->add_format(font=>'Arial', size=>10, italic=>1, color => 'red'); - - if( (($ca_NonTr+$ca_Tr)< 10) || (($cg_NonTr+$cg_Tr)< 10) || (($ct_NonTr+$ct_Tr)< 10) || (($ta_NonTr+$ta_Tr)< 10) || (($tc_NonTr+$tc_Tr)< 10) || (($tg_NonTr+$tg_Tr)< 10) ) - { - $ws->write(36, 11, "Warning message: chi-squared approximation may be incorrect because the number of SBS", $format_italic_red); - $ws->write(37, 11, "on Non-transcribed and transcribed strand is lower than 10", $format_italic_red); - } - - close SB; - - - ########################################################################################################################################################### - ################################################################### Write SBS Per Chr ##################################################################### - ########################################################################################################################################################### - # For the HTML report - open(SBSPerChr, ">", "$folderFigure/Distribution_SBS_Per_Chromosomes/$k_file-DistributionSNVS_per_chromosome.txt") or die "$!: $folderFigure/Distribution_SBS_Per_Chromosomes/$k_file-DistributionSNVS_per_chromosome.txt\n"; - print SBSPerChr "\tPearson\t$refH_file->{$k_file}{'SBSPerChr'}{'AllMutType'}\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>A:T"}{'Pearson'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>G:C"}{'Pearson'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>T:A"}{'Pearson'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>A:T"}{'Pearson'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>C:G"}{'Pearson'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>G:C"}{'Pearson'},"\n"; - print SBSPerChr "Chr\tSize\tAll SBS\tC:G>A:T\tC:G>G:C\tC:G>T:A\tT:A>A:T\tT:A>C:G\tT:A>G:C\n"; - - my $row_SBSPerChr = $row_SBSDistrBySegAndFunc_CG + 8; # Line 158 - - # Write the Pearson coefficient - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>A:T"}{'Pearson'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+4, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>G:C"}{'Pearson'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+5, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>T:A"}{'Pearson'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+6, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>A:T"}{'Pearson'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>C:G"}{'Pearson'}, $format_A10); - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+8, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>G:C"}{'Pearson'}, $formatT_right); - - # Write the chromosome number and their sizes / Write the total of SBS per chromosome - my $line=0; - - foreach my $chromosome (sort keys %chromosomes) - { - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg, $chromosome, $formatT_left); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+1, $chromosomes{$chromosome}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}, $format_A10); - - # Write the count per mutation - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>A:T"}{'CHR'}{$chromosome}{'chr'}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+4, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>G:C"}{'CHR'}{$chromosome}{'chr'}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+5, $refH_file->{$k_file}{'SBSPerChr'}{"C:G>T:A"}{'CHR'}{$chromosome}{'chr'}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+6, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>A:T"}{'CHR'}{$chromosome}{'chr'}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+7, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>C:G"}{'CHR'}{$chromosome}{'chr'}, $format_A10); - $ws->write($row_SBSPerChr+($line), $colStart_SBSdistrBySeg+8, $refH_file->{$k_file}{'SBSPerChr'}{"T:A>G:C"}{'CHR'}{$chromosome}{'chr'}, $formatT_right); - - - # For the HTML report - print SBSPerChr "$chromosome\t", $chromosomes{$chromosome},"\t", $refH_file->{$k_file}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>A:T"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>G:C"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"C:G>T:A"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>A:T"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>C:G"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$k_file}{'SBSPerChr'}{"T:A>G:C"}{'CHR'}{$chromosome}{'chr'},"\n"; - $line++; - } - - # Write the Pearson coefficient for the total number of SBS - $ws->write($row_SBSDistrBySegAndFunc_CG+6, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'SBSPerChr'}{'AllMutType'}, $format_A10); - $ws->write($row_SBSPerChr+(keys %chromosomes), $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'TotalSBSGenomic'}, $formatT_bottomHeader); - - print SBSPerChr "\t\t$refH_file->{$k_file}{'TotalSBSGenomic'}\n"; - close SBSPerChr; - - - - ########################################################################################################################################################### - ####################################################################### Impact on protein ################################################################# - ########################################################################################################################################################### - $ws->write(29, 6, "Table 2. Frequency and counts of functional impact", $format_A10Boldleft); - $ws->set_column(6, 6, 13); $ws->set_column(10, 10, 15); - $ws->write(30, 6, "RefSeq gene", $table_topleft); $ws->write(30, 7, "", $table_top); $ws->write(30, 8, "Percent", $table_top); $ws->write(30, 9, "Count", $table_topRight); - my $lImpactSBS = 31; - open(IMPACTSBS, ">", "$folderFigure/Impact_protein_sequence/$k_file/$k_file-DistributionExoFunc.txt") or die "$!: $folderFigure/Impact_protein_sequence/$k_file/$k_file-DistributionExoFunc.txt\n"; - print IMPACTSBS "AA_Change\tCount\tPercent\n"; - - # Pie chart with the distribution of SBS vs Indel - open(SBSINDEL, ">", "$folderFigure/Overall_mutation_distribution/$k_file/$k_file-OverallMutationDistribution.txt") or die "$!: $folderFigure/Overall_mutation_distribution/$k_file/$k_file-OverallMutationDistribution.txt\n"; - print SBSINDEL "Variant_type\tCount\tPercent\n"; - my ($deletion, $insertion) = (0, 0); - - - foreach my $k_exoFunc(sort keys $refH_file->{$k_file}{'ImpactSBS'}) - { - my $percent = ($refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc} / $refH_file->{$k_file}{'TotalMutGenomic'})*100; - $percent = sprintf("%.2f", $percent); - - if($k_exoFunc eq "NA") { print IMPACTSBS "Not_Applicable\t$percent\t$refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc}\n"; } - else { my $temp = $k_exoFunc; $temp =~ s/ /_/g; print IMPACTSBS "$temp\t$percent\t$refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc}\n"; } - - $ws->write($lImpactSBS, 6, $k_exoFunc, $table_left2); $ws->write($lImpactSBS, 8, $percent, $format_A10); $ws->write($lImpactSBS, 9, $refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc}, $table_right); - $lImpactSBS++; - - # Pie chart with the distribution of SBS vs Indel - if($k_exoFunc =~ /deletion/i) { $deletion += $refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc}; } - elsif($k_exoFunc =~ /insertion/i) { $insertion += $refH_file->{$k_file}{'ImpactSBS'}{$k_exoFunc}; } - } - close IMPACTSBS; - $ws->write($lImpactSBS, 9, $refH_file->{$k_file}{'TotalMutGenomic'}, $table_bottomrightHeader); - $ws->write($lImpactSBS, 6, "", $table_bottomleft); $ws->write($lImpactSBS, 7, "", $table_bottom); $ws->write($lImpactSBS, 8, "", $table_bottom); - - # Pie chart with the distribution of SBS vs Indel - my $percentSBSIndel = ($deletion/$refH_file->{$k_file}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); - print SBSINDEL "Deletion\t$deletion\t$percentSBSIndel\n"; - $percentSBSIndel = ($insertion/$refH_file->{$k_file}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); - print SBSINDEL "Insertion\t$insertion\t$percentSBSIndel\n"; - $percentSBSIndel = ($refH_file->{$k_file}{TotalSBSGenomic}/$refH_file->{$k_file}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); - print SBSINDEL "SBS\t$refH_file->{$k_file}{TotalSBSGenomic}\t$percentSBSIndel\n"; - close SBSINDEL; - - ########################################################################################################################################################### - ######################################################## SEQUENCE CONTEXT ON GENOMIC STRAND ############################################################### - ########################################################################################################################################################### - my $row_SeqContext6 = 4; - # Count the total of mutations for 6 mutation types on genomic strand - my ($c_ca6_g, $c_cg6_g, $c_ct6_g, $c_ta6_g, $c_tc6_g, $c_tg6_g) = (0,0,0, 0,0,0); - my ($p_ca6_g, $p_cg6_g, $p_ct6_g, $p_ta6_g, $p_tc6_g, $p_tg6_g) = (0,0,0, 0,0,0); - my $maxValue = 0; # For the heatmap - - # For checking if the total number of SBS is correct - my $total_SBS_genomic = 0; - - - open(HEATMAPCGENOMIC, ">", "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapCount-Genomic.txt") or die "$!: $folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapCount-Genomic.txt\n"; - print HEATMAPCGENOMIC "\tC>A\tC>G\tC>T\tT>A\tT>C\tT>G\n"; - open(HEATMAPPGENOMIC, ">", "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapPercent-Genomic.txt") or die "$!: $folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapPercent-Genomic.txt\n"; - print HEATMAPPGENOMIC "\tC>A\tC>G\tC>T\tT>A\tT>C\tT>G\n"; - - ## Bar plot NMF like - open(BARPLOTNMFLIKE, ">", "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-MutationSpectraPercent-Genomic.txt") or die "$!: $folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-MutationSpectraPercent-Genomic.txt\n"; - print BARPLOTNMFLIKE "alteration\tcontext\tvalue\n"; - - foreach my $k_context (sort keys $refH_file->{$k_file}{'SeqContextG'}) - { - if( ($k_context =~ /N/) || (length($k_context) != 3) ) { next; } - - # Write the context: 6 mut type on genomic strand - $ws->write($row_SeqContext6 , $colStart_matrixSeqContext+3, $k_context, $format_A10); $ws->write($row_SeqContext6 , $colStart_matrixSeqContext+13, $k_context, $format_A10); - - foreach my $k_mutation (sort keys $refH_file->{$k_file}{'SeqContextG'}{$k_context}) - { - # For checking the total number of SBS - $total_SBS_genomic += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - - # Calculate the percentages - my $percent = 0; - if($refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation} == 0) { $percent = 0; } - else - { - $percent = ($refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation} / $refH_file->{$k_file}{'TotalSBSGenomic'}) * 100; - $percent = sprintf("%.2f", $percent); - } - - # For representing the sequence context with a bar plot (NMF like style) - print BARPLOTNMFLIKE $k_mutation,"\t", $k_context,"\t", $percent,"\n"; - - if($k_mutation eq "C>A") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+4, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$k_context\t$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\t"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+14, $percent, $format_A10); - print HEATMAPPGENOMIC "$k_context\t$percent\t"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'C>A'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'C>A'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_ca6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_ca6_g += $percent; - } - if($k_mutation eq "C>G") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+5, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\t"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+15, $percent, $format_A10); - print HEATMAPPGENOMIC "$percent\t"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'C>G'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'C>G'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_cg6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_cg6_g += $percent; - } - if($k_mutation eq "C>T") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+6, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\t"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+16, $percent, $format_A10); - print HEATMAPPGENOMIC "$percent\t"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'C>T'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'C>T'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_ct6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_ct6_g += $percent; - } - if($k_mutation eq "T>A") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+7, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\t"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+17, $percent, $format_A10); - print HEATMAPPGENOMIC "$percent\t"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'T>A'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'T>A'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_ta6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_ta6_g += $percent; - } - if($k_mutation eq "T>C") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+8, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\t"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+18, $percent, $format_A10); - print HEATMAPPGENOMIC "$percent\t"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'T>C'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'T>C'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_tc6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_tc6_g += $percent; - } - if($k_mutation eq "T>G") - { - ### COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+9, $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}, $format_A10); - # Write the count for the heatmap - print HEATMAPCGENOMIC "$refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}\n"; - - ### PERCENTAGE - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+19, $percent, $format_A10); - print HEATMAPPGENOMIC "$percent\n"; - - # For NMF input - my $count = $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{'T>G'}}, $count); } - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{'T>G'}}, $percent); } - - # For the heatmap - if($percent >= $maxValue) { $maxValue = $percent; } - - # For the total amount per mutation types - $c_tg6_g += $refH_file->{$k_file}{'SeqContextG'}{$k_context}{$k_mutation}; - $p_tg6_g += $percent; - } - } - $row_SeqContext6++; - } - close HEATMAPCGENOMIC; close HEATMAPPGENOMIC; - close BARPLOTNMFLIKE; - - - # Write the total number of SBS per mutation type: COUNT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+4, $c_ca6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+5, $c_cg6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+6, $c_ct6_g, $formatT_bottomHeader2); - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+7, $c_ta6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+8, $c_tc6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+9, $c_tg6_g, $formatT_bottomHeader2); - if($total_SBS_genomic != $refH_file->{$k_file}{'TotalSBSGenomic'}) { print STDERR "Error in the calculation of the total number of SBS on the genomic strand!!!!\nFrom hash table $refH_file->{$k_file}{'TotalSBSGenomic'}\tVS\t$total_SBS_genomic\n"; exit; } - - # Write the total number of SBS per mutation type: PERCENT - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+14, $p_ca6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+15, $p_cg6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+16, $p_ct6_g, $formatT_bottomHeader2); - $ws->write($row_SeqContext6, $colStart_matrixSeqContext+17, $p_ta6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+18, $p_tc6_g, $formatT_bottomHeader2); $ws->write($row_SeqContext6, $colStart_matrixSeqContext+19, $p_tg6_g, $formatT_bottomHeader2); - my $totalPercent_genomic = $p_ca6_g + $p_cg6_g + $p_ct6_g + $p_ta6_g + $p_tc6_g + $p_tg6_g; $totalPercent_genomic = sprintf("%.0f", $totalPercent_genomic); - if($totalPercent_genomic != 100) { print STDERR "Error in the calculation of the total percentages on the genomic strand!!!\nThe total is equal to=\t$totalPercent_genomic\n"; exit; } - - - #----------------------------------------------------------------------------------------------------------------------------------------------------------------# - # For the input matrix for NMF - if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Sample'}}, $k_file); } - - - ########################################################################################################################################################### - ######################################################## SEQUENCE CONTEXT ON CODING STRAND ############################################################### - ########################################################################################################################################################### - my $row_SeqContext12 = $rowStart_SBSdistrBySeg+6; my $row_SeqContext12Percent = $rowStart_SBSdistrBySeg+27; - # Reset the total count and percent calculated for the strand bias - ($ca_NonTr, $ca_Tr, $cg_NonTr, $cg_Tr, $ct_NonTr, $ct_Tr, $ta_NonTr, $ta_Tr, $tc_NonTr, $tc_Tr, $tg_NonTr, $tg_Tr) = (0,0,0, 0,0,0, 0,0,0, 0,0,0); - ($percent_ca_NonTr, $percent_ca_Tr, $percent_cg_NonTr, $percent_cg_Tr, $percent_ct_NonTr, $percent_ct_Tr, $percent_ta_NonTr, $percent_ta_Tr, $percent_tc_NonTr, $percent_tc_Tr, $percent_tg_NonTr, $percent_tg_Tr) = (0,0,0, 0,0,0, 0,0,0, 0,0,0); - - # For checking if the total number of SBS is correct - my $total_SBS_coding = 0; - - open(COUNT, ">", "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignatureCount.txt") or die "$!: $folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignatureCount.txt\n"; - print COUNT "MutationTypeContext\tStrand\tValue\tSample\n"; - open(PERCENT, ">", "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignaturePercent.txt") or die "$!: $folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignaturePercent.txt\n"; - print PERCENT "MutationTypeContext\tStrand\tValue\tSample\n"; - - foreach my $k_context (sort keys $refH_file->{$k_file}{'SeqContextC'}) - { - if( ($k_context =~ /N/) || (length($k_context) != 3) ) { next; } - - # Write the context: 12 mut type on coding strand - $ws->write($row_SeqContext12 , $colStart_matrixSeqContext, $k_context, $formatT_left); $ws->write($row_SeqContext12Percent , $colStart_matrixSeqContext, $k_context, $formatT_left); - - foreach my $k_mutation (sort keys $refH_file->{$k_file}{'SeqContextC'}{$k_context}) - { - # Percent: 12 mut type on coding strand - my ($percent_NonTr, $percent_Tr) = (0, 0); - if($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} == 0) { $percent_NonTr = 0; } - else { $percent_NonTr = ( $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'TotalSBSCoding'} ) * 100 } - if($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'} == 0) { $percent_Tr = 0; } - else { $percent_Tr = ( $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'} / $refH_file->{$k_file}{'TotalSBSCoding'} ) * 100 } - - - # Calculate the total number for each mutation types - if($k_mutation eq "C>A") - { - $ca_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $ca_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+1, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+2, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_ca_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_ca_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+1, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+2, $percent_Tr, $format_A10); - } - if($k_mutation eq "C>G") - { - $cg_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $cg_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+3, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+4, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_cg_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_cg_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+3, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+4, $percent_Tr, $format_A10); - } - if($k_mutation eq "C>T") - { - $ct_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $ct_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+5, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+6, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_ct_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_ct_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+5, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+6, $percent_Tr, $format_A10); - } - if($k_mutation eq "T>A") - { - $ta_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $ta_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+7, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+8, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_ta_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_ta_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+7, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+8, $percent_Tr, $format_A10); - } - if($k_mutation eq "T>C") - { - $tc_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $tc_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+9, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+10, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_tc_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_tc_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+9, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+10, $percent_Tr, $format_A10); - } - if($k_mutation eq "T>G") - { - $tg_NonTr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}; - $tg_Tr += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - - # COUNT : 12 mutation type (stranded bar graph) - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+11, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$k_file\n"; - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+12, $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}, $format_A10); - print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$k_file\n"; - - ## PERCENT : 12 mutation type (stranded bar graph) - my $percent_NonTr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_NonTr = sprintf("%.2f", $percent_NonTr); $percent_tg_NonTr += $percent_NonTr; - my $percent_Tr = ($refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}/$refH_file->{$k_file}{'TotalSBSCoding'})*100; - $percent_Tr = sprintf("%.2f", $percent_Tr); $percent_tg_Tr += $percent_Tr; - print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$k_file\n"; - print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$k_file\n"; - - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+11, $percent_NonTr, $format_A10); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+12, $percent_Tr, $format_A10); - } - - # For checking if the total number of SBS is correct - $total_SBS_coding += $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} + $refH_file->{$k_file}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; - } - $row_SeqContext12++; $row_SeqContext12Percent++; - } - close COUNT; close PERCENT; - - ## Write the total of each mutation types: 12 mut type on coding strand - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+1, $ca_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+2, $ca_Tr, $formatT_bottomHeader2); - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+3, $cg_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+4, $cg_Tr, $formatT_bottomHeader2); - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+5, $ct_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+6, $ct_Tr, $formatT_bottomHeader2); - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+7, $ta_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+8, $ta_Tr, $formatT_bottomHeader2); - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+9, $tc_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+10, $tc_Tr, $formatT_bottomHeader2); - $ws->write($row_SeqContext12, $colStart_matrixSeqContext+11, $tg_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $colStart_matrixSeqContext+12, $tg_Tr, $formatT_bottomHeader2); - # Write the total percentages of each mutation types: 12 mut type on coding strand - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+1, $percent_ca_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+2, $percent_ca_Tr, $formatT_bottomHeader); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+3, $percent_cg_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+4, $percent_cg_Tr, $formatT_bottomHeader); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+5, $percent_ct_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+6, $percent_ct_Tr, $formatT_bottomHeader); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+7, $percent_ta_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+8, $percent_ta_Tr, $formatT_bottomHeader); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+9, $percent_tc_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+10, $percent_tc_Tr, $formatT_bottomHeader); - $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+11, $percent_tg_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $colStart_matrixSeqContext+12, $percent_tg_Tr, $formatT_bottomHeader); - - if($total_SBS_coding == $refH_file->{$k_file}{'TotalSBSCoding'}) { $ws->write($row_SeqContext12, $colStart_matrixSeqContext+13, $refH_file->{$k_file}{'TotalSBSCoding'}, $formatT_bottomHeader2) } - else { print STDERR "Error in the calculation of the total number of SBS on the coding strand!!!!\nFrom hash table $refH_file->{$k_file}{'TotalSBSCoding'}\tVS\t$total_SBS_coding\n"; exit; } - - - my $totalP_SBS_coding = $percent_ca_NonTr + $percent_ca_Tr + $percent_cg_NonTr + $percent_cg_Tr + $percent_ct_NonTr + $percent_ct_Tr + $percent_ta_NonTr + $percent_ta_Tr + $percent_tc_NonTr + $percent_tc_Tr + $percent_tg_NonTr + $percent_tg_Tr; $totalP_SBS_coding = sprintf("%.0f", $totalP_SBS_coding); - if($totalP_SBS_coding != 100) { print STDERR "The percentages for the trinucleotide sequence context on the coding strand for 12 mutation types is not equal to 100!!!\n$totalP_SBS_coding\n"; exit; } - - - ########################################################################################################################################################### - ################################################################### GRAPHS & TABLES ####################################################################### - ########################################################################################################################################################### - Create_Graph($folderFigure, $k_file, $maxValue); - - ## Distribution of SBS into the Excel report (Figure 1 + Table 1) - $ws->write(0, 0, "Graph 1. SBS distribution", $formatT_graphTitle); $ws->set_row(0, 18); - $ws->insert_image(1, 0, "$folder_temp/$k_file-SBS_distribution-Report.png", 0, 0, .2, .2); - $ws->write(29, 0, "Table 1. Frequency and counts of all SBS", $format_A10Boldleft); - $ws->write(30, 0, "Mutation type", $table_topleft); $ws->write(30, 1, "Percentage", $table_top); $ws->write(30, 2, "Count", $table_topRight); - $ws->write(31, 0, "C:G>A:T", $table_left); $ws->write(31, 1, $percent_ca, $format_A10); $ws->write(31, 2, $ca_genomique, $table_right); - $ws->write(32, 0, "C:G>G:C", $table_left); $ws->write(32, 1, $percent_cg, $format_A10); $ws->write(32, 2, $cg_genomique, $table_right); - $ws->write(33, 0, "C:G>T:A", $table_left); $ws->write(33, 1, $percent_ct, $format_A10); $ws->write(33, 2, $ct_genomique, $table_right); - $ws->write(34, 0, "T:A>A:T", $table_left); $ws->write(34, 1, $percent_ta, $format_A10); $ws->write(34, 2, $ta_genomique, $table_right); - $ws->write(35, 0, "T:A>C:G", $table_left); $ws->write(35, 1, $percent_tc, $format_A10); $ws->write(35, 2, $tc_genomique, $table_right); - $ws->write(36, 0, "T:A>G:C", $table_left); $ws->write(36, 1, $percent_tg, $format_A10); $ws->write(36, 2, $tg_genomique, $table_right); - $ws->write(37, 0, "", $table_bottomleft); $ws->write(37, 1, "", $table_bottom); $ws->write(37, 2, $refH_file->{$k_file}{'TotalSBSGenomic'}, $table_bottomrightHeader); - - ## Impact of the SBS on the protein - $ws->write(0, 6, "Graph 2. Impact on protein sequence", $formatT_graphTitle); - $ws->insert_image(1, 6, "$folder_temp/$k_file-DistributionExoFunc-Report.png", 0, 0, .2, .2); - - ## Strand Bias - $ws->write(0, 11, "Graph 3. Stranded distribution of SBS", $formatT_graphTitle); - $ws->insert_image(1, 11, "$folder_temp/$k_file-StrandBias-Report.png", 0, 0, .2, .2); - - ## Stranded signature (Scale the inserted image: width x 0.7, height x 0.8) - $ws->insert_image($rowStart_SBSdistrBySeg+3, $colStart_matrixSeqContext+15, "$folder_temp/$k_file-StrandedSignatureCount-Report.png", 0, 0, .16, .16); - $ws->insert_image($rowStart_SBSdistrBySeg+24, $colStart_matrixSeqContext+15, "$folder_temp/$k_file-StrandedSignaturePercent-Report.png", 0, 0, .16, .16); - - - # Heatamp for the sequence context on the genomic strand (6 mutation types) - $ws->insert_image(4, $colStart_matrixSeqContext, "$folder_temp/$k_file-HeatmapCount-Genomic-Report.png"); - $ws->insert_image(4, $colStart_matrixSeqContext+10, "$folder_temp/$k_file-HeatmapPercent-Genomic-Report.png"); - - - ## Bar plot for representing the sequence context (NMF like style) - `Rscript $pathRScriptMutSpectrum $folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-MutationSpectraPercent-Genomic.txt $k_file $folderFigure/Trinucleotide_Sequence_Context/$k_file $folder_temp $c_ca6_g $c_cg6_g $c_ct6_g $c_ta6_g $c_tc6_g $c_tg6_g 2>&1`; - - # Bar plot for the sequence context on the genomic strand (6 mutation types) - $ws->insert_image(27, $colStart_matrixSeqContext+3, "$folder_temp/$k_file-MutationSpectraPercent-Genomic-Report.png"); - - # Next sample - $row_SumSheet++; - } # End $k_file - - #----------------------------------------------------------------------------------------------------------------------------------------------------------------# - # Write the input matrix for NMF - open(OUTINPUTNMFC, ">", "$folderNMF/Input_NMF_Count.txt") or die "$!: $folderNMF/Input_NMF_Count.txt\n"; # with the count - open(OUTINPUTNMFP, ">", "$folderNMF/Input_NMF_Frequency.txt") or die "$!: $folderNMF/Input_NMF_Frequency.txt\n"; # With the frequency un-normalized - - foreach my $k_sample (@{$h_inputNMF{'Sample'}}) { print OUTINPUTNMFC "\t$k_sample"; print OUTINPUTNMFP "\t$k_sample"; } - print OUTINPUTNMFC "\n"; print OUTINPUTNMFP "\n"; - - my $row_inputNMF = 1; - foreach my $k_context (sort keys $h_inputNMF{'Count'}) - { - $k_context =~ /(\w)_(\w)/; my ($base5, $base3) = ($1, $2); - foreach my $k_mutation (sort keys $h_inputNMF{'Count'}{$k_context}) - { - my ($col_inputNMF_Count, $col_inputNMF_Percent) = (1, 1); - my $contextNMF = $base5."[$k_mutation]".$base3; - $ws_inputNMF_count->write($row_inputNMF, 0, $contextNMF); $ws_inputNMF_percent->write($row_inputNMF, 0, $contextNMF); - print OUTINPUTNMFC $contextNMF,"\t"; print OUTINPUTNMFP $contextNMF,"\t"; - - foreach (@{$h_inputNMF{'Count'}{$k_context}{$k_mutation}}) { print OUTINPUTNMFC "$_\t"; } print OUTINPUTNMFC "\n"; - foreach (@{$h_inputNMF{'Percent'}{$k_context}{$k_mutation}}) { print OUTINPUTNMFP "$_\t"; } print OUTINPUTNMFP "\n"; - - foreach (@{$h_inputNMF{'Count'}{$k_context}{$k_mutation}}) - { - # print "\t$k_context\t$k_mutation\t"; - # print "\t$row_inputNMF\t$col_inputNMF_Count\t$_\n"; - $ws_inputNMF_count->write($row_inputNMF, $col_inputNMF_Count, $_); $col_inputNMF_Count++; - } - foreach (@{$h_inputNMF{'Percent'}{$k_context}{$k_mutation}}) { $ws_inputNMF_percent->write($row_inputNMF, $col_inputNMF_Percent, $_); $col_inputNMF_Percent++; } - $row_inputNMF++; - } - } - close OUTINPUTNMFP; close OUTINPUTNMFC; - - - # Close the workbook - $wb->close(); - } - # Calculate the chi2 for the strand bias - sub CalculateChi2 - { - my ($refH_file, $folderChi2) = @_; - - # No value for the chi2 - if(scalar (keys $refH_file) == 0) { print STDERR "No value for calculating the chi2 for the strand bias\n"; exit; } - - # Strand bias for one mutation type for all the samples - my %h_tempchi2 = (); - my ($ca_NonTr, $ca_Tr, $cg_NonTr, $cg_Tr, $ct_NonTr, $ct_Tr, $ta_NonTr, $ta_Tr, $tc_NonTr, $tc_Tr, $tg_NonTr, $tg_Tr) = (0,0,0,0,0,0, 0,0,0,0,0,0); - - my $nb_file = 0; - - foreach my $k_file (sort keys $refH_file) - { - $nb_file++; - foreach my $k_func (sort keys $refH_file->{$k_file}{'6mutType'}) - { - foreach my $k_mutation (sort keys $refH_file->{$k_file}{'6mutType'}{$k_func}) - { - if($k_mutation eq "C:G>A:T") - { - $h_tempchi2{'C>A'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'C>A'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - if($k_mutation eq "C:G>G:C") - { - $h_tempchi2{'C>G'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'C>G'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - if($k_mutation eq "C:G>T:A") - { - $h_tempchi2{'C>T'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'C>T'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - if($k_mutation eq "T:A>A:T") - { - $h_tempchi2{'T>A'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'T>A'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - if($k_mutation eq "T:A>C:G") - { - $h_tempchi2{'T>C'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'T>C'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - if($k_mutation eq "T:A>G:C") - { - $h_tempchi2{'T>G'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; - $h_tempchi2{'T>G'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; - } - } - } - } - - # Create the input file for NMF - open(CHI2, ">", "$folderChi2/Input_chi2_strandBias.txt") or die "$!: $folderChi2/Input_chi2_strandBias.txt\n"; - print CHI2 "SampleName\tNonTr\tTr\tAlteration\n"; - - foreach my $k_mutation (sort keys %h_tempchi2) - { - foreach my $k_file (sort keys $h_tempchi2{$k_mutation}) - { - print CHI2 "$k_file\t$h_tempchi2{$k_mutation}{$k_file}{'NonTr'}\t$h_tempchi2{$k_mutation}{$k_file}{'Tr'}\t$k_mutation\n"; - } - } - close CHI2; - - - # Open the connection with R - my $R = Statistics::R->new() or die "Impossible to create a communication bridge with R\n"; - - $R->send(qq`## Load the data. There is one column with the mutation type and the sample name but it's just for knowing what is corresponding to each line. The two columns with the number of variant per strand would be sufficient. - strBias<-read.delim("$folderChi2/Input_chi2_strandBias.txt", dec=".");`); - $R->send(q`# Chi2 - pValChi2 <- c() # First I create an empty vector and then I apply a for on the data load - pValChi2_round <- c() # Empty vector with the rounded p-values - confInt <- c() # Empty vector for the confident interval - proportion <- c() # Empty vector for the proportion of NonTr compared to the (NonTr+Tr) - sampleSize <- c() # Empty vector for the count of samples in NonTr and Tr - # For Pool_Data save the p-values in a different vector for not having them for the FDR - pValChi2_PoolData <- c() - pValChi2_PoolData_Round <- c() - - j = 1 # Timer for pValChi2_PoolData vector - k = 1 # Timer for pValChi2 - - for(i in 1:nrow(strBias)) - { - if(! sum(strBias[i,2:3]) == 0) - { - # For Pool_Data - if(strBias[i,1] == "Pool_Data") - { - pValChi2_PoolData[j] <- prop.test(x=strBias[i,2],n=sum(strBias[i,2:3]),p=0.5)$p.value - j <- j+1 - } - # For the other sample(s) - else - { - # Calculate the p-value - pValChi2[k] <- prop.test(x=strBias[i,2],n=sum(strBias[i,2:3]),p=0.5)$p.value - k <- k+1 - } - - # Calculate the confidence interval - temp <- prop.test(x=strBias[i,2],n=sum(strBias[i,2:3]),p=0.5)$conf.int - confInt[i] <- paste0("[", round(temp[1],2), "-", round(temp[2],2), "]") # Same as paste(sep="") - - # Save the proportion - proportion[i] <- strBias[i,2] / sum(strBias[i,2:3]) - - # Save the sample size (count on NonTr and Tr) - sampleSize[i] <- paste(strBias[i,2], strBias[i,3], sep="-") - } else - { - if(strBias[i,1] == "Pool_Data") - { - pValChi2_PoolData[j] <- NA - pValChi2_PoolData_Round[j] <- NA - j <- j+1 - } - else - { - # Not enough effective for the test - pValChi2[k] <- NA - confInt[k] <- NA - proportion[k] <- NA - sampleSize[k] <- NA - pValChi2_round[k] <- NA - k <- k+1 - } - } - } - # Adjust with FDR - FDR<-p.adjust(pValChi2, method="BH") - - # Rount the p-value - for(i in 1:nrow(strBias)) - { - pValChi2_round[i] <- format(pValChi2[i], scientific=T, digits=3) - } - - # The option for the pool is specified - if(!is.null(pValChi2_PoolData)) - { - # Round the p-value for Pool_Data - for(i in 1:6) - { - pValChi2_PoolData_Round[i] <- format(pValChi2_PoolData[i], scientific=T, digits=3) - } - } - - - # I create a dataframe for add what I want - outputChi2 <- data.frame(round(strBias[,2]/strBias[,3], digits=2), sampleSize, round(proportion, 3), confInt) - outputChi2$Mut.type <- strBias$Alteration - outputChi2$SampleName <- strBias$SampleName - colnames(outputChi2)[1:6]<-c("Strand_Bias", "NonTr-Tr", "Proportion", "Confidence Interval", "Mutation_Type", "SampleName") - - # Transform the data frame into a matrix for adding the p-value for the samples and Pool_Data - matrix <- as.matrix(outputChi2) - tempColPValFDR <- matrix(, nrow=length(sampleSize), ncol = 2) # Create an empty matrix with 2 columns for adding the p-value and the FDR - matrix <- cbind(matrix, tempColPValFDR) - j = 1 # Timer for all the sample - k = 1 # Timer for Pool_Data - for(i in 1:nrow(matrix)) - { - if(matrix[i,6] == "Pool_Data") - { - matrix[i,7] <- pValChi2_PoolData_Round[k] - matrix[i,8] <- "NA" # No FDR for Pool_Data - k = k+1 - } - else - { - matrix[i,7] <- pValChi2_round[j] - matrix[i,8] <- round(FDR[j], 3) - j = j+1 - } - } - # Reorder the columns - matrix <- cbind(matrix[,1:3], matrix[,7], matrix[,8], matrix[,4:6]) - colnames(matrix)[4] <- "P-val-Chi2" - colnames(matrix)[5] <- "FDR"`); - - $R->send(qq`# Export the file - # dec=".": Set the separator for the decimal by "." - write.table(matrix,file="$folderChi2/Output_chi2_strandBias.txt",quote = FALSE,sep="\t",row.names = FALSE,dec=".");`); - - # Stop the connection with R - $R->stop(); - } - # Pearson correlation - sub PearsonCoefficient - { - our ($refH_file, $filename) = @_; - - #### Calculate the Pearson coefficient - my @total_SBS = (); # Pearson for all mutation types - - # Create a 2D array - foreach my $k_mutation (sort keys $refH_file->{$filename}{'SBSPerChr'}) - { - my $x = []; - my $correlation = 0; - - if($k_mutation eq "AllMutType") { next; } - elsif($k_mutation eq "TotalPerChr") { next; } - elsif($k_mutation eq "ChrSize") { next; } - else - { - my $testZero = 0; # The correlation function doesn't works if all the variables are equal to zero - # generate an anonymous 2D array where $x->[1] is the row - # $x->[1][1] is the value in row 1 column 1 and $x->[1][2] is the value of row 1 column 2 - # once you build the entire array, pass it to the correlation subroutine - my $i=1; - while ( my ($chromosome, $lenght) = each (%chromosomes)) - { - $x->[$i][1] = $lenght; # First column contains the chromosome size - $x->[$i][2] = $refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'}{$chromosome}{'chr'}; # Second column contains the count of SBS - if($refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'}{$chromosome}{'chr'}==0) { $testZero++; } - $i++; - } - if( $testZero == keys $refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'} ) { $correlation = 0; } - # Pass the 2D array to the correlation subroutine - else { $correlation = correlation($x); } - - $refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'Pearson'} = $correlation; # Pearson per mutation type - } - } - - #generate an anonymous 2D array for all mutation type - my $testZero = 0; - my $x = []; - my $correlation = 0; - my $i=1; - while ( my ($chromosome, $lenght) = each (%chromosomes)) - { - $x->[$i][1] = $lenght; - $x->[$i][2] = $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}; - $i++; - } - if($testZero == keys $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}) { $correlation = 0; } - else { $correlation = correlation($x); } - # Pass the 2D array to the correlation subroutine - $refH_file->{$filename}{'SBSPerChr'}{'AllMutType'} = $correlation; - - sub correlation - { - my ($x) = @_; - my ($mean_x,$mean_y) = mean($x); - my $ssxx=ss($x,$mean_x,$mean_y,1,1); - my $ssyy=ss($x,$mean_x,$mean_y,2,2); - my $ssxy=ss($x,$mean_x,$mean_y,1,2); - my $correl=correl($ssxx,$ssyy,$ssxy);; - my $xcorrel=sprintf("%.2f",$correl); - return($xcorrel); - - sub mean - { - my ($x)=@_; - my $num = scalar(@{$x}) - 2; - my $sum_x = '0'; - my $sum_y = '0'; - for (my $i = 2; $i < scalar(@{$x}); ++$i) - { - $sum_x += $x->[$i][1]; - $sum_y += $x->[$i][2]; - } - my $mu_x = $sum_x / $num; - my $mu_y = $sum_y / $num; - return($mu_x,$mu_y); - } - - ### ss = sum of squared (deviations to the mean) - sub ss - { - my ($x,$mean_x,$mean_y,$one,$two)=@_; - my $sum = '0'; - for (my $i=2;$i<scalar(@{$x});++$i) - { - $sum += ($x->[$i][$one]-$mean_x)*($x->[$i][$two]-$mean_y); - } - return $sum; - } - - sub correl - { - my($ssxx,$ssyy,$ssxy)=@_; - - my ($sign, $correl) = (0,0); - if(abs($ssxy) == 0) { $sign = 0 } - else { $sign=$ssxy/abs($ssxy); } - - if( ($ssxx==0) || ($ssyy==0) ) { $correl = 0 } - else { $correl=$sign*sqrt($ssxy*$ssxy/($ssxx*$ssyy)); } - - return $correl; - } - } - } - # Complement bases (for the sequence context) - sub complement - { - if($_[0] eq "A") { return "T"; } - if($_[0] eq "C") { return "G"; } - if($_[0] eq "G") { return "C"; } - if($_[0] eq "T") { return "A"; } - } - # Create and write some graphics - sub Create_Graph - { - our ($folderFigure, $filename, $maxValue) = @_; - - # Open the connection with R - my $R = Statistics::R->new() or die "Impossible to create a communication bridge with R\n"; - $R->startR() ; - # Load the Library - $R->send(q`library(ggplot2)`); - $R->send(q`library(gplots)`); - $R->send(q`library(gtable)`); - - - $R->send(qq`########################################## - ## OVERALL MUTATION DISTRIBUTION ## - ########################################## - distrMut <- read.table("$folderFigure/Overall_mutation_distribution/$filename/$filename-OverallMutationDistribution.txt", header=T)`); - $R->send(q`# Add the count of each category in the legend - distrMut$Legend[[1]] <- paste0(distrMut$Variant_type[[1]], " (", distrMut$Count[[1]], ")") - distrMut$Legend[[2]] <- paste0(distrMut$Variant_type[[2]], " (", distrMut$Count[[2]], ")") - distrMut$Legend[[3]] <- paste0(distrMut$Variant_type[[3]], " (", distrMut$Count[[3]], ")")`); - - $R->send(qq`# Base plot - pie <- ggplot(distrMut, aes(x=factor(""), fill=Legend, weight=Count)) + geom_bar(width=1) + coord_polar(theta="y") + scale_x_discrete("", breaks=NULL) + scale_y_continuous("", breaks=NULL) + labs(fill="") - # Background of the plot entire white - pie <- pie + theme(panel.grid.major = element_line(colour="white"), panel.grid.minor = element_line(colour="white"), panel.background = element_rect(fill="white")) - # Legend on right in 3 rows - pie <- pie + theme(legend.position="bottom") + guides(fill=guide_legend(nrow=3)) - # Change the color and the title of the legend - pie <- pie + scale_fill_brewer("Variant type", palette="Set1") - # Remove all the margins - pie <- pie + theme(plot.margin=unit(c(-1, 0, -1.5, 0), "cm")) - # Save the pie chart for the HTML page (higher resolution) - options(bitmapType='cairo') # Use cairo device as isn't possible to install X11 on the server... - png("$folderFigure/Overall_mutation_distribution/$filename/$filename-OverallMutationDistribution.png", width=700, height=1100, res=300) - print(pie) - dev.off() - - - ########################################## - ## SBS MUTATION DISTRIBUTION ## - ########################################## - distrSBS <- read.delim("$folderFigure/SBS_distribution/$filename/$filename-SBS_distribution.txt") - distrSBS <- data.frame(distrSBS) - bar <- ggplot(distrSBS, aes(x=Mutation_Type, y=Percentage, fill=Mutation_Type)) - bar <- bar + geom_bar(stat="identity", width=0.5) - # Theme classic - bar <- bar + theme_classic() - # Remove the axis legend - bar <- bar + xlab("") - # Set the color of the bars and Changing the labels in the legend - bar <- bar + scale_fill_manual(values=c("blue", "black", "red", "gray", "#00CC33", "pink"), - labels=c("C:G>A:T", "C:G>G:C", "C:G>T:A", "T:A>A:T", "T:A>C:G", "T:A>G:C") - ) - # Remove the label in x axis - bar <- bar + theme(axis.text.x = element_blank()) - # Change the name of the y label - bar <- bar + ylab("Percent") - # Save the plot for the HTML page (higher resolution) - options(bitmapType='cairo') - png("$folderFigure/SBS_distribution/$filename/$filename-SBS_distribution.png", width=1800, height=1500, res=300) - print(bar); - dev.off() - # Save the plot for the report - bar - ggsave("$folder_temp/$filename-SBS_distribution-Report.png") - - - ########################################## - ## IMPACT ON PROTEIN ## - ########################################## - impactProt <- read.table("$folderFigure/Impact_protein_sequence/$filename/$filename-DistributionExoFunc.txt", header=T) - # Custom palette: black, orange, dark green, yellow, light blue, dark blue, darkslateblue, red, purple, pink, light green, turquoise, gray - cb_palette <- c("#000000", "#E69F00", "#006600", "#660000", "#F0E442", "#56B4E9", "#3300FF", "#483D8B", "#FF0000", "#9900CC", "#FF66CC", "#00CC00", "#66FFFF", "#C0C0C0") - pie <- ggplot(impactProt, aes(x=factor(""), fill=AA_Change, weight=Count)) + geom_bar(width=1) + coord_polar(theta="y") + scale_x_discrete("", breaks=NULL)+ scale_y_continuous("", breaks=NULL) + scale_fill_manual(values=cb_palette) - # Background of the plot entire white - pie <- pie + theme(panel.grid.major = element_line(colour="white"), panel.grid.minor = element_line(colour="white"), panel.background = element_rect(fill="white")) - # Legend in two column - pie <- pie + guides(fill=guide_legend(ncol=2)) + theme(legend.position="bottom") - # Remove the legend title - pie <- pie + labs(fill="") - # Save the plot for the HTML page (higher resolution) - options(bitmapType='cairo') - png("$folderFigure/Impact_protein_sequence/$filename/$filename-DistributionExoFunc.png", width=1600, height=1800, res=300) - print(pie) - dev.off() - # Save the plot for the report - pie - ggsave("$folder_temp/$filename-DistributionExoFunc-Report.png") - - - ########################################## - ## STRAND BIAS ## - ########################################## - cb_palette_SB <- c("#0072B2", "#CC0000") - file_sb <- read.table("$folderFigure/Stranded_Analysis/$filename/$filename-StrandBias.txt", header=T); - p_sb <- ggplot(file_sb, aes(x=Alteration, y=Count, fill=Strand)) + theme_classic() + geom_bar(stat="identity", position="dodge") + scale_fill_manual(values=cb_palette_SB) + theme(axis.text.x = element_text(angle=60, hjust=1)) + xlab("") + theme(legend.position="bottom") - # Save the plot for the HTML page (higher resolution) - options(bitmapType='cairo') - png("$folderFigure/Stranded_Analysis/$filename/$filename-StrandBias.png", width=1000, height=1200, res=300) - print(p_sb) - dev.off() - # Save the plot for the report - p_sb - ggsave("$folder_temp/$filename-StrandBias-Report.png") - - - ########################################## - ## HEATMAP SEQUENCE CONTEXT ## - ## GENOMIC STRAND ## - ########################################## - ## COUNT - heatmap_G <- read.table("$folderFigure/Trinucleotide_Sequence_Context/$filename/$filename-HeatmapCount-Genomic.txt", header=T) - # Save the plot for the report - options(bitmapType='cairo') - png(filename="$folder_temp/$filename-HeatmapCount-Genomic-Report.png", bg="transparent", width=240, height=360) - # Heatmap with an absolute scale - heatmap.2(as.matrix(heatmap_G),Rowv=F,Colv=F,col=colorpanel(384,low="yellow",high="red"),dendrogram="none",scale="none",trace="none",key=F,labRow=rownames(as.matrix(heatmap_G)),labCol=colnames(as.matrix(heatmap_G)),lmat=rbind(c(5,1,4),c(3,1,2)), lhei=c(0.75,0.75),lwid=c(0.5,1.5,0.5)) - dev.off() - # Save the plot for the HTML page (higher resolution) - options(bitmapType='cairo') - png(filename="$folderFigure/Trinucleotide_Sequence_Context/$filename/$filename-HeatmapCount-Genomic.png", width=1100, height=1600, res=300) - heatmap.2(as.matrix(heatmap_G),Rowv=F,Colv=F,col=colorpanel(384,low="yellow",high="red"),dendrogram="none",scale="none",trace="none",key=F,labRow=rownames(as.matrix(heatmap_G)),labCol=colnames(as.matrix(heatmap_G)),lmat=rbind(c(5,1,4),c(3,1,2)), lhei=c(0.75,0.75),lwid=c(0.5,1.5,0.5)) - dev.off() - - ## PERCENT - heatmap_G <- read.table("$folderFigure/Trinucleotide_Sequence_Context/$filename/$filename-HeatmapPercent-Genomic.txt", header=T) - # Save the plot for the report - options(bitmapType='cairo') - png(filename="$folder_temp/$filename-HeatmapPercent-Genomic-Report.png",bg="transparent", width=240, height=360) - # Heatmap with an absolute scale - heatmap.2(as.matrix(heatmap_G),Rowv=F,Colv=F,col=colorpanel(384,low="yellow",high="red"),dendrogram="none",scale="none",trace="none",key=F,labRow=rownames(as.matrix(heatmap_G)),labCol=colnames(as.matrix(heatmap_G)),lmat=rbind(c(5,1,4),c(3,1,2)), lhei=c(0.75,0.75),lwid=c(0.5,1.5,0.5)) - dev.off() - # Save the plot for the HTML page (higher resolution) - options(bitmapType='cairo') - png(filename="$folderFigure/Trinucleotide_Sequence_Context/$filename/$filename-HeatmapPercent-Genomic.png", width=1100, height=1600, res=300) - heatmap.2(as.matrix(heatmap_G),Rowv=F,Colv=F,col=colorpanel(384,low="yellow",high="red"),dendrogram="none",scale="none",trace="none",key=F,labRow=rownames(as.matrix(heatmap_G)),labCol=colnames(as.matrix(heatmap_G)),lmat=rbind(c(5,1,4),c(3,1,2)), lhei=c(0.75,0.75),lwid=c(0.5,1.5,0.5)) - dev.off()`); - $R->stopR() ; - - ## Plot the transcriptional strand bias in mutation signature - `Rscript $pathRScriptTxnSB $folderFigure/Stranded_Analysis/$filename/$filename-StrandedSignatureCount.txt $folderFigure/Stranded_Analysis/$filename/$filename-StrandedSignatureCount $folder_temp/$filename-StrandedSignatureCount Count 2>&1`; - `Rscript $pathRScriptTxnSB $folderFigure/Stranded_Analysis/$filename/$filename-StrandedSignaturePercent.txt $folderFigure/Stranded_Analysis/$filename/$filename-StrandedSignaturePercent $folder_temp/$filename-StrandedSignaturePercent Percent 2>&1`; - } - # Write the titles of the different sections of the report - sub WriteBoderSection - { - our ($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext) = @_; - - our ($format_topLeft, $format_topRight, $format_bottomLeft, $format_bottomRight, $format_top, $format_right, $format_bottom, $format_left); - Format_section($wb, \$format_topLeft, \$format_topRight, \$format_bottomLeft, \$format_bottomRight, \$format_top, \$format_right, \$format_bottom, \$format_left); - - TableSBSDistrBySeg(); - TableStrandBiasBySegment(); - CountSBSPerChr(); - ShortTriNtContext(); # 6 mut type - LongTriNtContext(); # 12 mut type - - sub TableSBSDistrBySeg - { - # Top-Left - $ws->write($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, "Table 4. SBS distribution by functional region", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg, 18); # Set the height of the row to 0.25" - # Top - for(my $i=1; $i<=13; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+$i, $format_top); } - # Top-Right - $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+13, $format_topRight); - # Right - $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg+13, $format_right); - # Bottom-left - $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+5, $colStart_SBSdistrBySeg, $format_bottomLeft); - # Left - $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg, $format_left); - } - - sub TableStrandBiasBySegment - { - # Top-Left - $ws->write($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg, "Table 5. Strand bias by functional region", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg+$nb_func+8, 18); # Set the height of the row to 0.25" - # Top - for(my $i=1; $i<=10; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg+$i, $format_top); } - # Top-Right - $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg+11, $format_topRight); - # Right - $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+9, $colStart_SBSdistrBySeg+11, $format_right); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+13, $colStart_SBSdistrBySeg+11, $format_right); - # Left - $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+9, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+10, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+13, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+14, $colStart_SBSdistrBySeg, $format_left); - # Bottom - $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*3)+16, $colStart_SBSdistrBySeg+4, $format_bottom); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*3)+16, $colStart_SBSdistrBySeg+8, $format_bottom); - } - - sub CountSBSPerChr - { - #### Top-Left - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg, "Table 6. SBS distribution per chromosome", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, 18); # Set the height of the row to 0.25" - #### Top - for(my $i=1; $i<8; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg+$i, $format_top); } - #### Top-Right - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg+8, $format_topRight); - #### Right - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+5, $colStart_SBSdistrBySeg+8, $format_right); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+6, $colStart_SBSdistrBySeg+8, $format_right); - - #### Bottom-Right - # Human genome = 24 chromosomes - if($refGenome =~ /hg/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+8, $format_bottomRight); } - # Mouse genome = 21 chromosomes - if($refGenome =~ /mm/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+8, $format_bottomRight); } - # Rat genome = 22 chromosomes - if($refGenome =~ /rn/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+8, $format_bottomRight); } - - #### Bottom - if($refGenome =~ /hg/) - { - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+1, $format_bottom); - for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+$i, $format_bottom); } - } - if($refGenome =~ /mm/) - { - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+1, $format_bottom); - for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+$i, $format_bottom); } - } - if($refGenome =~ /rn/) - { - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+1, $format_bottom); - for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+$i, $format_bottom); } - } - - #### Left - $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+5, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+6, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+7, $colStart_SBSdistrBySeg, $format_left); - - #### Bottom-left - if($refGenome =~ /hg/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg, $format_bottomLeft); } - if($refGenome =~ /mm/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg, $format_bottomLeft); } - if($refGenome =~ /rn/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg, $format_bottomLeft); } - } - - sub ShortTriNtContext - { - my $format_headerSection = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); - $format_headerSection->set_left(2); $format_headerSection->set_left_color('blue'); - - # Top-left - $ws->write(0, $colStart_matrixSeqContext, "Panel 1. Trinucleotide sequence context of SBS on the genomic sequence", $format_topLeft); - # Top - for(my $i=1; $i<=19; $i++) { $ws->write_blank(0, $colStart_matrixSeqContext+$i, $format_top); } - # Top-right - $ws->write_blank(0, $colStart_matrixSeqContext+20, $format_topRight); - # Right - for(my $i=1; $i<=37; $i++) { $ws->write_blank($i, $colStart_matrixSeqContext+20, $format_right); } - # Bottom-right - $ws->write_blank(37, $colStart_matrixSeqContext+20, $format_bottomRight); - # Bottom - for(my $i=1; $i<=19; $i++) { $ws->write_blank(38, $colStart_matrixSeqContext+$i, $format_top); } - # Bottom-left - $ws->write_blank(37, $colStart_matrixSeqContext, $format_bottomLeft); - # Left - $ws->write(1, $colStart_matrixSeqContext, "", $format_left); - for(my $i=3; $i<=36; $i++) { $ws->write_blank($i, $colStart_matrixSeqContext, $format_left); } - } - - sub LongTriNtContext - { - # Top-left - $ws->write($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext, "Panel 2. Stranded analysis of trinucleotide sequence context of SBS", $format_topLeft); - # Top - for(my $i=1; $i<=28; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext+$i, $format_top); } - # Top-right - $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext+29, $format_topRight); - # Right - for(my $i=1; $i<=42; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+$i, $colStart_matrixSeqContext+29, $format_right); } - # Bottom-right - $ws->write_blank(91, $colStart_matrixSeqContext+29, $format_bottomRight); - # Bottom - for(my $i=13; $i<=28; $i++) { $ws->write_blank(92, $colStart_matrixSeqContext+$i, $format_top); } - # Bottom-left - $ws->write_blank(91, $colStart_matrixSeqContext, $format_bottomLeft); - # Left - $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext, $format_left); - $ws->write_blank($rowStart_SBSdistrBySeg+22, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+23, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext, $format_left); - } - } - # Write the header for the six mutation types - sub WriteHeaderSection - { - our ($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext) = @_; - - our ($format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG, $format_TG2, $format_LeftHeader, $format_RightHeader, $format_LeftHeader2); - Format_Header($wb, \$format_CA, \$format_CG, \$format_CT, \$format_TA, \$format_TC, \$format_TG, \$format_TG2, \$format_LeftHeader, \$format_RightHeader, \$format_LeftHeader2); - - our ($format_LeftCA, $format_LeftCG, $format_LeftCT, $format_LeftTA, $format_LeftTC, $format_LeftTG, $format_RightCA, $format_RightCG, $format_RightCT, $format_RightTA, $format_RightTC, $format_RightTG); - Format_HeaderSBSDistrBySegAndFunc($wb, \$format_LeftCA, \$format_LeftCG, \$format_LeftCT, \$format_LeftTA, \$format_LeftTC, \$format_LeftTG, \$format_RightCA, \$format_RightCG, \$format_RightCT, \$format_RightTA, \$format_RightTC, \$format_RightTG); - - our $format_A11Bold = ""; Format_A11Bold($wb, \$format_A11Bold); # Arial 11 bold and center - our $format_A11BoldLeft = ""; Format_A11BoldLeft($wb, \$format_A11BoldLeft); # Arial 11 bold and left - - our ($format_header12CA, $format_header12CG, $format_header12CT, $format_header12TA, $format_header12TC, $format_header12TG); - Format_Header12MutType($wb, \$format_header12CA, \$format_header12CG, \$format_header12CT, \$format_header12TA, \$format_header12TC, \$format_header12TG); - - ## Header for SBS distribution by segment - HeaderMutTypeSBSDistrBySeg(); - - ## Header for strand bias by function - $ws->set_column($colStart_SBSdistrBySeg+5, $colStart_SBSdistrBySeg+5, 11); - - my $row = $rowStart_SBSdistrBySeg+$nb_func+10; my $col = $colStart_SBSdistrBySeg; - $ws->write($row, $col+1, ' ', $format_CA); $ws->write($row, $col+2, "C>A", $format_CA); $ws->write($row, $col+3, ' ', $format_CA); - $ws->write($row, $col+5, ' ', $format_CG); $ws->write($row, $col+6, "C>G", $format_CG); $ws->write($row, $col+7, ' ', $format_CG); - $ws->write($row, $col+9, ' ', $format_CT); $ws->write($row, $col+10, "C>T", $format_CT); $ws->write($row, $col+11, ' ', $format_RightCT); - - $row = $rowStart_SBSdistrBySeg+($nb_func*2)+14; - $ws->write($row, $col+1, ' ', $format_TA); $ws->write($row, $col+2, "T>A", $format_TA); $ws->write($row, $col+3, ' ', $format_TA); - $ws->write($row, $col+5, ' ', $format_TC); $ws->write($row, $col+6, "T>C", $format_TC); $ws->write($row, $col+7, ' ', $format_TC); - $ws->write($row, $col+9, ' ', $format_TG2); $ws->write($row, $col+10, "T>G", $format_TG2); $ws->write($row, $col+11, ' ', $format_RightTG); - - $ws->set_row($rowStart_SBSdistrBySeg+$nb_func+11, 18); $ws->set_row($rowStart_SBSdistrBySeg+($nb_func*2)+15, 18); - $ws->set_column($colStart_SBSdistrBySeg+5, $colStart_SBSdistrBySeg+5, 13); $ws->set_column($colStart_SBSdistrBySeg+9, $colStart_SBSdistrBySeg+9, 13); - - for(my $i=$rowStart_SBSdistrBySeg+$nb_func+10; $i<=$rowStart_SBSdistrBySeg+($nb_func*2)+14; $i+=$nb_func+4) - { - $ws->write($i+1, $colStart_SBSdistrBySeg, 'Segment', $format_LeftHeader); $ws -> write($i+1, $colStart_SBSdistrBySeg+1, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+2, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+3, 'Tr', $format_A11Bold); - $ws -> write($i+1, $colStart_SBSdistrBySeg+5, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+6, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+7, 'Tr', $format_A11Bold); - $ws -> write($i+1, $colStart_SBSdistrBySeg+9, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+10, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+11, 'Tr', $format_RightHeader); - } - - - ## Header for Counts of SBS per chromosome and mutation type - HeaderCountSBSPerChr(); - - ## Header for the short sequence context - HeaderShortTriNtContext(); - - ## Header for the 12 mutation types with the sequence context (coding strand) - HeaderLongTriNtContext(); - - sub HeaderMutTypeSBSDistrBySeg - { - $ws->set_row($rowStart_SBSdistrBySeg+2, 18); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+2, "C:G>A:T", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+3, $format_CA); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+4, "C:G>G:C", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+5, $format_CG); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+6, "C:G>T:A", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+7, $format_CT); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+8, "T:A>A:T", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+9, $format_TA); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+10, "T:A>C:G", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+11, $format_TC); - $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+12, "T:A>G:C", $format_TG); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+13, $format_TG); - - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg, "Segment", $format_LeftHeader); $ws->set_column($colStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, 13); $ws->set_row($rowStart_SBSdistrBySeg+3, 18); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+1, "Total SBS", $format_A11Bold); $ws->set_column($colStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg+1, 11); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+2, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+3, "#", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+4, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+5, "#", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+6, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+7, "#", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+8, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+9, "#", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+10, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+11, "#", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+12, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, 13, "#", $format_RightHeader); - } - - sub HeaderCountSBSPerChr - { - $ws->set_column(3,3, 10); $ws->set_column(4,4, 10); - $ws->set_row($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, 18); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+7, $colStart_SBSdistrBySeg+1, "Pearson", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg, "Chr", $format_LeftHeader); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+1, "Size", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+2, "All SBS", $format_A11Bold); - - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+3, "C:G>A:T", $format_CA); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+4, "C:G>G:C", $format_CG); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+5, "C:G>T:A", $format_CT); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+6, "T:A>A:T", $format_TA); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+7, "T:A>C:G", $format_TC); - $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+8, "T:A>G:C", $format_TG); - } - - sub HeaderShortTriNtContext - { - ### GENOMIC STRAND - $ws->write(2, $colStart_matrixSeqContext, 'Count matrix', $format_LeftHeader2); - $ws->write(3, $colStart_matrixSeqContext+4, 'C>A', $format_CA); $ws->write(3, $colStart_matrixSeqContext+5, 'C>G', $format_CG); $ws->write(3, $colStart_matrixSeqContext+6, 'C>T', $format_CT); $ws->write(3, $colStart_matrixSeqContext+7, 'T>A', $format_TA); $ws->write(3, $colStart_matrixSeqContext+8, 'T>C', $format_TC); $ws->write(3, $colStart_matrixSeqContext+9, 'T>G', $format_TG2); - - $ws->write(2, $colStart_matrixSeqContext+11, 'Frequency matrix', $format_A11BoldLeft); - $ws->write(3, $colStart_matrixSeqContext+14, 'C>A', $format_CA); $ws->write(3, $colStart_matrixSeqContext+15, 'C>G', $format_CG); $ws->write(3, $colStart_matrixSeqContext+16, 'C>T', $format_CT); $ws->write(3, $colStart_matrixSeqContext+17, 'T>A', $format_TA); $ws->write(3, $colStart_matrixSeqContext+18, 'T>C', $format_TC); $ws->write(3, $colStart_matrixSeqContext+19, 'T>G', $format_TG2); - - ### sequence context with a bar graph - $ws->write(25, $colStart_matrixSeqContext+10, "Mutation spectra frequency", $format_A11Bold); - } - - sub HeaderLongTriNtContext - { - $ws->set_row($rowStart_SBSdistrBySeg+3, 15); $ws->set_row($rowStart_SBSdistrBySeg+4, 15); $ws->set_row($rowStart_SBSdistrBySeg+5, 15); - $ws->write($rowStart_SBSdistrBySeg+3, $colStart_matrixSeqContext, "Count matrix", $format_LeftHeader2); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+1, "C>A", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+2, $format_CA); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+1, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+2, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+3, "C>G", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+4, $format_CG); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+3, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+4, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+5, "C>T", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+6, $format_CT); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+5, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+6, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+7, "T>A", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+8, $format_TA); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+7, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+8, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+9, "T>C", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+10, $format_TC); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+9, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+10, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+11, "T>G", $format_TG2); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+12, $format_TG2); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+11, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+12, "Tr", $format_A11Bold); - - - $ws->set_row($rowStart_SBSdistrBySeg+24, 15); $ws->set_row($rowStart_SBSdistrBySeg+25, 15); $ws->set_row($rowStart_SBSdistrBySeg+26, 15); - $ws->write($rowStart_SBSdistrBySeg+24, $colStart_matrixSeqContext, "Frequency matrix", $format_LeftHeader2); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+1, "C>A", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+2, $format_CA); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+1, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+2, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+3, "C>G", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+4, $format_CG); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+3, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+4, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+5, "C>T", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+6, $format_CT); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+5, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+6, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+7, "T>A", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+8, $format_TA); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+7, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+8, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+9, "T>C", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+10, $format_TC); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+9, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+10, "Tr", $format_A11Bold); - $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+11, "T>G", $format_TG2); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+12, $format_TG2); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+11, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+12, "Tr", $format_A11Bold); - } - } - # Create logo for representing the sequence context with n bases - sub CreateLogo - { - my ($refH_file, $folderWebLogo) = @_; - - my $folderSample = ""; - - foreach my $k_file (sort keys $refH_file) - { - $folderSample = "$folderWebLogo/$k_file"; - if(!-e $folderSample) { mkdir($folderSample) or die "Can't create the directory $folderSample\n"; } - - my $test_lengthSeqContext = 0; - - foreach my $k_mutation (sort keys $refH_file->{$k_file}{'WebLogo3'}) - { - open(WEBLOGO, ">", "$folderSample/$k_file-$k_mutation.fa") or die "$!: $folderSample/$k_file-$k_mutation.fa\n"; - foreach (@{$refH_file->{$k_file}{'WebLogo3'}{$k_mutation}}) - { - print WEBLOGO ">$k_file\n$_\n"; - - if(length($_) < 10) { $test_lengthSeqContext = 0; } - else { $test_lengthSeqContext = 1; } - } - close WEBLOGO; - } - - ## Generate the logo - foreach my $fastaFile (`ls $folderSample/*.fa`) - { - chomp($fastaFile); - my ($filename, $directories, $suffix) = fileparse("$folderSample/$fastaFile", qr/\.[^.]*/); - - $filename =~ /(.+)\-/; - my $title = $1; - - ## Test if there is fasta sequence for the mutation type - my $nbLigne_temp = `wc -l $fastaFile`; - my @nbLigne = split(" ", $nbLigne_temp); - - if($nbLigne[0] == 0) { print "WARNING: No sequence for $filename\n"; next; } - - # When length sequence context is lower than 10 the image is to small for adding a title - if($test_lengthSeqContext == 1) { system("weblogo -c classic -F png -U probability --title $title < $fastaFile > $folderSample/$filename-Probability.png"); } - else { system("weblogo -c classic -F png -U probability < $fastaFile > $folderSample/$filename-Probability.png"); } - } - } - } - - - # Define the format of the worksheet: Arial font size=10 - sub Format_A10 - { - my ($wb, $format) = @_; - $$format = $wb->add_format(font=>'Arial', size=>10); $$format->set_align('center'); - } - # Define the format of the worksheet: Arial font size=11 bold and center - sub Format_A11Bold - { - my ($wb, $format) = @_; - $$format = $wb->add_format(font=>'Arial', size=>11, bold=>1); $$format->set_align('center'); - } - # Define the format of the worksheet: Arial font size=10 italic red and center - sub Format_A10ItalicRed - { - my ($wb, $format) = @_; - $$format = $wb->add_format(font=>'Arial', size=>10, italic=>1, color => 'red'); $$format->set_align('center'); - } - # Defile the format of the worksheet: Arialt font size=11 bold and left - sub Format_A11BoldLeft - { - my ($wb, $format) = @_; - $$format = $wb->add_format(valign =>'left', font=>'Arial', size=>11, bold=>1); - } - # Defile the format of the worksheet: Arialt font size=10 bold and left - sub Format_A10BoldLeft - { - my ($wb, $format) = @_; - $$format = $wb->add_format(valign =>'left', font=>'Arial', size=>10, bold=>1); - } - # Define the format of the border of the section (for delimiting the different section of the report) - sub Format_section - { - my ($wb, $format_topLeft, $format_topRight, $format_bottomLeft, $format_bottomRight, $format_top, $format_right, $format_bottom, $format_left) = @_; - - $$format_topLeft = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); - $$format_topLeft->set_top(2); $$format_topLeft->set_top_color('blue'); - $$format_topLeft->set_left(2); $$format_topLeft->set_left_color('blue'); - - $$format_topRight = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); - $$format_topRight->set_top(2); $$format_topRight->set_top_color('blue'); - $$format_topRight->set_right(2); $$format_topRight->set_right_color('blue'); - - $$format_bottomLeft = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); - $$format_bottomLeft->set_bottom(2); $$format_bottomLeft->set_bottom_color('blue'); - $$format_bottomLeft->set_left(2); $$format_bottomLeft->set_left_color('blue'); - - $$format_bottomRight = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); - $$format_bottomRight->set_bottom(2); $$format_bottomRight->set_bottom_color('blue'); - $$format_bottomRight->set_right(2); $$format_bottomRight->set_right_color('blue'); - - $$format_top = $wb->add_format(); $$format_top->set_top(2); $$format_top->set_top_color('blue'); - $$format_right = $wb->add_format(); $$format_right->set_right(2); $$format_right->set_right_color('blue'); - $$format_bottom = $wb->add_format(); $$format_bottom->set_bottom(2); $$format_bottom->set_bottom_color('blue'); - $$format_left = $wb->add_format(); $$format_left->set_left(2); $$format_left->set_left_color('blue'); - } - # Define the header - sub Format_Header - { - my ($wb, $format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG, $format_TG2, $format_LeftHeader, $format_RightHeader, $format_LeftHeader2) = @_; - - my ($blue, $black, $red, $gray, $green, $pink); - Color($wb, \$blue, \$black, \$red, \$gray, \$green, \$pink); - - my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); - BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); - - - $$format_CA = $wb->add_format(bg_color => $blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CA->set_align('center'); $$format_CA->set_center_across(); - $$format_CG = $wb->add_format(bg_color => $black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CG->set_align('center'); $$format_CG->set_center_across(); - $$format_CT = $wb->add_format(bg_color => $red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CT->set_align('center'); $$format_CT->set_center_across(); - $$format_TA = $wb->add_format(bg_color => $gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TA->set_align('center'); $$format_TA->set_center_across(); - $$format_TC = $wb->add_format(bg_color => $green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TC->set_align('center'); $$format_TC->set_center_across(); - $$format_TG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG->set_align('center'); $$format_TG->set_center_across(); - $$format_TG->set_right(2); $$format_TG->set_right_color('blue'); - - $$format_TG2 = $wb->add_format(bg_color => $pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG2->set_align('center'); $$format_TG2->set_center_across(); - - $$format_LeftHeader = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_LeftHeader->set_align('center'); $$format_LeftHeader->set_left(2); $$format_LeftHeader->set_left_color('blue'); - $$format_LeftHeader2 = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_LeftHeader2->set_left(2); $$format_LeftHeader2->set_left_color('blue'); - $$format_RightHeader = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_RightHeader->set_align('center'); $$format_RightHeader->set_right(2); $$format_RightHeader->set_right_color('blue'); - } - # Define the mutation type header for the Strand bias by segment - sub Format_HeaderSBSDistrBySegAndFunc - { - my ($wb, $format_LeftCA, $format_LeftCG, $format_LeftCT, $format_LeftTA, $format_LeftTC, $format_LeftTG, $format_RightCA, $format_RightCG, $format_RightCT, $format_RightTA, $format_RightTC, $format_RightTG) = @_; - - my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); - BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); - - $$format_LeftCA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCA->set_align('center'); $$format_LeftCA->set_left(2); $$format_LeftCA->set_left_color('blue'); - $$format_LeftCG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCG->set_align('center'); $$format_LeftCG->set_left(2); $$format_LeftCG->set_left_color('blue'); - $$format_LeftCT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCT->set_align('center'); $$format_LeftCT->set_left(2); $$format_LeftCT->set_left_color('blue'); - $$format_LeftTA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTA->set_align('center'); $$format_LeftTA->set_left(2); $$format_LeftTA->set_left_color('blue'); - $$format_LeftTC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTC->set_align('center'); $$format_LeftTC->set_left(2); $$format_LeftTC->set_left_color('blue'); - $$format_LeftTG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTG->set_align('center'); $$format_LeftTG->set_left(2); $$format_LeftTG->set_left_color('blue'); - - - $$format_RightCA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCA->set_align('center'); $$format_RightCA->set_right(2); $$format_RightCA->set_right_color('blue'); - $$format_RightCG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCG->set_align('center'); $$format_RightCG->set_right(2); $$format_RightCG->set_right_color('blue'); - $$format_RightCT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCT->set_align('center'); $$format_RightCT->set_right(2); $$format_RightCT->set_right_color('blue'); - $$format_RightTA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTA->set_align('center'); $$format_RightTA->set_right(2); $$format_RightTA->set_right_color('blue'); - $$format_RightTC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTC->set_align('center'); $$format_RightTC->set_right(2); $$format_RightTC->set_right_color('blue'); - $$format_RightTG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTG->set_align('center'); $$format_RightTG->set_right(2); $$format_RightTG->set_right_color('blue'); - } - # Define the mutation type header for the trinucleotide sequence context on the coding strand - sub Format_Header12MutType - { - my ($wb, $format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG) = @_; - - my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); - BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); - - $$format_CA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CA->set_align('center'); - $$format_CG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CG->set_align('center'); - $$format_CT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CT->set_align('center'); - $$format_TA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TA->set_align('center'); - $$format_TC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TC->set_align('center'); - $$format_TG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG->set_align('center'); - } - # Define the format for the text that needs a section border - sub Format_TextSection - { - my ($wb, $formatT_left, $formatT_right, $formatT_bottomRight, $formatT_bottomLeft, $formatT_bottom, $formatT_bottomHeader, $formatT_bottomRightHeader, $formatT_bottomHeader2, $formatT_rightHeader) = @_; - - $$formatT_left = $wb->add_format(valign=>'center', font=>'Arial', size=>10); - $$formatT_left->set_left(2); $$formatT_left->set_left_color('blue'); - - $$formatT_right = $wb->add_format(valign=>'center', font=>'Arial', size=>10); - $$formatT_right->set_right(2); $$formatT_right->set_right_color('blue'); - - $$formatT_bottomRight = $wb->add_format(valign=>'center', font=>'Arial', size=>10); - $$formatT_bottomRight->set_bottom(2); $$formatT_bottomRight->set_bottom_color('blue'); - $$formatT_bottomRight->set_right(2); $$formatT_bottomRight->set_right_color('blue'); - - $$formatT_bottomLeft = $wb->add_format(valign=>'center', font=>'Arial', size=>10); - $$formatT_bottomLeft->set_bottom(2); $$formatT_bottomLeft->set_bottom_color('blue'); - $$formatT_bottomLeft->set_left(2); $$formatT_bottomLeft->set_left_color('blue'); - - $$formatT_bottom = $wb->add_format(valign=>'center', font=>'Arial', size=>10); - $$formatT_bottom->set_bottom(2); $$formatT_bottom->set_bottom_color('blue'); - - my $bgColor_totallighGray = $wb->set_custom_color(54, 230, 230, 230); - $$formatT_bottomHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomHeader->set_align('center'); - $$formatT_bottomHeader->set_bottom(2); $$formatT_bottomHeader->set_bottom_color('blue'); - - $$formatT_bottomRightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomRightHeader->set_align('center'); - $$formatT_bottomRightHeader->set_bottom(2); $$formatT_bottomRightHeader->set_bottom_color('blue'); - $$formatT_bottomRightHeader->set_right(2); $$formatT_bottomRightHeader->set_right_color('blue'); - - $$formatT_bottomHeader2 = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomHeader2->set_align('center'); - - $$formatT_rightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_rightHeader->set_align('center'); - $$formatT_rightHeader->set_right(2); $$formatT_rightHeader->set_right_color('blue'); - } - # Define the format for the graphs titles - sub Format_GraphTitle - { - my ($wb, $formatT_graphTitle) = @_; - - $$formatT_graphTitle = $wb->add_format(font=>'Arial', size=>12, bold=>1); - } - # Define the format of the border of the tables - sub Format_Table - { - my ($wb, $table_topleft, $table_topRight, $table_bottomleft, $table_bottomRight, $table_top, $table_right, $table_bottom, $table_bottomItalicRed, $table_left, $table_bottomrightHeader, $table_left2, $table_middleHeader, $table_middleHeader2) = @_; - - $$table_topleft = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_topleft->set_top(1); $$table_topleft->set_left(1); - $$table_topRight = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_topRight->set_top(1); $$table_topRight->set_right(1); - $$table_bottomleft = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_bottomleft->set_bottom(1); $$table_bottomleft->set_left(1); - $$table_bottomRight = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_bottomRight->set_bottom(1); $$table_bottomRight->set_right(1); - - $$table_top = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_top->set_top(1); - $$table_right = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_right->set_right(1); - $$table_bottom = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_bottom->set_bottom(1); - $$table_bottomItalicRed = $wb->add_format(valign=>'center', font=>'Arial', size=>10, italic=>1, color => 'red'); $$table_bottomItalicRed->set_bottom(1); - $$table_left = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_left->set_left(1); - - my $bgColor_totallighGray = $wb->set_custom_color(54, 230, 230, 230); - $$table_bottomrightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); $$table_bottomrightHeader->set_bottom(1); $$table_bottomrightHeader->set_right(1); - - $$table_left2 = $wb->add_format(valign=>'left', font=>'Arial', size=>10); $$table_left2->set_left(1); - - $$table_middleHeader = $wb->add_format(valign=>'center', bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); - $$table_middleHeader2 = $wb->add_format(valign=>'center', bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); $$table_middleHeader2->set_bottom(1); - } - - # Define the color - sub Color - { - my ($wb, $blue, $black, $red, $gray, $green, $pink) = @_; - - $$blue = $wb->set_custom_color(40, 0, 0, 204);# C:G>A:T in blue - $$black = $wb->set_custom_color(41, 0, 0, 0);# C:G>G:C in black - $$red = $wb->set_custom_color(42, 255, 0, 0);# C:G>T:A in red - $$gray = $wb->set_custom_color(43, 205, 205, 205); # T:A>A:T in light gray - $$green = $wb->set_custom_color(44, 0, 204, 51);# T:A>C:G in green - $$pink = $wb->set_custom_color(45, 255, 192, 203);# T:A>G:C in pink - } - sub BackgroundColor - { - my ($wb, $bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink) = @_; - - $$bgColor_blue = $wb->set_custom_color(48, 0, 0, 204); - $$bgColor_black = $wb->set_custom_color(49, 0, 0, 0); - $$bgColor_red = $wb->set_custom_color(50, 255, 0, 0); - $$bgColor_gray = $wb->set_custom_color(51, 205, 205, 205); - $$bgColor_green = $wb->set_custom_color(52, 0, 204, 51); - $$bgColor_pink = $wb->set_custom_color(53, 255, 192, 203); - } -} - - -sub recoverNumCol -{ - my ($input, $name_of_column) = @_; - - open(F1,$input) or die "recoverNumCol: $!: $input\n"; - # For having the name of the columns - my $search_header = <F1>; $search_header =~ s/[\r\n]+$//; my @tab_search_header = split("\t",$search_header); - close F1; - # The number of the column - my $name_of_column_NB = "toto"; - for(my $i=0; $i<=$#tab_search_header; $i++) - { - if($tab_search_header[$i] eq $name_of_column) { $name_of_column_NB = $i; last; } - } - if($name_of_column_NB eq "toto") { print STDERR "Error recoverNumCol(): the column named $name_of_column doesn't exits in the input file $input!!!!!\n"; exit; } - else { return $name_of_column_NB; } -} - - - - -=head1 NAME - -mutSpec-Stat - -=head1 SYNOPSIS - - mutSpecstat.pl [arguments] <query-file> - - <query-file> can be a folder with multiple VCF or a single VCF - - Arguments: - -h, --help print help message - -m, --man print complete documentation - -v, --verbose use verbose output - --refGenome the reference genome to use (human, mouse or rat genomes) - -o, --outfile <string> output directory for the result. If none is specify the result will be write in the same directory as the input file - -temp --pathTemporary <string> the path for saving the temporary files - --pathSeqRefGenome the path to the fasta reference sequences - --poolData generate the pool of all the samples (optional) - --reportSample generate a report for each sample (optional) - - -Function: automatically run a pipeline and calculate various statistics on mutations - - Example: mutSpecstat.pl --refGenome hg19 --outfile output_directory --temp path_to_temporary_directory --pathRscript path_to_R_scripts --pathSeqRefGenome path_fasta_ref_seq --poolData --reportSample input - - Version: 04-2016 (April 2016) - - -=head1 OPTIONS - -=over 8 - -=item B<--help> - -print a brief usage message and detailed explanation of options. - -=item B<--man> - -print the complete manual of the program. - -=item B<--verbose> - -use verbose output. - -=item B<--refGenome> - -the reference genome to use, could be human, mouse or rat genomes. - -=item B<--outfile> - -the directory of output file names. If it is nor specify the same directory as the input file is used. - -=item B<--pathTemporary> - -the path for saving temporary files generated by the script. -If any is specify a temporary folder is created in the same directory where the script is running. -Deleted when the script is finish - -=item B<--pathSeqRefGenome> - -The path to the fasta reference sequences - -=item B<--poolData only for the report> - -calculate the statistics on the pool of all the data pass in input - -=item B<--reportSample only for the report> - -generate a report for each samples - -=head1 DESCRIPTION - -mutSpecstat is a perl script for calculated various statistics on mutations -An Excel report containing the mutation type distribution per functional region, the strand bias and the sequence context on genomic and coding sequence is created. -The different statistics are illustrated using ggplot2. - -=cut +#!/usr/bin/env perl + +#-----------------------------------# +# Author: Maude # +# Script: mutspecStat.pl # +# Last update: 09/02/17 # +#-----------------------------------# + +use strict; +use warnings; +use Getopt::Long; +use Pod::Usage; +use File::Basename; # my ($filename, $directories, $suffix) = fileparse($file, qr/\.[^.]*/); +use File::Path; +use Spreadsheet::WriteExcel; + +our ($verbose, $man, $help) = (0, 0, 0); # Parse options and print usage if there is a syntax error, or if usage was explicitly requested. +our ($refGenome, $output, $folder_temp, $path_R_Scripts, $path_SeqrefGenome) = ("empty", "empty", "empty", "empty", "empty", "empty"); # The reference genome to use; The path for saving the result; The path for saving the temporary files; The path to R scripts; The path to the fasta reference sequences +our ($poolData, $oneReportPerSample) = (2, 2); # If a folder is pass as input file pool all the data and generate the report on the pool and for each samples; # Generate one report for each samples + + +GetOptions('verbose|v'=>\$verbose, 'help|h'=>\$help, 'man|m'=>\$man, 'refGenome=s'=>\$refGenome, 'outfile|o=s' => \$output, 'temp=s' => \$folder_temp, 'pathRscript=s' => \$path_R_Scripts, 'pathSeqRefGenome=s' => \$path_SeqrefGenome, 'poolData' => \$poolData, 'reportSample' => \$oneReportPerSample) or pod2usage(2); + +our ($input) = @ARGV; + +pod2usage(-verbose=>1, -exitval=>1, -output=>\*STDERR) if ($help); +pod2usage(-verbose=>2, -exitval=>1, -output=>\*STDERR) if ($man); +pod2usage(-verbose=>0, -exitval=>1, -output=>\*STDERR) if(@ARGV == 0); # No argument is pass to the command line print the usage of the script +pod2usage(-verbose=>0, -exitval=>1, -output=>\*STDERR) if(@ARGV == 2); # Only one argument is expected to be pass to @ARGV (the input) + + + + +####### The input must be a folder with one or several annotated files +if(!-d $input) +{ + print STDERR "Error: The input must be a Dataset List\n"; + print STDERR "Even for 1 file, please create a Dataset List\n"; + exit; +} + + +###################################################################################################################################################### +# GLOBAL VARIABLES # +###################################################################################################################################################### +# Recover the current path +our $pwd = `pwd`; +chomp($pwd); + + +# Path to R scripts +our $pathRscriptChi2test = "$path_R_Scripts/R/chi2test_MutSpecStat_Galaxy.r"; +our $pathRScriptFigs = "$path_R_Scripts/R/figs_MutSpecStat_Galaxy.r"; +our $pathRScriptTxnSB = "$path_R_Scripts/R/transciptionalStrandBias.r"; +our $pathRScriptMutSpectrum = "$path_R_Scripts/R/mutationSpectra_Galaxy.r"; + + +# The path for saving the files with enough mutations for calculating the statistics; +our $folderCheckedForStat = "$pwd/folder_checked"; +if(!-e $folderCheckedForStat) { mkdir($folderCheckedForStat) or die "$!: $folderCheckedForStat\n"; } + +# Output dir with all the results +our $folderMutAnalysis = ""; + +# Hash table with the length of each chromosomes +our %chromosomes; +# Define the name of the column containing the chromosome, start, ref and alt alleles (based on Annovar output) +our ($chr_name, $start_name, $ref_name, $alt_name) = qw(Chr Start Ref Alt); +# Annovar annotation used +our $func_name = "Func.refGene"; +our $exonicFunc_name = "ExonicFunc.refGene"; +our $strand_name = "Strand"; +our $context_name = "context"; +# Font formats +our ($format_A10, $format_A10Boldleft, $format_A10ItalicRed) = ("", "", ""); +our ($formatT_left, $formatT_right, $formatT_bottomRight, $formatT_bottomLeft, $formatT_bottom, $formatT_bottomHeader, $formatT_bottomRightHeader, $formatT_bottomHeader2, $formatT_rightHeader); +our ($formatT_graphTitle); +our ($table_topleft, $table_topRight, $table_bottomleft, $table_bottomRight, $table_top, $table_right, $table_bottom, $table_bottomItalicRed, $table_left, $table_bottomrightHeader, $table_left2, $table_middleHeader, $table_middleHeader2); +# Hash table with the result of chi2 test for the strand bias +our %h_chi2 = (); +# For NMF input +our %h_inputNMF = (); + + +###################################################################################################################################################### +# MAIN # +###################################################################################################################################################### +# Check the presence of the flags and create the output and temp directories +CheckFlags(); + +# First check if the files are annotated or not. +# If the files are annotated check there is enough mutations for generating the statistics, otherwise remove the samples from the analysis +checkVariants(); + +# Retrieve chromosomes length +checkChrDir(); + +# Calculate the statistics and generate the report +ReportMutDist(); + +# Remove the temporary directory +rmtree($folder_temp); +rmtree($folderCheckedForStat); + + +###################################################################################################################################################### +# FUNCTIONS # +###################################################################################################################################################### + +# Check the presence of the flags and create the output and temp directories +sub CheckFlags +{ + # Check the reference genome + if($refGenome eq "empty") + { + print STDERR "Missing flag !\n"; + print STDERR "You forget to specify the name for the reference genome!!!\nPlease specify it with the flag --refGenome\n"; + exit; + } + + # If no output is specified write the result as the same place as the input file + if($output eq "empty") + { + # The input is a folder with one or more annotated files + my $directory = dirname( $input ); + + $folderMutAnalysis = "$directory/Mutational_Analysis"; + if(!-e $folderMutAnalysis) { mkdir($folderMutAnalysis) or die "$!: $folderMutAnalysis\n"; } + } + else + { + if(!-e $output) { mkdir($output) or die "$!: $output\n"; } + + $folderMutAnalysis = "$output/Mutational_Analysis"; + if(!-e $folderMutAnalysis) { mkdir($folderMutAnalysis) or die "$!: $folderMutAnalysis\n"; } + } + + # If no temp folder is specified write the result in the current path + my ($filename, $directories, $suffix) = fileparse($input, qr/\.[^.]*/); + if($folder_temp eq "empty") { $folder_temp = "$pwd/TEMP_MutationalAnalysis_$filename"; } + if(!-e $folder_temp) { mkdir($folder_temp) or die "$!: $folder_temp\n"; } + + # Check the path to the R scripts + if($path_R_Scripts eq "empty") + { + print STDERR "Missing flag !\n"; + print STDERR "You forget to specify the path for the R scripts!!!\nPlease specify it with the flag --pathRscript\n"; + exit; + } + + + foreach my $file (`ls $input/*`) + { + chomp($file); + + ## Verify the name of file, must be <= 31 chars for the sheet name + my ($filename, $directories, $suffix) = fileparse($file, qr/\.[^.]*/); + + if(length($filename) > 31) + { + print STDERR "Error: The filename of: $file\nMust be <= 31 chars\nPlease modify it before running the script\n"; + exit; + } + } +} + +# Check input file(s) +sub checkVariants +{ + # Count the number of file(s) with enough mutations (at least 1 with a strand orientation) + my $timerFile = 0; + my @listRemovedFile = (); + + foreach my $file (`ls $input/*`) + { + chomp($file); + + ### Check if the file is annotated + my $testAnnotation = "toto"; + $testAnnotation = `grep 'Func.refGene' $file`; + + if($testAnnotation eq "toto") + { + print STDERR "Error: The input file you specify is not annotated!\nThe file concerned is: $file !!!!\nPlease first annotate your file before trying to generate the report on mutation spectra\n"; + exit; + } + else + { + ### check if there is at least 1 mutation with a strand info + my $strand_value = recoverNumCol($file, "Strand"); + my $nbSBScoding = 0; + + open(F1, $file) or die "$!: $file\n"; + my $header = <F1>; + while(<F1>) + { + $_ =~ s/[\r\n]+$//; + my @tab = split("\t", $_); + + if($tab[$strand_value] ne "NA") + { + $nbSBScoding++; + } + } + close F1; + + if($nbSBScoding != 0) + { + $timerFile++; + `cp $file $folderCheckedForStat/`; + } + else + { + print STDOUT "\n\nWarning: There is no variant to compute statistics for $file\n\n"; + push(@listRemovedFile, $file); + } + } + } + + if($timerFile == 0) + { + print STDERR "\n\nError: No variants to compute statistics for:\n"; + + foreach (@listRemovedFile) + { + print STDERR $_."\n"; + } + exit; + } +} + +# Retrieve chromosomes length +sub checkChrDir +{ + my @files = `ls $path_SeqrefGenome/$refGenome"_seq"/*.fa`; + foreach my $file (@files) + { + if ($file !~ /chr(\d+|x|y)\.fa/i){next;} + open(FILE,$file); + <FILE>; + my $seq=""; + while (<FILE>){ chomp; $seq.=$_;} + $file =~ /chr(.*)\.fa/; + $chromosomes{"chr".$1}=length($seq); + } +} + +# Calculate the statistics and generate the report +sub ReportMutDist +{ + print STDOUT "-----------------------------------------------------------------\n"; + print STDOUT "-----------------Report Mutational Analysis----------------------\n"; + print STDOUT "-----------------------------------------------------------------\n"; + + my $folderFigure = "$folderMutAnalysis/Figures"; + if(-e $folderFigure) { rmtree($folderFigure); mkdir($folderFigure) or die "Can't create the directory $folderFigure\n"; } + else { mkdir($folderFigure) or die "Can't create the directory $folderFigure\n"; } + my $folderChi2 = "$folderFigure/Chi2"; + if(!-e $folderChi2) { mkdir($folderChi2) or die "Can't create the directory $folderChi2\n"; } + my $folderWebLogo = "$folderFigure/WebLogo"; + if(!-e $folderWebLogo) { mkdir($folderWebLogo) or die "Can't create the directory $folderWebLogo\n"; } + my $folderNMF = "$folderFigure/Input_NMF"; + if(!-e $folderNMF) { mkdir($folderNMF) or die "Can't create the directory $folderNMF\n"; } + + + ################################################################################################ + ### Calculates all the statistics ### + ################################################################################################ + + ########### Recover the functional region for all the samples. Allows to thave the same annotations for the pie chart "Impact on protein sequence" + my @tab_func = recoverAnnovarAnnotation($func_name); + if(@tab_func == 0) + { + print STDERR "Error: the table for the functional region is empty!!!!! check $folderCheckedForStat\n$func_name\n"; + exit; + } + + ############ Calculate the different statistics present in the report + my %h_file = (); + CalculateStatistics(\%h_file, \@tab_func); + + ############ Calculate the chi2 for the strand bias + CalculateChi2(\%h_file, $folderChi2); + + ############ Write the different statistics present in the report + WriteStatistics(\%h_file, $#tab_func, $folderFigure, $folderChi2, $folderNMF); + + ############ Create logo for studying the 10 flanking bases of the mutation + CreateLogo(\%h_file, $folderWebLogo); +} + + +# Calculate the different statistics present in the report +sub CalculateStatistics +{ + my ($refH_file, $refT_func) = @_; + + our ($chr_value, $start_value, $ref_value, $alt_value, $func_value, $exonicFunc_value, $strand_value, $contextSeq_value) = ("", "", "", "", "", "", "", "", "", ""); + + # Generate the pool of all the data + if($poolData == 1) + { + my @listFile = `ls $folderCheckedForStat`; + + # For keeping the header only one time + chomp($listFile[0]); + system("cp $folderCheckedForStat/$listFile[0] $folderCheckedForStat/Pool_Data.txt"); + + open(OUT, ">>", "$folderCheckedForStat/Pool_Data.txt") or die "$!: $folderCheckedForStat/Pool_Data.txt\n"; + + for(my $i=1; $i<=$#listFile; $i++) + { + chomp($listFile[$i]); + open(F1, "$folderCheckedForStat/$listFile[$i]") or die "$!: $folderCheckedForStat/$listFile[$i]\n"; + my $header = <F1>; + while(<F1>) { print OUT $_; } + close F1; + } + close OUT; + } + + foreach my $file (`ls $folderCheckedForStat/*`) + { + chomp($file); + ############ Recover the number of the columns of interest + $chr_value = recoverNumCol($file, $chr_name); + $start_value = recoverNumCol($file, $start_name); + $ref_value = recoverNumCol($file, $ref_name); + $alt_value = recoverNumCol($file, $alt_name); + $func_value = recoverNumCol($file, $func_name); + $exonicFunc_value = recoverNumCol($file, $exonicFunc_name); + $strand_value = recoverNumCol($file, $strand_name); + $contextSeq_value = recoverNumCol($file, $context_name); + ############ Recover the number of the columns of interest + + ############ Calculate the statistics for each file + File2Hash($file, $func_value, $exonicFunc_value, $chr_value, $ref_value, $alt_value, $strand_value, $contextSeq_value, $refH_file, $refT_func); + } +} + +# Calculate the chi2 for the strand bias +sub CalculateChi2 +{ + my ($refH_file, $folderChi2) = @_; + + # No value for the chi2 + if(scalar (keys %{$refH_file}) == 0) + { + print STDERR "Error: No value for calculating the chi2 for the strand bias\n"; + exit; + } + + # Strand bias for one mutation type for all the samples + my %h_tempchi2 = (); + my ($ca_NonTr, $ca_Tr, $cg_NonTr, $cg_Tr, $ct_NonTr, $ct_Tr, $ta_NonTr, $ta_Tr, $tc_NonTr, $tc_Tr, $tg_NonTr, $tg_Tr) = (0,0,0,0,0,0, 0,0,0,0,0,0); + + my $nb_file = 0; + + foreach my $k_file (sort keys %{$refH_file}) + { + $nb_file++; + foreach my $k_func (sort keys %{$refH_file->{$k_file}{'6mutType'}}) + { + foreach my $k_mutation (sort keys %{$refH_file->{$k_file}{'6mutType'}{$k_func}}) + { + if($k_mutation eq "C:G>A:T") + { + $h_tempchi2{'C>A'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'C>A'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + if($k_mutation eq "C:G>G:C") + { + $h_tempchi2{'C>G'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'C>G'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + if($k_mutation eq "C:G>T:A") + { + $h_tempchi2{'C>T'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'C>T'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + if($k_mutation eq "T:A>A:T") + { + $h_tempchi2{'T>A'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'T>A'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + if($k_mutation eq "T:A>C:G") + { + $h_tempchi2{'T>C'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'T>C'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + if($k_mutation eq "T:A>G:C") + { + $h_tempchi2{'T>G'}{$k_file}{'NonTr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}; + $h_tempchi2{'T>G'}{$k_file}{'Tr'} += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + } + } + } + + # Create the input file for NMF + open(CHI2, ">", "$folderChi2/Input_chi2_strandBias.txt") or die "$!: $folderChi2/Input_chi2_strandBias.txt\n"; + print CHI2 "SampleName\tNonTr\tTr\tAlteration\n"; + + foreach my $k_mutation (sort keys %h_tempchi2) + { + foreach my $k_file (sort keys %{$h_tempchi2{$k_mutation}}) + { + print CHI2 "$k_file\t$h_tempchi2{$k_mutation}{$k_file}{'NonTr'}\t$h_tempchi2{$k_mutation}{$k_file}{'Tr'}\t$k_mutation\n"; + } + } + close CHI2; + + + `Rscript $pathRscriptChi2test --folderChi2 $folderChi2 2>&1`; + # `Rscript $pathRscriptChi2test $folderChi2 2>&1`; + + + if(!-e "$folderChi2/Output_chi2_strandBias.txt") + { + print STDERR "Error: Chi2 test didn't work !!!\n"; + exit; + } +} + +# Write the different statistics in the report +sub WriteStatistics +{ + my ($refH_file, $nb_func, $folderFigure, $folderChi2, $folderNMF) = @_; + + # Save the different graphs in specific folde + if(!-e "$folderFigure/Overall_mutation_distribution") { mkdir("$folderFigure/Overall_mutation_distribution") or die "Can't create the directory $folderFigure/Overall_mutation_distribution\n"; } + if(!-e "$folderFigure/Impact_protein_sequence") { mkdir("$folderFigure/Impact_protein_sequence") or die "Can't create the directory $folderFigure/Impact_protein_sequence\n"; } + if(!-e "$folderFigure/SBS_distribution") { mkdir("$folderFigure/SBS_distribution") or die "Can't create the directory $folderFigure/SBS_distribution\n"; } + if(!-e "$folderFigure/Stranded_Analysis") { mkdir("$folderFigure/Stranded_Analysis") or die "Can't create the directory $folderFigure/Stranded_Analysis\n"; } + if(!-e "$folderFigure/Trinucleotide_Sequence_Context") { mkdir("$folderFigure/Trinucleotide_Sequence_Context") or die "Can't create the directory $folderFigure/Trinucleotide_Sequence_Context\n"; } + if(!-e "$folderFigure/Distribution_SBS_Per_Chromosomes") { mkdir("$folderFigure/Distribution_SBS_Per_Chromosomes") or die "Can't create the directory $folderFigure/Distribution_SBS_Per_Chromosomes\n"; } + + + # Create a workbook with all the samples + my $wb = ""; my $ws_sum = ""; + my ($ws_inputNMF_count, $ws_inputNMF_percent) = ("", ""); + + # Create one Excel file with all the samples + if($oneReportPerSample == 2) + { + $wb = Spreadsheet::WriteExcel->new("$folderMutAnalysis/Report_Mutation_Spectra.xls"); + + ############## Set the variables for font formats in the Excel report + Format_A10($wb, \$format_A10); # Text center in Arial 10 + Format_A10BoldLeft($wb, \$format_A10Boldleft); # Text on the left in Arial 10 bold + Format_TextSection($wb, \$formatT_left, \$formatT_right, \$formatT_bottomRight, \$formatT_bottomLeft, \$formatT_bottom, \$formatT_bottomHeader, \$formatT_bottomRightHeader, \$formatT_bottomHeader2, \$formatT_rightHeader); + Format_GraphTitle($wb, \$formatT_graphTitle); + Format_Table($wb, \$table_topleft, \$table_topRight, \$table_bottomleft, \$table_bottomRight, \$table_top, \$table_right, \$table_bottom, \$table_bottomItalicRed, \$table_left, \$table_bottomrightHeader, \$table_left2, \$table_middleHeader, \$table_middleHeader2); + Format_A10ItalicRed($wb, \$format_A10ItalicRed); + + ############### Worksheet with a summary of the samples + $ws_sum = $wb->add_worksheet("Sample_List"); + $ws_sum->write(0, 0, "Samples", $format_A10); $ws_sum->write(0, 1, "Total number SBS", $format_A10); $ws_sum->write(0, 2, "Total number of Indel", $format_A10); $ws_sum->write(0, 3, "Total number of mutations", $format_A10); + $ws_sum->set_column(0,0, 50); $ws_sum->set_column(1,1, 20); $ws_sum->set_column(2,2, 20); $ws_sum->set_column(3,3, 22); + + ############### Write the input matrix for NMF for the count and the un-normalized frequency + $ws_inputNMF_count = $wb->add_worksheet("Input_NMF_Count"); + $ws_inputNMF_percent = $wb->add_worksheet("Input_NMF_Percent"); + } + + + ################################################ Set the Rows and columns of the different part of the report + my $row_SumSheet = 1; # First row for the summary sheet of the report + my $rowStart_SBSdistrBySeg = 48; my $colStart_SBSdistrBySeg = 0; # For the table SBS distribution by segment + my $colStart_matrixSeqContext = 19; # Sequence context + my $col_inputNMF = 0; # Write the names of the samples with at least 33 SBS + + + ## For each file + foreach my $k_file (sort keys %{$refH_file}) + { + print "File in process: $k_file\n"; + + # Count the total of mutations for 6 mutation types on genomic strand + my ($c_ca6_g, $c_cg6_g, $c_ct6_g, $c_ta6_g, $c_tc6_g, $c_tg6_g) = (0,0,0, 0,0,0); + + if($k_file ne "Pool_Data") { $col_inputNMF++; } + + ############### Save the chi2 values into a hash table + if(-e "$folderChi2/Output_chi2_strandBias.txt") + { + chi2hash("$folderChi2/Output_chi2_strandBias.txt", $k_file); + } + + # Create one workbook for each sample + if($oneReportPerSample == 1) + { + $wb = Spreadsheet::WriteExcel->new("$folderMutAnalysis/Report_Mutation_Spectra-$k_file.xls"); + + ############## Set the variables for font formats in the Excel report + Format_A10($wb, \$format_A10); # Text center in Arial 10 + Format_A10BoldLeft($wb, \$format_A10Boldleft); # Text on the left in Arial 10 bold + Format_TextSection($wb, \$formatT_left, \$formatT_right, \$formatT_bottomRight, \$formatT_bottomLeft, \$formatT_bottom, \$formatT_bottomHeader, \$formatT_bottomRightHeader, \$formatT_bottomHeader2, \$formatT_rightHeader); + Format_GraphTitle($wb, \$formatT_graphTitle); + Format_Table($wb, \$table_topleft, \$table_topRight, \$table_bottomleft, \$table_bottomRight, \$table_top, \$table_right, \$table_bottom, \$table_bottomItalicRed, \$table_left, \$table_bottomrightHeader, \$table_left2, \$table_middleHeader, \$table_middleHeader2); + Format_A10ItalicRed($wb, \$format_A10ItalicRed); + + ############### Worksheet with a summary of the samples + $ws_sum = $wb->add_worksheet("Sample_List"); + $ws_sum->write(0, 0, "Samples", $format_A10); $ws_sum->write(0, 1, "Total number SBS", $format_A10); $ws_sum->write(0, 2, "Total number of Indel", $format_A10); $ws_sum->write(0, 3, "Total number of mutations", $format_A10); + $ws_sum->set_column(0,0, 50); $ws_sum->set_column(1,1, 20); $ws_sum->set_column(2,2, 20); $ws_sum->set_column(3,3, 22); + # Write in the Samples sheet the name and the total number of SBS + $ws_sum->write(1, 0, "$k_file", $format_A10); + $ws_sum->write(1, 1, $refH_file->{$k_file}{'TotalSBSGenomic'}, $format_A10); $ws_sum->write(1, 2, $refH_file->{$k_file}{'TotalIndelGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 3, $refH_file->{$k_file}{'TotalMutGenomic'}, $format_A10); + } + # One workbook with all the samples + else + { + # Write in the Samples sheet the name and the total number of SBS + $ws_sum->write($row_SumSheet, 0, $k_file, $format_A10); + $ws_sum->write($row_SumSheet, 1, $refH_file->{$k_file}{'TotalSBSGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 2, $refH_file->{$k_file}{'TotalIndelGenomic'}, $format_A10); $ws_sum->write($row_SumSheet, 3, $refH_file->{$k_file}{'TotalMutGenomic'}, $format_A10); + + # For NMF don't consider the pool of the samples + if($k_file ne "Pool_Data") + { + # Write in the input NMF sheet the name of the samples + $ws_inputNMF_count->write(0, $col_inputNMF, $k_file); + $ws_inputNMF_percent->write(0, $col_inputNMF, $k_file); + } + } + + # Calculate the correlation between the number of SBS and the size of the chromosome + PearsonCoefficient($refH_file, $k_file); + + # Add a worksheet to the workbook + my $ws = $wb->add_worksheet($k_file); + + # Write the titles of the different sections of the report + WriteBorderSection($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext); + + # Write the mutation types (6 types) + WriteHeaderSection($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext); + + + # Save the figures of each samples in a different folder + if(!-e "$folderFigure/Overall_mutation_distribution/$k_file") { mkdir("$folderFigure/Overall_mutation_distribution/$k_file") or die "Can't create the directory $folderFigure/Overall_mutation_distribution/$k_file\n"; } + if(!-e "$folderFigure/Impact_protein_sequence/$k_file") { mkdir("$folderFigure/Impact_protein_sequence/$k_file") or die "Can't create the directory $folderFigure/Impact_protein_sequence/$k_file\n"; } + if(!-e "$folderFigure/SBS_distribution/$k_file") { mkdir("$folderFigure/SBS_distribution/$k_file") or die "Can't create the directory $folderFigure/SBS_distribution\n"; } + if(!-e "$folderFigure/Stranded_Analysis/$k_file") { mkdir("$folderFigure/Stranded_Analysis/$k_file") or die "Can't create the directory $folderFigure/Stranded_Analysis/$k_file\n"; } + if(!-e "$folderFigure/Trinucleotide_Sequence_Context/$k_file") { mkdir("$folderFigure/Trinucleotide_Sequence_Context/$k_file") or die "Can't create the directory $folderFigure/Trinucleotide_Sequence_Context/$k_file\n"; } + + + + ################################################################################################################################################## + ################################################################# Write the statistics ########################################################## + ################################################################################################################################################## + my $row_SBSDistrBySegAndFunc_CG = $rowStart_SBSdistrBySeg+($nb_func*2)+16; + + + ######## Count of SBS by functional impact on the protein (Table 2) + Create the input for ggplot2 (pie chart with functional impact) + Create the input for ggplot2 (pie chart of SBS vs. Indels) + writeDistrFuncImpact($ws, $refH_file, $k_file, "$folderFigure/Impact_protein_sequence/$k_file/$k_file-DistributionExoFunc.txt", "$folderFigure/Overall_mutation_distribution/$k_file/$k_file-OverallMutationDistribution.txt"); + + + ######## Result of the chi2 for the strand bias (Table 3) + Create the input for ggplot2 (Strand bias bar graph) + writeChi2result($wb, $ws, "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandBias.txt", $refH_file, $k_file); + + + ######## SBS distribution by functional region (Table 4) + Strand bias by functional region (Table 5) + Create the input for ggplot2 (SBS distribution) + Overall count and percent of SBS (Table 1) + writeStatbyFuncRegion($refH_file, $k_file, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, \$row_SBSDistrBySegAndFunc_CG, "$folderFigure/SBS_distribution/$k_file/$k_file-SBS_distribution.txt"); + + + ######## Distribution of SBS per chromosomes and the result of Pearson test (Table 6) + writeDistrByChr($ws, $refH_file, $k_file, $row_SBSDistrBySegAndFunc_CG, $colStart_SBSdistrBySeg, "$folderFigure/Distribution_SBS_Per_Chromosomes/$k_file-DistributionSNVS_per_chromosome.txt"); + + + ######## Trinucleotide sequence context on genomic strand (Panel 1) + # Represent the trinucleotide with a heatmap with count of SBS + my $heatmapCountggplot2 = "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapCount-Genomic.txt"; + my $heatmapPercentggplot2 = "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-HeatmapPercent-Genomic.txt"; + my $triNtBarChartggplot2 = "$folderFigure/Trinucleotide_Sequence_Context/$k_file/$k_file-MutationSpectraPercent-Genomic.txt"; + + writeTriNtGenomic($ws, $refH_file, $k_file, $colStart_matrixSeqContext, $heatmapCountggplot2, $heatmapPercentggplot2, $triNtBarChartggplot2, \$c_ca6_g, \$c_cg6_g, \$c_ct6_g, \$c_ta6_g, \$c_tc6_g, \$c_tg6_g); + + # For the input matrix for NMF + if($k_file ne "Pool_Data") { push(@{$h_inputNMF{'Sample'}}, $k_file); } + + + ######## Trinucleotide sequence context on genomic strand (Panel 2) + my $triNtBarChartCodingCountggplot2 = "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignatureCount.txt"; + my $triNtBarChartCodingPercentggplot2 = "$folderFigure/Stranded_Analysis/$k_file/$k_file-StrandedSignaturePercent.txt"; + + writeTriNtCoding($ws, $rowStart_SBSdistrBySeg, $colStart_matrixSeqContext, $refH_file, $k_file, $triNtBarChartCodingCountggplot2, $triNtBarChartCodingPercentggplot2); + + + ######## Generate the figures and include them in the report + createWriteFigs($ws, $rowStart_SBSdistrBySeg, $colStart_matrixSeqContext, $folderFigure, $k_file, $c_ca6_g, $c_cg6_g, $c_ct6_g, $c_ta6_g, $c_tc6_g, $c_tg6_g); + + + # Next sample + $row_SumSheet++; + } # End $k_file + + ######## Write the input matrix for NMF + # One workbook with all the samples + writeInputNMF($ws_inputNMF_count, $ws_inputNMF_percent, "$folderNMF/Input_NMF_Count.txt", "$folderNMF/Input_NMF_Frequency.txt"); + + + # Close the workbook + $wb->close(); +} + +# Create logo for representing the sequence context with n bases +sub CreateLogo +{ + my ($refH_file, $folderWebLogo) = @_; + + my $folderSample = ""; + + foreach my $k_file (sort keys %{$refH_file}) + { + $folderSample = "$folderWebLogo/$k_file"; + if(!-e $folderSample) { mkdir($folderSample) or die "Can't create the directory $folderSample\n"; } + + my $test_lengthSeqContext = 0; + + + foreach my $k_mutation (sort keys %{$refH_file->{$k_file}{'WebLogo3'}}) + { + $k_mutation =~ /(\w)>(\w)/; + my ($ref, $alt) = ($1, $2); + + open(WEBLOGO, ">", "$folderSample/$k_file-$ref$alt.fa") or die "$!: $folderSample/$k_file-$ref$alt.fa\n"; + foreach (@{$refH_file->{$k_file}{'WebLogo3'}{$k_mutation}}) + { + print WEBLOGO ">$k_file\n$_\n"; + + if(length($_) < 10) { $test_lengthSeqContext = 0; } + else { $test_lengthSeqContext = 1; } + } + close WEBLOGO; + } + + ## Generate the logo + foreach my $fastaFile (`ls $folderSample/*.fa`) + { + chomp($fastaFile); + my ($filename, $directories, $suffix) = fileparse("$folderSample/$fastaFile", qr/\.[^.]*/); + + $filename =~ /(.+)\-/; + my $title = $1; + + ## Test if there is fasta sequences for the mutation type + my $nbLigne_temp = `wc -l $fastaFile`; + my @nbLigne = split(" ", $nbLigne_temp); + + + if($nbLigne[0] == 0) { print "WARNING: No sequence for $filename\n"; next; } + + # When length sequence context is lower than 10 the image is to small for adding a title + if($test_lengthSeqContext == 1) { system("weblogo -c classic -F png_print -U probability --title $title < $fastaFile > $folderSample/$filename-Probability.png"); } + else { system("weblogo -c classic -F png_print -U probability < $fastaFile > $folderSample/$filename-Probability.png"); } + } + } +} + + + +### Save the count of SBS for each file into a hash table +sub File2Hash +{ + my ($inputFile, $func_value, $exonicFunc_value, $chr_value, $ref_value, $alt_value, $strand_value, $contextSeq_value, $refH_file, $refT_func) = @_; + + my ($filename, $directories, $suffix) = fileparse($inputFile, qr/\.[^.]*/); + + # Initialisation of the hash + my @tab_mutation = qw(C:G>A:T C:G>G:C C:G>T:A T:A>A:T T:A>C:G T:A>G:C); + my @tab_aaChange = ("NonTr", "Tr", "TotalMutG"); + my @tabExoFunc = ("frameshift insertion", "frameshift deletion", "frameshift block substitution", "frameshift substitution", "stopgain", "stoploss", "nonframeshift insertion", "nonframeshift deletion", "nonframeshift substitution", "nonframeshift block substitution", "nonsynonymous SNV", "synonymous SNV", "unknown", "NA"); + + # Total number of SBS on the genomic strand + $refH_file->{$filename}{'TotalSBSGenomic'} = 0; + # Total number of Indel on the genomic strand + $refH_file->{$filename}{'TotalIndelGenomic'} = 0; + # Total number of SBS on the coding strand + $refH_file->{$filename}{'TotalSBSCoding'} = 0; + # Total number of SBS and Indel on the genomic strand + $refH_file->{$filename}{'TotalMutGenomic'} = 0; + + + ##################################################### + # Initialisation of the tables and hash tables # + ##################################################### + + ## SBS by segment (6 mutation types) + foreach my $elt_tabFunc (@$refT_func) + { + foreach my $elt_tabMutation (@tab_mutation) + { + foreach my $elt_aaChange (@tab_aaChange) + { + $refH_file->{$filename}{'6mutType'}{$elt_tabFunc}{$elt_tabMutation}{$elt_aaChange} = 0; + } + } + } + + ## Pearson correlation + $refH_file->{$filename}{'SBSPerChr'}{'AllMutType'} = 0; + # Count of SBS per chromosome foreach mutation types + foreach my $elt_tabMutation (@tab_mutation) + { + foreach my $chromosome (sort keys %chromosomes){ $refH_file->{$filename}{'SBSPerChr'}{$elt_tabMutation}{'CHR'}{$chromosome}{'chr'} = 0;} + $refH_file->{$filename}{'SBSPerChr'}{$elt_tabMutation}{'Pearson'} = 0; + } + foreach my $chromosome (sort keys %chromosomes) + { + $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}=0; + } + + ## Impact of SBS on protein + foreach my $elt_exoFunc (@tabExoFunc) + { + $refH_file->{$filename}{'ImpactSBS'}{$elt_exoFunc} = 0; + } + + ## Sequence context (genomic strand) + my @tab_mutation2 = qw(C>A C>G C>T T>A T>C T>G); + my @tab_context = qw(A_A A_C A_G A_T C_A C_C C_G C_T G_A G_C G_G G_T T_A T_C T_G T_T); + foreach my $elt_context (@tab_context) + { + foreach my $elt_mutation3 (@tab_mutation2) + { + $refH_file->{$filename}{'SeqContextG'}{$elt_context}{$elt_mutation3} = 0; + } + } + + ## Sequence context (coding strand) + my @tab_TrNonTr = qw(NonTr Tr); + foreach my $elt_context (@tab_context) + { + foreach my $elt_mutation2 (@tab_mutation2) + { + foreach my $trNonTr (@tab_TrNonTr) + { + $refH_file->{$filename}{'SeqContextC'}{$elt_context}{$elt_mutation2}{$trNonTr} = 0; + } + } + } + + + ##################################################### + # Parse the intput file # + ##################################################### + + open(F1,$inputFile) or die "$!: $inputFile\n"; + my $header = <F1>; + while(<F1>) + { + $_ =~ s/[\r\n]+$//; + my @tab = split("\t", $_); + + ### Don't consider random chromosomes and chromosome M + if( ($tab[$chr_value] =~ /random/i) || ($tab[$chr_value] =~ /M/i) ) { next; } + + + ### Recover the trinucleotide sequence context: Extract the base just before and after the mutation + my $context = ""; + my $contextSequence = $tab[$contextSeq_value]; $contextSequence =~ tr/a-z/A-Z/; + my @tempContextSequence = split("", $contextSequence); + my $total_nbBaseContext = $#tempContextSequence; + my $midlle_totalNbBaseContext = $total_nbBaseContext/2; # For having the middle of the sequence + my $before = $midlle_totalNbBaseContext - 1; my $after = $midlle_totalNbBaseContext + 1; + $context = $tempContextSequence[$before]."_".$tempContextSequence[$after]; + + + + ### Recover the annotations on the impact on the protein for creating the pie chart + my $exoFunc = ""; + # Sometimes the annotation is repeated frameshift deletion;frameshift deletion + if($tab[$exonicFunc_value] =~ /\;/) + { + my @temp = split(";", $tab[$exonicFunc_value]); + if($temp[0] eq $temp[1]) { $exoFunc = $temp[0]; } + } + # The annotations have changed after MAJ Annovar 2014Jul22 (stopgain SNV => stopgain) + elsif($tab[$exonicFunc_value] eq "stopgain SNV") { $exoFunc = "stopgain"; } + elsif($tab[$exonicFunc_value] eq "stoploss SNV") { $exoFunc = "stoploss"; } + elsif($tab[$exonicFunc_value] eq "nonsynonymous_SNV") { $exoFunc = "nonsynonymous SNV"; } + elsif($tab[$exonicFunc_value] eq "stopgain_SNV") { $exoFunc = "stopgain SNV"; } + elsif($tab[$exonicFunc_value] eq "synonymous_SNV") { $exoFunc = "synonymous SNV"; } + else { $exoFunc = $tab[$exonicFunc_value]; } + + if(exists $refH_file->{$filename}{'ImpactSBS'}{$exoFunc}) + { + # If the sequence context if not recovered correctly don't considered the variants + if( ($context =~ /N/) || (length($context) != 3) ) { next; } + + $refH_file->{$filename}{'ImpactSBS'}{$exoFunc}++; + $refH_file->{$filename}{'TotalMutGenomic'}++; + } + else { print "WARNING: Exonic function not considered: $exoFunc\n"; } + + #### Only SBS are considered for the statistics + if( ($tab[$ref_value] =~ /^[ACGT]$/i) && ($tab[$alt_value] =~ /^[ACGT]$/i) ) + { + # If the sequence context if not recovered correctly don't considered the variants + if( ($context =~ /N/) || (length($context) != 3) ) { next; } + + # Total number of SBS on the genomic strand + $refH_file->{$filename}{'TotalSBSGenomic'}++; + + # Total number of SBS on the coding strand with a sequence context + if( ($tab[$strand_value] eq "+") || ($tab[$strand_value] eq "-") ) + { + if( ($context ne "NA") && (($context =~ /N/) || (length($context) != 3)) ) { next; } + $refH_file->{$filename}{'TotalSBSCoding'}++; + } + } + else { $refH_file->{$filename}{'TotalIndelGenomic'}++; } + + ### Number of SBS per chromosome: remove the "chr" + my $chrNameForH=$tab[$chr_value]; + if(exists $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chrNameForH}{'chr'}) + { + $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chrNameForH}{'chr'}++; + } + + + #### Some func value are repeated and separated by ";" + my $funcSegment = ""; + if($tab[$func_value] =~ /;/) { my @temp = split(";", $tab[$func_value]); $funcSegment = $temp[0]; } + else { $funcSegment = $tab[$func_value]; } + + + + ##################################################### + # Calculate the statistics for each mutation type # + ##################################################### + if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "A")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "T") ) ) + { + my $mutation = "C:G>A:T"; + + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "G")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "C") ) ) + { + my $mutation = "C:G>G:C"; + + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + if( (($tab[$ref_value] eq "C") && ($tab[$alt_value] eq "T")) || ( ($tab[$ref_value] eq "G") && ($tab[$alt_value] eq "A") ) ) + { + my $mutation = "C:G>T:A"; + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "A")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "T") ) ) + { + my $mutation = "T:A>A:T"; + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "C")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "G")) ) + { + my $mutation = "T:A>C:G"; + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + if( (($tab[$ref_value] eq "T") && ($tab[$alt_value] eq "G")) || ( ($tab[$ref_value] eq "A") && ($tab[$alt_value] eq "C")) ) + { + my $mutation = "T:A>G:C"; + statPerMutType($filename, \@tab, $ref_value, $alt_value, \@tempContextSequence, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext); + } + } + close F1; +} + +### Count the number of SBS for 12 and 6 categories +sub statPerMutType +{ + my ($filename, $refTab, $ref_value, $alt_value, $refTab_tempSeqContext, $before, $after, $context, $funcSegment, $mutation, $refH_file, $chrNameForH, $strand_value, $midlle_totalNbBaseContext) = @_; + + my @tab = @$refTab; + my @tempContextSequence = @$refTab_tempSeqContext; + + + # Split the mutations + $mutation =~ /(\w)\:(\w)\>(\w)\:(\w)/; + my ($ref1, $ref2, $alt1, $alt2) = ($1, $2, $3, $4); + + # Count the total number of mutations + $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'TotalMutG'}++; + + # Pearson correlation + if(exists $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}) + { + $refH_file->{$filename}{'SBSPerChr'}{$mutation}{'CHR'}{$chrNameForH}{'chr'}++; + } + + #### Sequence context - 6 mutation types - genomic strand + my $mutationSeqContext6mutType = "$ref1>$alt1"; + # We want to express the mutation in C> or T> + if( ($tab[$ref_value] eq $ref2) && ($tab[$alt_value] eq $alt2) ) + { + my $base3 = complement($tempContextSequence[$before]); + my $base5 = complement($tempContextSequence[$after]); + my $context_reverse = $base5."_".$base3; + if(exists $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}) + { + $refH_file->{$filename}{'SeqContextG'}{$context_reverse}{$mutationSeqContext6mutType}++; + } + } + elsif(exists $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}) + { + $refH_file->{$filename}{'SeqContextG'}{$context}{$mutationSeqContext6mutType}++; + } + + + #### Strand analysis C>N and T>N on NonTr strand + if( (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq $ref1)&&($tab[$alt_value] eq $alt1))) || (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq $ref2)&&($tab[$alt_value] eq $alt2))) ) + { + if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}) + { + $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'NonTr'}++; + } + + # C>A With the sequence context (C>N and T>N on strand +) + if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq $ref1)&&($tab[$alt_value] eq $alt1)) ) + { + if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{$mutationSeqContext6mutType}{'NonTr'}) + { + $refH_file->{$filename}{'SeqContextC'}{$context}{$mutationSeqContext6mutType}{'NonTr'}++; + } + } + # C>A With the sequence context (G>N and A>N on strand -) + else + { + my $base3 = complement($tempContextSequence[$before]); + my $base5 = complement($tempContextSequence[$after]); + my $context_reverse = $base5."_".$base3; + + if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{$mutationSeqContext6mutType}{'NonTr'}) + { + $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{$mutationSeqContext6mutType}{'NonTr'}++; + } + } + } + + #### Strand analysis C>N and T>N on Tr strand + if( (($tab[$strand_value] eq "-") && (($tab[$ref_value] eq $ref1)&&($tab[$alt_value] eq $alt1))) || (($tab[$strand_value] eq "+") && (($tab[$ref_value] eq $ref2)&&($tab[$alt_value] eq $alt2))) ) + { + if(exists $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}) + { + $refH_file->{$filename}{'6mutType'}{$funcSegment}{$mutation}{'Tr'}++; + } + + # C>N and T>N With the sequence context (strand -) + if( ($tab[$strand_value] eq "-") && (($tab[$ref_value] eq $ref1)&&($tab[$alt_value] eq $alt1)) ) + { + if(exists $refH_file->{$filename}{'SeqContextC'}{$context}{$mutationSeqContext6mutType}{'Tr'}) + { + $refH_file->{$filename}{'SeqContextC'}{$context}{$mutationSeqContext6mutType}{'Tr'}++; + } + } + # C>N and T>N with the sequence context (strand +) + if( ($tab[$strand_value] eq "+") && (($tab[$ref_value] eq $ref2)&&($tab[$alt_value] eq $alt2)) ) + { + my $base3 = complement($tempContextSequence[$before]); + my $base5 = complement($tempContextSequence[$after]); + my $context_reverse = $base5."_".$base3; + if(exists $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{$mutationSeqContext6mutType}{'Tr'}) + { + $refH_file->{$filename}{'SeqContextC'}{$context_reverse}{$mutationSeqContext6mutType}{'Tr'}++; + } + } + } + + #### WebLogo-3 + if(($tab[$ref_value] eq $ref1) && ($tab[$alt_value] eq $alt1)) + { + # For the logo all the sequences must have the same length + if(scalar(@tempContextSequence) == 2) { next; } + my ($contextTemp1, $contextTemp2) = ("", ""); + for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= $tempContextSequence[$i]; } + for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= $tempContextSequence[$i]; } + my $context = $contextTemp1.$ref1.$contextTemp2; + push(@{$refH_file->{$filename}{'WebLogo3'}{$mutationSeqContext6mutType}}, $context); + } + else + { + + if(scalar(@tempContextSequence) == 2) { next; } + my ($contextTemp1, $contextTemp2) = ("", ""); + for(my $i=0; $i<$midlle_totalNbBaseContext; $i++) { $contextTemp1 .= complement($tempContextSequence[$i]); } + for(my $i=$midlle_totalNbBaseContext+1; $i<=$#tempContextSequence; $i++) { $contextTemp2 .= complement($tempContextSequence[$i]); } + my $context = $contextTemp1.$ref1.$contextTemp2; $context = reverse $context; + push(@{$refH_file->{$filename}{'WebLogo3'}{$mutationSeqContext6mutType}}, $context); + } +} + +# Calculate the correlation between the number of SBS and the size of the chromosome +sub PearsonCoefficient +{ + our ($refH_file, $filename) = @_; + + #### Calculate the Pearson coefficient + my @total_SBS = (); # Pearson for all mutation types + + # Create a 2D array + foreach my $k_mutation (sort keys %{$refH_file->{$filename}{'SBSPerChr'}}) + { + my $x = []; + my $correlation = 0; + + if($k_mutation eq "AllMutType") { next; } + elsif($k_mutation eq "TotalPerChr") { next; } + elsif($k_mutation eq "ChrSize") { next; } + else + { + my $testZero = 0; # The correlation function doesn't works if all the variables are equal to zero + # generate an anonymous 2D array where $x->[1] is the row + # $x->[1][1] is the value in row 1 column 1 and $x->[1][2] is the value of row 1 column 2 + # once you build the entire array, pass it to the correlation subroutine + my $i=1; + while ( my ($chromosome, $lenght) = each (%chromosomes)) + { + $x->[$i][1] = $lenght; # First column contains the chromosome size + $x->[$i][2] = $refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'}{$chromosome}{'chr'}; # Second column contains the count of SBS + if($refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'}{$chromosome}{'chr'}==0) { $testZero++; } + $i++; + } + if( $testZero == keys %{$refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'CHR'}} ) + { + $correlation = 0; + } + # Pass the 2D array to the correlation subroutine + else + { + $correlation = correlation($x); + } + + $refH_file->{$filename}{'SBSPerChr'}{$k_mutation}{'Pearson'} = $correlation; # Pearson per mutation type + } + } + + #generate an anonymous 2D array for all mutation type + my $testZero = 0; + my $x = []; + my $correlation = 0; + my $i=1; + while ( my ($chromosome, $lenght) = each (%chromosomes)) + { + $x->[$i][1] = $lenght; + $x->[$i][2] = $refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}; + $i++; + } + if($testZero == keys %{$refH_file->{$filename}{'SBSPerChr'}{'TotalPerChr'}} ) { $correlation = 0; } + else { $correlation = correlation($x); } + # Pass the 2D array to the correlation subroutine + $refH_file->{$filename}{'SBSPerChr'}{'AllMutType'} = $correlation; + + sub correlation + { + my ($x) = @_; + my ($mean_x,$mean_y) = mean($x); + my $ssxx=ss($x,$mean_x,$mean_y,1,1); + my $ssyy=ss($x,$mean_x,$mean_y,2,2); + my $ssxy=ss($x,$mean_x,$mean_y,1,2); + my $correl=correl($ssxx,$ssyy,$ssxy);; + my $xcorrel=sprintf("%.2f",$correl); + return($xcorrel); + + sub mean + { + my ($x)=@_; + my $num = scalar(@{$x}) - 2; + my $sum_x = '0'; + my $sum_y = '0'; + for (my $i = 2; $i < scalar(@{$x}); ++$i) + { + $sum_x += $x->[$i][1]; + $sum_y += $x->[$i][2]; + } + my $mu_x = $sum_x / $num; + my $mu_y = $sum_y / $num; + return($mu_x,$mu_y); + } + + ### ss = sum of squared (deviations to the mean) + sub ss + { + my ($x,$mean_x,$mean_y,$one,$two)=@_; + my $sum = '0'; + for (my $i=2;$i<scalar(@{$x});++$i) + { + $sum += ($x->[$i][$one]-$mean_x)*($x->[$i][$two]-$mean_y); + } + return $sum; + } + + sub correl + { + my($ssxx,$ssyy,$ssxy)=@_; + + my ($sign, $correl) = (0,0); + if(abs($ssxy) == 0) { $sign = 0 } + else { $sign=$ssxy/abs($ssxy); } + + if( ($ssxx==0) || ($ssyy==0) ) { $correl = 0 } + else { $correl=$sign*sqrt($ssxy*$ssxy/($ssxx*$ssyy)); } + + return $correl; + } + } +} + +# Save the output of the chi2 into a hash table for writing the results in the Excel file +sub chi2hash +{ + my ($outputChi2, $k_file) = @_; + + open(F1, $outputChi2) or die "$!: $outputChi2\n"; + my $header = <F1>; + # Strand_Bias($tab[0]) NonTr-Tr($tab[1]) Proportion($tab[2]) P-val-Chi2($tab[3]) FDR($tab[4]) Confidence Interval($tab[5]) Mutation_Type($tab[6]) SampleName($tab[7]) + while(<F1>) + { + $_ =~ s/[\r\n]+$//; + my @tab = split("\t", $_); + + if($tab[7] eq $k_file) + { + if($tab[1] eq "NA") + { + $h_chi2{$tab[7]}{$tab[6]}{'NonTr'} = "NA"; + $h_chi2{$tab[7]}{$tab[6]}{'Tr'} = "NA"; + } + else + { + my ($nonTr, $tr) = split("-", $tab[1]); + + $h_chi2{$tab[7]}{$tab[6]}{'NonTr'} = $nonTr; + $h_chi2{$tab[7]}{$tab[6]}{'Tr'} = $tr; + } + + + $h_chi2{$tab[7]}{$tab[6]}{'p-value'} = $tab[3]; + $h_chi2{$tab[7]}{$tab[6]}{'ConfInt'} = $tab[5]; + + # For the pool data the FDR isn't calculated so replace the NA (=Missing values) by "-" + if($tab[7] eq "Pool_Data") + { + $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = "-"; + } + else + { + $h_chi2{$tab[7]}{$tab[6]}{'FDR'} = $tab[4]; + } + } + } + close F1; +} + +### Complement bases (for the sequence context) +sub complement +{ + if($_[0] eq "A") { return "T"; } + if($_[0] eq "C") { return "G"; } + if($_[0] eq "G") { return "C"; } + if($_[0] eq "T") { return "A"; } +} + +### Recover the functional region for all the samples. Allows to thave the same annotations for the pie chart "Impact on protein sequence" +sub recoverAnnovarAnnotation +{ + my ($func_name) = @_; + + my %hash = (); + + # The input is a folder + foreach my $file (`ls $folderCheckedForStat/*`) + { + chomp($file); + my $AV_annotation_value = recoverNumCol($file, $func_name); + + open(F1, $file) or die "$!: $file\n"; + my $header = <F1>; + while(<F1>) + { + $_ =~ s/[\r\n]+$//; + my @tab = split("\t", $_); + + # Some files can have an empty line at the end and WE DON'T WANT to consider it + if(! defined $tab[0]) { next; } + # Some func value are repeated and separated by ";" + my $funcSegment = ""; + if($tab[$AV_annotation_value] =~ /;/) + { + my @temp = split(";", $tab[$AV_annotation_value]); + $funcSegment = $temp[0]; + } + else { $funcSegment = $tab[$AV_annotation_value]; } + + $hash{$funcSegment} = ""; + } + close F1; + } + my @tab_AVAnnotation = (); + foreach my $k (sort keys %hash) { push(@tab_AVAnnotation, $k); } + return @tab_AVAnnotation; +} + +sub recoverNumCol +{ + my ($input, $name_of_column) = @_; + + open(F1,$input) or die "recoverNumCol: $!: $input\n"; + # For having the name of the columns + my $search_header = <F1>; $search_header =~ s/[\r\n]+$//; my @tab_search_header = split("\t",$search_header); + close F1; + # The number of the column + my $name_of_column_NB = "toto"; + for(my $i=0; $i<=$#tab_search_header; $i++) + { + if($tab_search_header[$i] eq $name_of_column) { $name_of_column_NB = $i; last; } + } + if($name_of_column_NB eq "toto") { print STDERR "Error recoverNumCol(): the column named $name_of_column doesn't exits in the input file $input!!!!!\n"; exit 3; } + else { return $name_of_column_NB; } +} + + + +###################################################################################################################################################### +# Functions for writing in the Excel report # +###################################################################################################################################################### +# Write the header for the six mutation types +sub WriteHeaderSection +{ + our ($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext) = @_; + + our ($format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG, $format_TG2, $format_LeftHeader, $format_RightHeader, $format_LeftHeader2); + Format_Header($wb, \$format_CA, \$format_CG, \$format_CT, \$format_TA, \$format_TC, \$format_TG, \$format_TG2, \$format_LeftHeader, \$format_RightHeader, \$format_LeftHeader2); + + our ($format_LeftCA, $format_LeftCG, $format_LeftCT, $format_LeftTA, $format_LeftTC, $format_LeftTG, $format_RightCA, $format_RightCG, $format_RightCT, $format_RightTA, $format_RightTC, $format_RightTG); + Format_HeaderSBSDistrBySegAndFunc($wb, \$format_LeftCA, \$format_LeftCG, \$format_LeftCT, \$format_LeftTA, \$format_LeftTC, \$format_LeftTG, \$format_RightCA, \$format_RightCG, \$format_RightCT, \$format_RightTA, \$format_RightTC, \$format_RightTG); + + our $format_A11Bold = ""; Format_A11Bold($wb, \$format_A11Bold); # Arial 11 bold and center + our $format_A11BoldLeft = ""; Format_A11BoldLeft($wb, \$format_A11BoldLeft); # Arial 11 bold and left + + our ($format_header12CA, $format_header12CG, $format_header12CT, $format_header12TA, $format_header12TC, $format_header12TG); + Format_Header12MutType($wb, \$format_header12CA, \$format_header12CG, \$format_header12CT, \$format_header12TA, \$format_header12TC, \$format_header12TG); + + ## Header for SBS distribution by segment + HeaderMutTypeSBSDistrBySeg(); + + ## Header for strand bias by function + $ws->set_column($colStart_SBSdistrBySeg+5, $colStart_SBSdistrBySeg+5, 11); + + my $row = $rowStart_SBSdistrBySeg+$nb_func+10; my $col = $colStart_SBSdistrBySeg; + $ws->write($row, $col+1, ' ', $format_CA); $ws->write($row, $col+2, "C>A", $format_CA); $ws->write($row, $col+3, ' ', $format_CA); + $ws->write($row, $col+5, ' ', $format_CG); $ws->write($row, $col+6, "C>G", $format_CG); $ws->write($row, $col+7, ' ', $format_CG); + $ws->write($row, $col+9, ' ', $format_CT); $ws->write($row, $col+10, "C>T", $format_CT); $ws->write($row, $col+11, ' ', $format_RightCT); + + $row = $rowStart_SBSdistrBySeg+($nb_func*2)+14; + $ws->write($row, $col+1, ' ', $format_TA); $ws->write($row, $col+2, "T>A", $format_TA); $ws->write($row, $col+3, ' ', $format_TA); + $ws->write($row, $col+5, ' ', $format_TC); $ws->write($row, $col+6, "T>C", $format_TC); $ws->write($row, $col+7, ' ', $format_TC); + $ws->write($row, $col+9, ' ', $format_TG2); $ws->write($row, $col+10, "T>G", $format_TG2); $ws->write($row, $col+11, ' ', $format_RightTG); + + $ws->set_row($rowStart_SBSdistrBySeg+$nb_func+11, 18); $ws->set_row($rowStart_SBSdistrBySeg+($nb_func*2)+15, 18); + $ws->set_column($colStart_SBSdistrBySeg+5, $colStart_SBSdistrBySeg+5, 13); $ws->set_column($colStart_SBSdistrBySeg+9, $colStart_SBSdistrBySeg+9, 13); + + for(my $i=$rowStart_SBSdistrBySeg+$nb_func+10; $i<=$rowStart_SBSdistrBySeg+($nb_func*2)+14; $i+=$nb_func+4) + { + $ws->write($i+1, $colStart_SBSdistrBySeg, 'Segment', $format_LeftHeader); $ws -> write($i+1, $colStart_SBSdistrBySeg+1, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+2, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+3, 'Tr', $format_A11Bold); + $ws -> write($i+1, $colStart_SBSdistrBySeg+5, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+6, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+7, 'Tr', $format_A11Bold); + $ws -> write($i+1, $colStart_SBSdistrBySeg+9, 'Non-Tr/Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+10, 'Non-Tr', $format_A11Bold); $ws -> write($i+1, $colStart_SBSdistrBySeg+11, 'Tr', $format_RightHeader); + } + + + ## Header for Counts of SBS per chromosome and mutation type + HeaderCountSBSPerChr(); + + ## Header for the short sequence context + HeaderShortTriNtContext(); + + ## Header for the 12 mutation types with the sequence context (coding strand) + HeaderLongTriNtContext(); + + sub HeaderMutTypeSBSDistrBySeg + { + $ws->set_row($rowStart_SBSdistrBySeg+2, 18); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+2, "C:G>A:T", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+3, $format_CA); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+4, "C:G>G:C", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+5, $format_CG); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+6, "C:G>T:A", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+7, $format_CT); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+8, "T:A>A:T", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+9, $format_TA); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+10, "T:A>C:G", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+11, $format_TC); + $ws->write($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+12, "T:A>G:C", $format_TG); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg+13, $format_TG); + + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg, "Segment", $format_LeftHeader); $ws->set_column($colStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, 13); $ws->set_row($rowStart_SBSdistrBySeg+3, 18); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+1, "Total SBS", $format_A11Bold); $ws->set_column($colStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg+1, 11); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+2, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+3, "#", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+4, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+5, "#", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+6, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+7, "#", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+8, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+9, "#", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+10, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+11, "#", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_SBSdistrBySeg+12, "%", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+3, 13, "#", $format_RightHeader); + } + + sub HeaderCountSBSPerChr + { + $ws->set_column(3,3, 10); $ws->set_column(4,4, 10); + $ws->set_row($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, 18); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+7, $colStart_SBSdistrBySeg+1, "Pearson", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg, "Chr", $format_LeftHeader); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+1, "Size", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+2, "All SBS", $format_A11Bold); + + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+3, "C:G>A:T", $format_CA); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+4, "C:G>G:C", $format_CG); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+5, "C:G>T:A", $format_CT); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+6, "T:A>A:T", $format_TA); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+7, "T:A>C:G", $format_TC); + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+8, $colStart_SBSdistrBySeg+8, "T:A>G:C", $format_TG); + } + + sub HeaderShortTriNtContext + { + ### GENOMIC STRAND + $ws->write(2, $colStart_matrixSeqContext, 'Count matrix', $format_LeftHeader2); + $ws->write(3, $colStart_matrixSeqContext+4, 'C>A', $format_CA); $ws->write(3, $colStart_matrixSeqContext+5, 'C>G', $format_CG); $ws->write(3, $colStart_matrixSeqContext+6, 'C>T', $format_CT); $ws->write(3, $colStart_matrixSeqContext+7, 'T>A', $format_TA); $ws->write(3, $colStart_matrixSeqContext+8, 'T>C', $format_TC); $ws->write(3, $colStart_matrixSeqContext+9, 'T>G', $format_TG2); + + $ws->write(2, $colStart_matrixSeqContext+11, 'Frequency matrix', $format_A11BoldLeft); + $ws->write(3, $colStart_matrixSeqContext+14, 'C>A', $format_CA); $ws->write(3, $colStart_matrixSeqContext+15, 'C>G', $format_CG); $ws->write(3, $colStart_matrixSeqContext+16, 'C>T', $format_CT); $ws->write(3, $colStart_matrixSeqContext+17, 'T>A', $format_TA); $ws->write(3, $colStart_matrixSeqContext+18, 'T>C', $format_TC); $ws->write(3, $colStart_matrixSeqContext+19, 'T>G', $format_TG2); + + ### sequence context with a bar graph + $ws->write(25, $colStart_matrixSeqContext+10, "Mutation spectra frequency", $format_A11Bold); + } + + sub HeaderLongTriNtContext + { + $ws->set_row($rowStart_SBSdistrBySeg+3, 15); $ws->set_row($rowStart_SBSdistrBySeg+4, 15); $ws->set_row($rowStart_SBSdistrBySeg+5, 15); + $ws->write($rowStart_SBSdistrBySeg+3, $colStart_matrixSeqContext, "Count matrix", $format_LeftHeader2); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+1, "C>A", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+2, $format_CA); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+1, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+2, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+3, "C>G", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+4, $format_CG); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+3, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+4, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+5, "C>T", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+6, $format_CT); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+5, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+6, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+7, "T>A", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+8, $format_TA); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+7, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+8, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+9, "T>C", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+10, $format_TC); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+9, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+10, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+11, "T>G", $format_TG2); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext+12, $format_TG2); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+11, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext+12, "Tr", $format_A11Bold); + + + $ws->set_row($rowStart_SBSdistrBySeg+24, 15); $ws->set_row($rowStart_SBSdistrBySeg+25, 15); $ws->set_row($rowStart_SBSdistrBySeg+26, 15); + $ws->write($rowStart_SBSdistrBySeg+24, $colStart_matrixSeqContext, "Frequency matrix", $format_LeftHeader2); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+1, "C>A", $format_CA); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+2, $format_CA); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+1, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+2, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+3, "C>G", $format_CG); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+4, $format_CG); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+3, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+4, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+5, "C>T", $format_CT); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+6, $format_CT); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+5, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+6, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+7, "T>A", $format_TA); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+8, $format_TA); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+7, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+8, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+9, "T>C", $format_TC); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+10, $format_TC); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+9, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+10, "Tr", $format_A11Bold); + $ws->write($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+11, "T>G", $format_TG2); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext+12, $format_TG2); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+11, "NonTr", $format_A11Bold); $ws->write($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext+12, "Tr", $format_A11Bold); + } +} +# Write the titles of the different sections of the report +sub WriteBorderSection +{ + our ($wb, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $colStart_matrixSeqContext) = @_; + + our ($format_topLeft, $format_topRight, $format_bottomLeft, $format_bottomRight, $format_top, $format_right, $format_bottom, $format_left); + Format_section($wb, \$format_topLeft, \$format_topRight, \$format_bottomLeft, \$format_bottomRight, \$format_top, \$format_right, \$format_bottom, \$format_left); + + TableSBSDistrBySeg(); + TableStrandBiasBySegment(); + CountSBSPerChr(); + ShortTriNtContext(); # 6 mut type + LongTriNtContext(); # 12 mut type + + sub TableSBSDistrBySeg + { + # Top-Left + $ws->write($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, "Table 4. SBS distribution by functional region", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg, 18); # Set the height of the row to 0.25" + # Top + for(my $i=1; $i<=13; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+$i, $format_top); } + # Top-Right + $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+13, $format_topRight); + # Right + $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg+13, $format_right); + # Bottom-left + $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+5, $colStart_SBSdistrBySeg, $format_bottomLeft); + # Left + $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_SBSdistrBySeg, $format_left); + } + + sub TableStrandBiasBySegment + { + # Top-Left + $ws->write($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg, "Table 5. Strand bias by functional region", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg+$nb_func+8, 18); # Set the height of the row to 0.25" + # Top + for(my $i=1; $i<=10; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg+$i, $format_top); } + # Top-Right + $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+8, $colStart_SBSdistrBySeg+11, $format_topRight); + # Right + $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+9, $colStart_SBSdistrBySeg+11, $format_right); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+13, $colStart_SBSdistrBySeg+11, $format_right); + # Left + $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+9, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+$nb_func+10, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+13, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*2)+14, $colStart_SBSdistrBySeg, $format_left); + # Bottom + $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*3)+16, $colStart_SBSdistrBySeg+4, $format_bottom); $ws->write_blank($rowStart_SBSdistrBySeg+($nb_func*3)+16, $colStart_SBSdistrBySeg+8, $format_bottom); + } + + sub CountSBSPerChr + { + #### Top-Left + $ws->write($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg, "Table 6. SBS distribution per chromosome", $format_topLeft); $ws->set_row($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, 18); # Set the height of the row to 0.25" + #### Top + for(my $i=1; $i<8; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg+$i, $format_top); } + #### Top-Right + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+4, $colStart_SBSdistrBySeg+8, $format_topRight); + #### Right + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+5, $colStart_SBSdistrBySeg+8, $format_right); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+6, $colStart_SBSdistrBySeg+8, $format_right); + + #### Bottom-Right + # Human genome = 24 chromosomes + if($refGenome =~ /hg/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+8, $format_bottomRight); } + # Mouse genome = 21 chromosomes + if($refGenome =~ /mm/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+8, $format_bottomRight); } + # Rat genome = 22 chromosomes + if($refGenome =~ /rn/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+8, $format_bottomRight); } + + #### Bottom + if($refGenome =~ /hg/) + { + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+1, $format_bottom); + for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg+$i, $format_bottom); } + } + if($refGenome =~ /mm/) + { + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+1, $format_bottom); + for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg+$i, $format_bottom); } + } + if($refGenome =~ /rn/) + { + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+1, $format_bottom); + for(my $i=3; $i<=7; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg+$i, $format_bottom); } + } + + #### Left + $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+5, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+6, $colStart_SBSdistrBySeg, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+7, $colStart_SBSdistrBySeg, $format_left); + + #### Bottom-left + if($refGenome =~ /hg/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+33, $colStart_SBSdistrBySeg, $format_bottomLeft); } + if($refGenome =~ /mm/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+30, $colStart_SBSdistrBySeg, $format_bottomLeft); } + if($refGenome =~ /rn/) { $ws->write_blank($rowStart_SBSdistrBySeg+8+$nb_func+(($nb_func+4)*2)+31, $colStart_SBSdistrBySeg, $format_bottomLeft); } + } + + sub ShortTriNtContext + { + my $format_headerSection = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); + $format_headerSection->set_left(2); $format_headerSection->set_left_color('blue'); + + # Top-left + $ws->write(0, $colStart_matrixSeqContext, "Panel 1. Trinucleotide sequence context of SBS on the genomic sequence", $format_topLeft); + # Top + for(my $i=1; $i<=19; $i++) { $ws->write_blank(0, $colStart_matrixSeqContext+$i, $format_top); } + # Top-right + $ws->write_blank(0, $colStart_matrixSeqContext+20, $format_topRight); + # Right + for(my $i=1; $i<=37; $i++) { $ws->write_blank($i, $colStart_matrixSeqContext+20, $format_right); } + # Bottom-right + $ws->write_blank(37, $colStart_matrixSeqContext+20, $format_bottomRight); + # Bottom + for(my $i=1; $i<=19; $i++) { $ws->write_blank(38, $colStart_matrixSeqContext+$i, $format_top); } + # Bottom-left + $ws->write_blank(37, $colStart_matrixSeqContext, $format_bottomLeft); + # Left + $ws->write(1, $colStart_matrixSeqContext, "", $format_left); + for(my $i=3; $i<=36; $i++) { $ws->write_blank($i, $colStart_matrixSeqContext, $format_left); } + } + + sub LongTriNtContext + { + # Top-left + $ws->write($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext, "Panel 2. Stranded analysis of trinucleotide sequence context of SBS", $format_topLeft); + # Top + for(my $i=1; $i<=28; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext+$i, $format_top); } + # Top-right + $ws->write_blank($rowStart_SBSdistrBySeg, $colStart_matrixSeqContext+29, $format_topRight); + # Right + for(my $i=1; $i<=42; $i++) { $ws->write_blank($rowStart_SBSdistrBySeg+$i, $colStart_matrixSeqContext+29, $format_right); } + # Bottom-right + $ws->write_blank(91, $colStart_matrixSeqContext+29, $format_bottomRight); + # Bottom + for(my $i=13; $i<=28; $i++) { $ws->write_blank(92, $colStart_matrixSeqContext+$i, $format_top); } + # Bottom-left + $ws->write_blank(91, $colStart_matrixSeqContext, $format_bottomLeft); + # Left + $ws->write_blank($rowStart_SBSdistrBySeg+1, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+2, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+4, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+5, $colStart_matrixSeqContext, $format_left); + $ws->write_blank($rowStart_SBSdistrBySeg+22, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+23, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+25, $colStart_matrixSeqContext, $format_left); $ws->write_blank($rowStart_SBSdistrBySeg+26, $colStart_matrixSeqContext, $format_left); + } +} + +############################################################################################################ +# Write count of SBS by functional impact on the protein (Table 2) + Create the input for ggplot2 (pie chart with functional impact) + Create the input for ggplot2 (pie chart of SBS vs. Indels) +sub writeDistrFuncImpact +{ + my ($ws, $refH_file, $sample, $funcImpactggplot2, $overallDistrggplot2) = @_; + + # Set the row for the table 2 + my $lImpactSBS = 31; + my ($deletion, $insertion) = (0, 0); + + + $ws->write(29, 6, "Table 2. Frequency and counts of functional impact", $format_A10Boldleft); + $ws->set_column(6, 6, 13); $ws->set_column(10, 10, 15); + $ws->write(30, 6, "RefSeq gene", $table_topleft); + $ws->write(30, 7, "", $table_top); + $ws->write(30, 8, "Percent", $table_top); + $ws->write(30, 9, "Count", $table_topRight); + + + + open(IMPACTSBS, ">", $funcImpactggplot2) or die "$!: $funcImpactggplot2\n"; + print IMPACTSBS "AA_Change\tCount\tPercent\n"; + + # Pie chart with the distribution of SBS vs Indel + open(SBSINDEL, ">", $overallDistrggplot2) or die "$!: $overallDistrggplot2\n"; + print SBSINDEL "Variant_type\tCount\tPercent\n"; + + + foreach my $k_exoFunc(sort keys %{$refH_file->{$sample}{'ImpactSBS'}}) + { + my $percent = ($refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc} / $refH_file->{$sample}{'TotalMutGenomic'})*100; + $percent = sprintf("%.2f", $percent); + + if($k_exoFunc eq "NA") + { + print IMPACTSBS "Not_Applicable\t$percent\t$refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc}\n"; + } + else + { + my $temp = $k_exoFunc; + $temp =~ s/ /_/g; + print IMPACTSBS "$temp\t$percent\t$refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc}\n"; + } + + $ws->write($lImpactSBS, 6, $k_exoFunc, $table_left2); + $ws->write($lImpactSBS, 8, $percent, $format_A10); + $ws->write($lImpactSBS, 9, $refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc}, $table_right); + + $lImpactSBS++; + + # Pie chart with the distribution of SBS vs Indel + if($k_exoFunc =~ /deletion/i) { $deletion += $refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc}; } + elsif($k_exoFunc =~ /insertion/i) { $insertion += $refH_file->{$sample}{'ImpactSBS'}{$k_exoFunc}; } + } + close IMPACTSBS; + + $ws->write($lImpactSBS, 9, $refH_file->{$sample}{'TotalMutGenomic'}, $table_bottomrightHeader); + $ws->write($lImpactSBS, 6, "", $table_bottomleft); $ws->write($lImpactSBS, 7, "", $table_bottom); $ws->write($lImpactSBS, 8, "", $table_bottom); + + # Pie chart with the distribution of SBS vs Indel + my $percentSBSIndel = ($deletion/$refH_file->{$sample}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); + print SBSINDEL "Deletion\t$deletion\t$percentSBSIndel\n"; + $percentSBSIndel = ($insertion/$refH_file->{$sample}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); + print SBSINDEL "Insertion\t$insertion\t$percentSBSIndel\n"; + $percentSBSIndel = ($refH_file->{$sample}{TotalSBSGenomic}/$refH_file->{$sample}{'TotalMutGenomic'})*100; $percentSBSIndel = sprintf("%.2f", $percentSBSIndel); + print SBSINDEL "SBS\t$refH_file->{$sample}{TotalSBSGenomic}\t$percentSBSIndel\n"; + close SBSINDEL; +} + + +############################################################################################################ +# Write the result of the chi2 for the strand bias (Table 3) +sub writeChi2result +{ + my ($wb, $ws, $strandBiasggplot2, $refH_file, $k_file) = @_; + + # Define the header of the table 3 + $ws->write(28, 11, "Table 3. Significance of the strand biases", $format_A10Boldleft); + $ws->set_column(11, 11, 13); $ws->set_column(16, 16, 15); $ws->set_column(17, 17, 10); + $ws->write(29, 11, "Mutation Type", $table_topleft); $ws->write(29, 12, "Non-Tr/Tr", $table_top); $ws->write(29, 13, "Non-Tr", $table_top); $ws->write(29, 14, "Tr", $table_top); $ws->write(29, 15, "P-value", $table_top); $ws->write(29, 16, "FDR q value", $table_top); $ws->write(29, 17, "95% CI", $table_topRight); + $ws->write(39, 11, "Table 3. Significance of the strand biases", $format_A10Boldleft); + $ws->write(40, 11, "Mutation Type", $table_topleft); $ws->write(40, 12, "Non-Tr/Tr", $table_top); $ws->write(40, 13, "Non-Tr", $table_top); $ws->write(40, 14, "Tr", $table_top); $ws->write(40, 15, "P-value", $table_top); $ws->write(40, 16, "FDR q value", $table_top); $ws->write(40, 17, "95% CI", $table_topRight); + + + # Define the count on non-transcribed and transcribed strand + # C>A + my ($ca_NonTr, $ca_Tr) = (0,0); + if( ($h_chi2{$k_file}{"C>A"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"C>A"}{'Tr'} eq "NA") ) + { + $ca_NonTr = 0; + $ca_Tr = 0; + } + else + { + $ca_NonTr = $h_chi2{$k_file}{"C>A"}{'NonTr'}; + $ca_Tr = $h_chi2{$k_file}{"C>A"}{'Tr'}; + } + # C>G + my ($cg_NonTr, $cg_Tr) = (0,0); + if( ($h_chi2{$k_file}{"C>G"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"C>G"}{'Tr'} eq "NA") ) + { + $cg_NonTr = 0; + $cg_Tr = 0; + } + else + { + $cg_NonTr = $h_chi2{$k_file}{"C>G"}{'NonTr'}; + $cg_Tr = $h_chi2{$k_file}{"C>G"}{'Tr'}; + } + # C>T + my ($ct_NonTr, $ct_Tr) = (0,0); + if( ($h_chi2{$k_file}{"C>T"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"C>T"}{'Tr'} eq "NA") ) + { + $ct_NonTr = 0; + $ct_Tr = 0; + } + else + { + $ct_NonTr = $h_chi2{$k_file}{"C>T"}{'NonTr'}; + $ct_Tr = $h_chi2{$k_file}{"C>T"}{'Tr'}; + } + # T>A + my ($ta_NonTr, $ta_Tr) = (0,0); + if( ($h_chi2{$k_file}{"T>A"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"T>A"}{'Tr'} eq "NA") ) + { + $ta_NonTr = 0; + $ta_Tr = 0; + } + else + { + $ta_NonTr = $h_chi2{$k_file}{"T>A"}{'NonTr'}; + $ta_Tr = $h_chi2{$k_file}{"T>A"}{'Tr'}; + } + # T>C + my ($tc_NonTr, $tc_Tr) = (0,0); + if( ($h_chi2{$k_file}{"T>C"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"T>C"}{'Tr'} eq "NA") ) + { + $tc_NonTr = 0; + $tc_Tr = 0; + } + else + { + $tc_NonTr = $h_chi2{$k_file}{"T>C"}{'NonTr'}; + $tc_Tr = $h_chi2{$k_file}{"T>C"}{'Tr'}; + } + # T>G + my ($tg_NonTr, $tg_Tr) = (0,0); + if( ($h_chi2{$k_file}{"T>G"}{'NonTr'} eq "NA") || ($h_chi2{$k_file}{"T>G"}{'Tr'} eq "NA") ) + { + $tg_NonTr = 0; + $tg_Tr = 0; + } + else + { + $tg_NonTr = $h_chi2{$k_file}{"T>G"}{'NonTr'}; + $tg_Tr = $h_chi2{$k_file}{"T>G"}{'Tr'}; + } + + + + # Create an input for representing the strand bias with ggplot2 + open(SB, ">", $strandBiasggplot2) or die "$!: $strandBiasggplot2\n"; + print SB "Alteration\tStrand\tCount\n"; + + print SB "C>A\tNonTranscribed\t".$ca_NonTr."\n"."C>A\tTranscribed\t".$ca_Tr."\n"; + print SB "C>G\tNonTranscribed\t".$cg_NonTr."\n"."C>G\tTranscribed\t".$cg_Tr."\n"; + print SB "C>T\tNonTranscribed\t".$ct_NonTr."\n"."C>T\tTranscribed\t".$ct_Tr."\n"; + print SB "T>A\tNonTranscribed\t".$ta_NonTr."\n"."T>A\tTranscribed\t".$ta_Tr."\n"; + print SB "T>C\tNonTranscribed\t".$tc_NonTr."\n"."T>C\tTranscribed\t".$tc_Tr."\n"; + print SB "T>G\tNonTranscribed\t".$tg_NonTr."\n"."T>G\tTranscribed\t".$tg_Tr."\n"; + close SB; + + + ### Calcul the ratio for the strand bias and write it on the Excel file (Table 3) + writeRatioSB($ca_NonTr, $ca_Tr, $ws, 30, $refH_file, $k_file, "C>A", "G>T", $format_A10, $table_left, $table_middleHeader, $table_right); + writeRatioSB($cg_NonTr, $cg_Tr, $ws, 31, $refH_file, $k_file, "C>G", "G>C", $format_A10, $table_left, $table_middleHeader, $table_right); + writeRatioSB($ct_NonTr, $ct_Tr, $ws, 32, $refH_file, $k_file, "C>T", "G>A", $format_A10, $table_left, $table_middleHeader, $table_right); + writeRatioSB($ta_NonTr, $ta_Tr, $ws, 33, $refH_file, $k_file, "T>A", "A>T", $format_A10, $table_left, $table_middleHeader, $table_right); + writeRatioSB($tc_NonTr, $tc_Tr, $ws, 34, $refH_file, $k_file, "T>C", "A>G", $format_A10, $table_left, $table_middleHeader, $table_right); + writeRatioSB($tg_NonTr, $tg_Tr, $ws, 35, $refH_file, $k_file, "T>G", "A>C", $table_bottom, $table_bottomleft, $table_middleHeader2, $table_bottomRight); + + + ### Write a warning message when NonTr+Tr < 10 + my $format_italic_red = $wb->add_format(font=>'Arial', size=>10, italic=>1, color => 'red'); + + if( (($ca_NonTr+$ca_Tr)< 10) || (($cg_NonTr+$cg_Tr)< 10) || (($ct_NonTr+$ct_Tr)< 10) || (($ta_NonTr+$ta_Tr)< 10) || (($tc_NonTr+$tc_Tr)< 10) || (($tg_NonTr+$tg_Tr)< 10) ) + { + $ws->write(36, 11, "Warning message: chi-squared approximation may be incorrect because the number of SBS", $format_italic_red); + $ws->write(37, 11, "on Non-transcribed and transcribed strand is lower than 10", $format_italic_red); + } +} +# Write values in Table 3 (Sub function of writeChi2result) +sub writeRatioSB +{ + my ($count_NonTr, $count_Tr, $ws, $row, $refH_file, $k_file, $mut1, $mut2, $formatText, $formatTextMut1, $formatTextRatio, $formatTable) = @_; + + my ($ratio_mut1, $ratio_mut2, $percent_NonTr, $percent_Tr) = (0, 0, 0, 0, 0); + if( ($count_NonTr==0) || ($count_Tr==0) ) { $ratio_mut1 = 0; $ratio_mut2 = 0; $percent_NonTr = 0; $percent_Tr = 0; } + else + { + $ratio_mut1 = $count_NonTr/$count_Tr; $ratio_mut1 = sprintf("%.2f", $ratio_mut1); + $ratio_mut2 = $count_Tr/$count_NonTr; $ratio_mut2 = sprintf("%.2f", $ratio_mut2); + $percent_NonTr = ($count_NonTr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; + $percent_Tr = ($count_Tr/$refH_file->{$k_file}{'TotalSBSGenomic'})*100; + } + + # C>N and T>N + $ws->write($row, 11, $mut1, $formatTextMut1); + $ws->write($row, 12, $ratio_mut1, $formatTextRatio); + $ws->write($row, 13, $count_NonTr, $formatText); + $ws->write($row, 14, $count_Tr, $formatText); + # Write in italic and red (= warning message) when the count of NonTr + Tr is lower than 10 + if( ($count_NonTr+$count_Tr) < 10 ) + { + if($mut1 eq "T>G") + { + if(! exists $h_chi2{$k_file}{$mut1}{'p-value'}) { $ws->write_string($row, 15, ""); } + elsif($h_chi2{$k_file}{$mut1}{'p-value'} eq "NA") { $ws->write_string($row, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $table_bottom); } + else { $ws->write_string($row, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $table_bottomItalicRed); } + } + else + { + if(! exists $h_chi2{$k_file}{$mut1}{'p-value'}) { $ws->write_string($row, 15, ""); } + elsif($h_chi2{$k_file}{$mut1}{'p-value'} eq "NA") { $ws->write_string($row, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $formatText); } + else { $ws->write_string($row, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $format_A10ItalicRed); } + } + } + else + { + $ws->write_string($row, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $formatText); + } + + $ws->write($row, 16, $h_chi2{$k_file}{$mut1}{'FDR'}, $formatText); + $ws->write($row, 17, $h_chi2{$k_file}{$mut1}{'ConfInt'}, $formatTable); + + + # G>N and A>N + $ws->write($row+11, 11, $mut2, $formatTextMut1); + $ws->write($row+11, 12, $ratio_mut2, $formatTextRatio); + $ws->write($row+11, 13, $count_Tr, $formatText); + $ws->write($row+11, 14, $count_NonTr, $formatText); + if( ($count_NonTr+$count_Tr) < 10 ) + { + if($mut1 eq "T>G") + { + if(! exists $h_chi2{$k_file}{$mut1}{'p-value'}) { $ws->write_string($row+11, 15, ""); } + elsif($h_chi2{$k_file}{$mut1}{'p-value'} eq "NA") { $ws->write_string($row+11, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $table_bottom); } + else { $ws->write_string($row+11, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $table_bottomItalicRed); } + } + else + { + if(! exists $h_chi2{$k_file}{$mut1}{'p-value'}) { $ws->write_string($row+11, 15, ""); } + elsif($h_chi2{$k_file}{$mut1}{'p-value'} eq "NA") { $ws->write_string($row+11, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $formatText); } + else { $ws->write_string($row+11, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $format_A10ItalicRed); } + } + } + else + { + $ws->write_string($row+11, 15, $h_chi2{$k_file}{$mut1}{'p-value'}, $formatText); + } + + $ws->write($row+11, 16, $h_chi2{$k_file}{$mut1}{'FDR'}, $formatText); + $ws->write($row+11, 17, $h_chi2{$k_file}{$mut1}{'ConfInt'}, $formatTable); +} + + +############################################################################################################ +# SBS distribution by functional region (Table 4) & Strand bias by functional region (Table 5) & Overall count and percent of SBS (Table 1) +sub writeStatbyFuncRegion +{ + my ($refH_file, $sample, $ws, $rowStart_SBSdistrBySeg, $colStart_SBSdistrBySeg, $nb_func, $ref_RowSBSDistrBySegAndFuncCG, $mutDistrggplot2) = @_; + + my $row_SBSdistrBySeg = $rowStart_SBSdistrBySeg+4; + my $row_SBSDistrBySegAndFunc_CA = $rowStart_SBSdistrBySeg + $nb_func + 12; + my $rowEndCG_SBSDistrBySegAndFunc_CG = $$ref_RowSBSDistrBySegAndFuncCG + $nb_func; + my $row_SBSDistrBySegAndFunc_CT = $rowStart_SBSdistrBySeg + ($nb_func*3) + 20; + my $colTable4 = 0; + + my ($count_ca, $count_cg, $count_ct, $count_ta, $count_tc, $count_tg) = (0,0,0,0,0,0); + + + ## 6 mutation types by segment + foreach my $k_func (sort keys %{$refH_file->{$sample}{'6mutType'}}) + { + my $totalSBS_bySegment = 0; + + # Write the functional region for the section SBS distribution by segment (Table 4) + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg, $k_func, $formatT_left); + # Write the exonic func for the section strand bias by segment (Table 5) + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg, $k_func, $formatT_left); + # Write the last functional element in the table + if($$ref_RowSBSDistrBySegAndFuncCG == $rowEndCG_SBSDistrBySegAndFunc_CG) + { + $ws->write($$ref_RowSBSDistrBySegAndFuncCG, $colStart_SBSdistrBySeg, $k_func, $formatT_bottomLeft); + } + else + { + $ws->write($$ref_RowSBSDistrBySegAndFuncCG, $colStart_SBSdistrBySeg, $k_func, $formatT_left); + } + + foreach my $k_mutation (sort keys %{$refH_file->{$sample}{'6mutType'}{$k_func}}) + { + ### Write the count of SBS per mutation on genomic (Table 4) and coding strand (Table 5) + if($k_mutation eq "C:G>A:T") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+3, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_ca); + } + if($k_mutation eq "C:G>G:C") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+4, $colStart_SBSdistrBySeg+5, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_cg); + } + if($k_mutation eq "C:G>T:A") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+8, $colStart_SBSdistrBySeg+7, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_ct); + } + if($k_mutation eq "T:A>A:T") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $$ref_RowSBSDistrBySegAndFuncCG, $colStart_SBSdistrBySeg, $colStart_SBSdistrBySeg+9, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_ta); + } + if($k_mutation eq "T:A>C:G") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $$ref_RowSBSDistrBySegAndFuncCG, $colStart_SBSdistrBySeg+4, $colStart_SBSdistrBySeg+11, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_tc); + } + if($k_mutation eq "T:A>G:C") + { + writeCountSBS($refH_file, $sample, $k_func, $k_mutation, $ws, $$ref_RowSBSDistrBySegAndFuncCG, $colStart_SBSdistrBySeg+8, $colStart_SBSdistrBySeg+13, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, \$count_tg); + } + + # Calculate the total number of SBS on the genomic strand for each mutation types by exonic region + $totalSBS_bySegment += $refH_file->{$sample}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } # End $k_mutation + + $row_SBSDistrBySegAndFunc_CA++; $$ref_RowSBSDistrBySegAndFuncCG++; #$row_SBSDistrBySegAndFunc_CT++; + + # Write the percent by exonic region + writePercentSBS($refH_file, $sample, $k_func, "C:G>A:T", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+2, $ws, $totalSBS_bySegment); + writePercentSBS($refH_file, $sample, $k_func, "C:G>G:C", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+4, $ws, $totalSBS_bySegment); + writePercentSBS($refH_file, $sample, $k_func, "C:G>T:A", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+6, $ws, $totalSBS_bySegment); + writePercentSBS($refH_file, $sample, $k_func, "T:A>A:T", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+8, $ws, $totalSBS_bySegment); + writePercentSBS($refH_file, $sample, $k_func, "T:A>C:G", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+10, $ws, $totalSBS_bySegment); + writePercentSBS($refH_file, $sample, $k_func, "T:A>G:C", $row_SBSdistrBySeg, $colStart_SBSdistrBySeg+12, $ws, $totalSBS_bySegment); + + # Write the count of SBS by segment + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+1, $totalSBS_bySegment, $format_A10); + + $row_SBSdistrBySeg++; + } # End $k_func + + + # Write the total number of SBS on the genomic strand (Table 4) + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+1, $refH_file->{$sample}{'TotalSBSGenomic'}, $formatT_bottomHeader); + + + ##### Calculate the total percentages by mutation type + my $percent_ca = ($count_ca / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_ca = sprintf("%.2f", $percent_ca); + my $percent_cg = ($count_cg / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_cg = sprintf("%.2f", $percent_cg); + my $percent_ct = ($count_ct / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_ct = sprintf("%.2f", $percent_ct); + my $percent_ta = ($count_ta / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_ta = sprintf("%.2f", $percent_ta); + my $percent_tc = ($count_tc / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_tc = sprintf("%.2f", $percent_tc); + my $percent_tg = ($count_tg / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; $percent_tg = sprintf("%.2f", $percent_tg); + + + # Write the total percentage (Table 4) + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+2, $percent_ca, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+3, $count_ca, $formatT_bottomHeader); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+4, $percent_cg, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+5, $count_cg, $formatT_bottomHeader); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+6, $percent_ct, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+7, $count_ct, $formatT_bottomHeader); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+8, $percent_ta, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+9, $count_ta, $formatT_bottomHeader); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+10, $percent_tc, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+11, $count_tc, $formatT_bottomHeader); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+12, $percent_tg, $formatT_bottom); + $ws->write($row_SBSdistrBySeg, $colStart_SBSdistrBySeg+13, $count_tg, $formatT_bottomRightHeader); + + + + # Overall distribution of SBS (Table 1) + $ws->write(0, 0, "Graph 1. SBS distribution", $formatT_graphTitle); $ws->set_row(0, 18); + $ws->write(29, 0, "Table 1. Frequency and counts of all SBS", $format_A10Boldleft); + $ws->write(30, 0, "Mutation type", $table_topleft); + $ws->write(30, 1, "Percentage", $table_top); + $ws->write(30, 2, "Count", $table_topRight); + $ws->write(31, 0, "C:G>A:T", $table_left); $ws->write(31, 1, $percent_ca, $format_A10); $ws->write(31, 2, $count_ca, $table_right); + $ws->write(32, 0, "C:G>G:C", $table_left); $ws->write(32, 1, $percent_cg, $format_A10); $ws->write(32, 2, $count_cg, $table_right); + $ws->write(33, 0, "C:G>T:A", $table_left); $ws->write(33, 1, $percent_ct, $format_A10); $ws->write(33, 2, $count_ct, $table_right); + $ws->write(34, 0, "T:A>A:T", $table_left); $ws->write(34, 1, $percent_ta, $format_A10); $ws->write(34, 2, $count_ta, $table_right); + $ws->write(35, 0, "T:A>C:G", $table_left); $ws->write(35, 1, $percent_tc, $format_A10); $ws->write(35, 2, $count_tc, $table_right); + $ws->write(36, 0, "T:A>G:C", $table_left); $ws->write(36, 1, $percent_tg, $format_A10); $ws->write(36, 2, $count_tg, $table_right); + $ws->write(37, 0, "", $table_bottomleft); $ws->write(37, 1, "", $table_bottom); $ws->write(37, 2, $refH_file->{$sample}{'TotalSBSGenomic'}, $table_bottomrightHeader); + + # Create an input for ggplot2 for representing the distribution of SBS for each mutation types (Figure 1) + open(DISTRSBS, ">", $mutDistrggplot2) or die "$!: $mutDistrggplot2\n"; + print DISTRSBS "Mutation_Type\tCount\tPercentage\tSample\n"; + print DISTRSBS "C:G>A:T\t$count_ca\t$percent_ca\t$sample\n"; + print DISTRSBS "C:G>G:C\t$count_cg\t$percent_cg\t$sample\n"; + print DISTRSBS "C:G>T:A\t$count_ct\t$percent_ct\t$sample\n"; + print DISTRSBS "T:A>A:T\t$count_ta\t$percent_ta\t$sample\n"; + print DISTRSBS "T:A>C:G\t$count_tc\t$percent_tc\t$sample\n"; + print DISTRSBS "T:A>G:C\t$count_tg\t$percent_tg\t$sample\n"; + close DISTRSBS; +} +# Write the percentage in table 4 of the Excel report (Sub function of writeStatbyFuncRegion) +sub writePercentSBS +{ + my ($refH_file, $k_file, $k_func, $mutation, $row, $col, $ws, $totalSBS) = @_; + + my $percent = 0; + if($refH_file->{$k_file}{'6mutType'}{$k_func}{$mutation}{'TotalMutG'} == 0) + { + $percent = 0; + } + else + { + $percent = ($refH_file->{$k_file}{'6mutType'}{$k_func}{$mutation}{'TotalMutG'} / $totalSBS ) * 100; + $percent = sprintf("%.2f", $percent); + } + $ws->write($row, $col, $percent, $format_A10); +} +# Write the count in table 4 and table 5 of the Excel report (Sub function of writeStatbyFuncRegion) +sub writeCountSBS +{ + my ($refH_file, $k_file, $k_func, $k_mutation, $ws, $row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg, $colTable4, $row_SBSdistrBySeg, $rowEndCG_SBSDistrBySegAndFunc_CG, $refCount) = @_; + + my $ratioSB = 0; + if( ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} == 0) || ($refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'} == 0) ) + { + $ratioSB = 0; + } + else + { + $ratioSB = $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'} / $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}; + } + $ratioSB = sprintf("%.2f", $ratioSB); + + + if($row_SBSDistrBySegAndFunc_CA == $rowEndCG_SBSDistrBySegAndFunc_CG) + { + # Write the ratio of NonTr / Tr (Table 5) + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+1, $ratioSB, $formatT_bottom); + # Write the count of SBS in the NonTr and Tr strand + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $formatT_bottom); + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_bottom); + + if( ($k_mutation eq "C:G>T:A") || ($k_mutation eq "T:A>G:C") ) + { + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_bottomRight); + } + } + else + { + # Write the ratio of NonTr / Tr (Table 5) + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+1, $ratioSB, $format_A10); + # Write the count of SBS in the NonTr and Tr strand + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+2, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'NonTr'}, $format_A10); + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $format_A10); + + if( ($k_mutation eq "C:G>T:A") || ($k_mutation eq "T:A>G:C") ) + { + $ws->write($row_SBSDistrBySegAndFunc_CA, $colStart_SBSdistrBySeg+3, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'Tr'}, $formatT_right); + } + } + + if($k_mutation eq "C:G>A:T") + { + # Calculate the total number of SBS per mut type (genomic strand) + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + elsif($k_mutation eq "C:G>G:C") + { + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + elsif($k_mutation eq "C:G>T:A") + { + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + elsif($k_mutation eq "T:A>A:T") + { + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + elsif($k_mutation eq "T:A>C:G") + { + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + elsif($k_mutation eq "T:A>G:C") + { + $$refCount += $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}; + } + + + # Write the count by exonic region (Table 4) + $ws->write($row_SBSdistrBySeg, $colTable4, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $format_A10); + + if($k_mutation eq "T:A>G:C") + { + $ws->write($row_SBSdistrBySeg, $colTable4, $refH_file->{$k_file}{'6mutType'}{$k_func}{$k_mutation}{'TotalMutG'}, $formatT_right); + } +} + + +############################################################################################################ +# SBS distribution by chromosomes (Table 6) +sub writeDistrByChr +{ + my ($ws, $refH_file, $sample, $row, $col, $distrByChrggplot2) = @_; + + + # For the HTML report + open(SBSPerChr, ">", $distrByChrggplot2) or die "$!: $distrByChrggplot2\n"; + print SBSPerChr "\tPearson\t$refH_file->{$sample}{'SBSPerChr'}{'AllMutType'}\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>A:T"}{'Pearson'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>G:C"}{'Pearson'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>T:A"}{'Pearson'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>A:T"}{'Pearson'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>C:G"}{'Pearson'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>G:C"}{'Pearson'},"\n"; + print SBSPerChr "Chr\tSize\tAll SBS\tC:G>A:T\tC:G>G:C\tC:G>T:A\tT:A>A:T\tT:A>C:G\tT:A>G:C\n"; + + + my $row_SBSPerChr = $row + 8; + + + # Write the Pearson coefficient + $ws->write($row+6, $col+3, $refH_file->{$sample}{'SBSPerChr'}{"C:G>A:T"}{'Pearson'}, $format_A10); + $ws->write($row+6, $col+4, $refH_file->{$sample}{'SBSPerChr'}{"C:G>G:C"}{'Pearson'}, $format_A10); + $ws->write($row+6, $col+5, $refH_file->{$sample}{'SBSPerChr'}{"C:G>T:A"}{'Pearson'}, $format_A10); + $ws->write($row+6, $col+6, $refH_file->{$sample}{'SBSPerChr'}{"T:A>A:T"}{'Pearson'}, $format_A10); + $ws->write($row+6, $col+7, $refH_file->{$sample}{'SBSPerChr'}{"T:A>C:G"}{'Pearson'}, $format_A10); + $ws->write($row+6, $col+8, $refH_file->{$sample}{'SBSPerChr'}{"T:A>G:C"}{'Pearson'}, $formatT_right); + + # Write the chromosome number and their sizes / Write count SBS per chromosomes + my $line=0; + + foreach my $chromosome (sort keys %chromosomes) + { + $ws->write($row_SBSPerChr+($line), $col, $chromosome, $formatT_left); + $ws->write($row_SBSPerChr+($line), $col+1, $chromosomes{$chromosome}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+2, $refH_file->{$sample}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'}, $format_A10); + + # Write the count per mutation + $ws->write($row_SBSPerChr+($line), $col+3, $refH_file->{$sample}{'SBSPerChr'}{"C:G>A:T"}{'CHR'}{$chromosome}{'chr'}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+4, $refH_file->{$sample}{'SBSPerChr'}{"C:G>G:C"}{'CHR'}{$chromosome}{'chr'}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+5, $refH_file->{$sample}{'SBSPerChr'}{"C:G>T:A"}{'CHR'}{$chromosome}{'chr'}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+6, $refH_file->{$sample}{'SBSPerChr'}{"T:A>A:T"}{'CHR'}{$chromosome}{'chr'}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+7, $refH_file->{$sample}{'SBSPerChr'}{"T:A>C:G"}{'CHR'}{$chromosome}{'chr'}, $format_A10); + $ws->write($row_SBSPerChr+($line), $col+8, $refH_file->{$sample}{'SBSPerChr'}{"T:A>G:C"}{'CHR'}{$chromosome}{'chr'}, $formatT_right); + + + # For the HTML report + print SBSPerChr "$chromosome\t", $chromosomes{$chromosome},"\t", $refH_file->{$sample}{'SBSPerChr'}{'TotalPerChr'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>A:T"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>G:C"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"C:G>T:A"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>A:T"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>C:G"}{'CHR'}{$chromosome}{'chr'},"\t", $refH_file->{$sample}{'SBSPerChr'}{"T:A>G:C"}{'CHR'}{$chromosome}{'chr'},"\n"; + + $line++; + } + + # Write the Pearson coefficient for the total number of SBS + $ws->write($row+6, $col+2, $refH_file->{$sample}{'SBSPerChr'}{'AllMutType'}, $format_A10); + $ws->write($row_SBSPerChr+(keys %chromosomes), $col+2, $refH_file->{$sample}{'TotalSBSGenomic'}, $formatT_bottomHeader); + + print SBSPerChr "\t\t$refH_file->{$sample}{'TotalSBSGenomic'}\n"; + close SBSPerChr; +} + + +############################################################################################################ +# Trinucleotide sequence context on genomic strand (Panel 1) +sub writeTriNtGenomic +{ + my ($ws, $refH_file, $sample, $col, $heatmapCountggplot2, $heatmapPercentggplot2, $triNtBarChartggplot2, $ref_c_ca6_g, $ref_c_cg6_g, $ref_c_ct6_g, $ref_c_ta6_g, $ref_c_tc6_g, $ref_c_tg6_g) = @_; + + # Initialise the row of the panel 1 + my $row_SeqContext6 = 4; + # Percent total of mutations for 6 mutation types on genomic strand + my ($p_ca6_g, $p_cg6_g, $p_ct6_g, $p_ta6_g, $p_tc6_g, $p_tg6_g) = (0,0,0, 0,0,0); + my $maxValue = 0; # For the heatmap + + # For checking if the total number of SBS is correct + my $total_SBS_genomic = 0; + + + open(HEATMAPCGENOMIC, ">", $heatmapCountggplot2) or die "$!: $heatmapCountggplot2\n"; + print HEATMAPCGENOMIC "\tC>A\tC>G\tC>T\tT>A\tT>C\tT>G\n"; + open(HEATMAPPGENOMIC, ">", $heatmapPercentggplot2) or die "$!: $heatmapPercentggplot2\n"; + print HEATMAPPGENOMIC "\tC>A\tC>G\tC>T\tT>A\tT>C\tT>G\n"; + + ## Bar plot NMF like + open(BARPLOTNMFLIKE, ">", $triNtBarChartggplot2) or die "$!: $triNtBarChartggplot2\n"; + print BARPLOTNMFLIKE "alteration\tcontext\tvalue\n"; + + + foreach my $k_context (sort keys %{$refH_file->{$sample}{'SeqContextG'}}) + { + if( ($k_context =~ /N/) || (length($k_context) != 3) ) { next; } + + # Write the context: 6 mut type on genomic strand + $ws->write($row_SeqContext6 , $col+3, $k_context, $format_A10); + $ws->write($row_SeqContext6 , $col+13, $k_context, $format_A10); + + # Count for the heatmap + print HEATMAPCGENOMIC $k_context."\t"; + print HEATMAPPGENOMIC $k_context."\t"; + + foreach my $k_mutation (sort keys %{$refH_file->{$sample}{'SeqContextG'}{$k_context}}) + { + # For checking the total number of SBS + $total_SBS_genomic += $refH_file->{$sample}{'SeqContextG'}{$k_context}{$k_mutation}; + + # Calculate the percentages + my $percent = 0; + if($refH_file->{$sample}{'SeqContextG'}{$k_context}{$k_mutation} == 0) { $percent = 0; } + else + { + $percent = ($refH_file->{$sample}{'SeqContextG'}{$k_context}{$k_mutation} / $refH_file->{$sample}{'TotalSBSGenomic'}) * 100; + $percent = sprintf("%.2f", $percent); + } + + # For representing the sequence context with a bar plot (NMF like style) + print BARPLOTNMFLIKE $k_mutation,"\t", $k_context,"\t", $percent,"\n"; + + # Write the count for the heatmap + print HEATMAPCGENOMIC $refH_file->{$sample}{'SeqContextG'}{$k_context}{$k_mutation}."\t"; + print HEATMAPPGENOMIC "$percent\t"; + + + # For NMF input + my $count = $refH_file->{$sample}{'SeqContextG'}{$k_context}{$k_mutation}; + if($sample ne "Pool_Data") { push(@{$h_inputNMF{'Count'}{$k_context}{$k_mutation}}, $count); } + if($sample ne "Pool_Data") { push(@{$h_inputNMF{'Percent'}{$k_context}{$k_mutation}}, $percent); } + + + if($k_mutation eq "C>A") + { + triNtByMut($ws, $row_SeqContext6, $col+4, $col+14, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_ca6_g, \$p_ca6_g); + } + elsif($k_mutation eq "C>G") + { + triNtByMut($ws, $row_SeqContext6, $col+5, $col+15, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_cg6_g, \$p_cg6_g); + } + elsif($k_mutation eq "C>T") + { + triNtByMut($ws, $row_SeqContext6, $col+6, $col+16, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_ct6_g, \$p_ct6_g); + } + elsif($k_mutation eq "T>A") + { + triNtByMut($ws, $row_SeqContext6, $col+7, $col+17, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_ta6_g, \$p_ta6_g); + } + elsif($k_mutation eq "T>C") + { + triNtByMut($ws, $row_SeqContext6, $col+8, $col+18, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_tc6_g, \$p_tc6_g); + } + elsif($k_mutation eq "T>G") + { + triNtByMut($ws, $row_SeqContext6, $col+9, $col+19, $refH_file, $sample, $k_context, $k_mutation, $percent, $maxValue, $ref_c_tg6_g, \$p_tg6_g); + } + else + { + print STDERR "Error: Mutation type not considered for: $k_mutation\n"; + exit; + } + } + $row_SeqContext6++; + + print HEATMAPCGENOMIC "\n"; + print HEATMAPPGENOMIC "\n"; + } + close HEATMAPCGENOMIC; close HEATMAPPGENOMIC; + close BARPLOTNMFLIKE; + + + # Write the total number of SBS per mutation type: COUNT + $ws->write($row_SeqContext6, $col+4, $$ref_c_ca6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+5, $$ref_c_cg6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+6, $$ref_c_ct6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+7, $$ref_c_ta6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+8, $$ref_c_tc6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+9, $$ref_c_tg6_g, $formatT_bottomHeader2); + if($total_SBS_genomic != $refH_file->{$sample}{'TotalSBSGenomic'}) + { + print STDERR "Error in the calculation of the total number of SBS on the genomic strand!!!!\n"; + print STDERR "From hash table $refH_file->{$sample}{'TotalSBSGenomic'}\tVS\t$total_SBS_genomic\n"; + exit; + } + + # Write the total number of SBS per mutation type: PERCENT + $ws->write($row_SeqContext6, $col+14, $p_ca6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+15, $p_cg6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+16, $p_ct6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+17, $p_ta6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+18, $p_tc6_g, $formatT_bottomHeader2); + $ws->write($row_SeqContext6, $col+19, $p_tg6_g, $formatT_bottomHeader2); + + my $totalPercent_genomic = $p_ca6_g + $p_cg6_g + $p_ct6_g + $p_ta6_g + $p_tc6_g + $p_tg6_g; + $totalPercent_genomic = sprintf("%.0f", $totalPercent_genomic); + + if($totalPercent_genomic != 100) + { + print STDERR "Error in the calculation of the total percentages on the genomic strand!!!\n"; + print STDERR "The total is equal to=\t$totalPercent_genomic\n"; + exit; + } +} +# Trinucleotide count and percentage by mutation type (Sub function of writeTriNtGenomic) +sub triNtByMut +{ + my ($ws, $row, $colC, $colP, $refH_file, $sample, $context, $mutation, $percent, $maxValue, $refCountG, $refPercentG) = @_; + + ### COUNT + $ws->write($row, $colC, $refH_file->{$sample}{'SeqContextG'}{$context}{$mutation}, $format_A10); + + ### PERCENTAGE + $ws->write($row, $colP, $percent, $format_A10); + + # For the heatmap + if($percent >= $maxValue) { $maxValue = $percent; } + + # For the total amount per mutation types + $$refCountG += $refH_file->{$sample}{'SeqContextG'}{$context}{$mutation}; + $$refPercentG += $percent; +} + + +############################################################################################################ +# Trinucleotide sequence context on coding strand (Panel 2) +sub writeTriNtCoding +{ + my ($ws, $row, $col, $refH_file, $sample, $triNtBarChartCodingCountggplot2, $triNtBarChartCodingPercentggplot2) = @_; + + # Initialise the row + my $row_SeqContext12 = $row+6; + my $row_SeqContext12Percent = $row+27; + + # Total count and percent calculated for the strand bias + my ($ca_NonTr, $ca_Tr, $cg_NonTr, $cg_Tr, $ct_NonTr, $ct_Tr, $ta_NonTr, $ta_Tr, $tc_NonTr, $tc_Tr, $tg_NonTr, $tg_Tr) = (0,0,0, 0,0,0, 0,0,0, 0,0,0); + my ($percent_ca_NonTr, $percent_ca_Tr, $percent_cg_NonTr, $percent_cg_Tr, $percent_ct_NonTr, $percent_ct_Tr, $percent_ta_NonTr, $percent_ta_Tr, $percent_tc_NonTr, $percent_tc_Tr, $percent_tg_NonTr, $percent_tg_Tr) = (0,0,0, 0,0,0, 0,0,0, 0,0,0); + + # For checking if the total number of SBS is correct + my $total_SBS_coding = 0; + + + open(COUNT, ">", $triNtBarChartCodingCountggplot2) or die "$!: $triNtBarChartCodingCountggplot2\n"; + print COUNT "MutationTypeContext\tStrand\tValue\tSample\n"; + open(PERCENT, ">", $triNtBarChartCodingPercentggplot2) or die "$!: $triNtBarChartCodingPercentggplot2\n"; + print PERCENT "MutationTypeContext\tStrand\tValue\tSample\n"; + + foreach my $k_context (sort keys %{$refH_file->{$sample}{'SeqContextC'}}) + { + if( ($k_context =~ /N/) || (length($k_context) != 3) ) { next; } + + # Write the context: 12 mut type on coding strand + $ws->write($row_SeqContext12 , $col, $k_context, $formatT_left); + $ws->write($row_SeqContext12Percent , $col, $k_context, $formatT_left); + + foreach my $k_mutation (sort keys %{$refH_file->{$sample}{'SeqContextC'}{$k_context}}) + { + # Percent: 12 mut type on coding strand + my ($percent_NonTr, $percent_Tr) = (0, 0); + + if($refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} != 0) + { + $percent_NonTr = ( $refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} / $refH_file->{$sample}{'TotalSBSCoding'} ) * 100; + } + + if($refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'} != 0) + { + $percent_Tr = ( $refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'} / $refH_file->{$sample}{'TotalSBSCoding'} ) * 100; + } + + + # Counts + print COUNT "$k_mutation:$k_context\tNonTranscribed\t$refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'}\t$sample\n"; + print COUNT "$k_mutation:$k_context\tTranscribed\t$refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}\t$sample\n"; + + # Percentages + $percent_NonTr = sprintf("%.2f", $percent_NonTr); + $percent_Tr = sprintf("%.2f", $percent_Tr); + print PERCENT "$k_mutation:$k_context\tNonTranscribed\t$percent_NonTr\t$sample\n"; + print PERCENT "$k_mutation:$k_context\tTranscribed\t$percent_Tr\t$sample\n"; + + # Calculate the total number for each mutation types + if($k_mutation eq "C>A") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+1, $row_SeqContext12Percent, \$ca_NonTr, \$ca_Tr, $percent_NonTr, $percent_Tr); + + $percent_ca_NonTr += $percent_NonTr; + $percent_ca_Tr += $percent_Tr; + } + if($k_mutation eq "C>G") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+3, $row_SeqContext12Percent, \$cg_NonTr, \$cg_Tr, $percent_NonTr, $percent_Tr); + + $percent_cg_NonTr += $percent_NonTr; + $percent_cg_Tr += $percent_Tr; + } + if($k_mutation eq "C>T") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+5, $row_SeqContext12Percent, \$ct_NonTr, \$ct_Tr, $percent_NonTr, $percent_Tr); + + $percent_ct_NonTr += $percent_NonTr; + $percent_ct_Tr += $percent_Tr; + } + if($k_mutation eq "T>A") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+7, $row_SeqContext12Percent, \$ta_NonTr, \$ta_Tr, $percent_NonTr, $percent_Tr); + + $percent_ta_NonTr += $percent_NonTr; + $percent_ta_Tr += $percent_Tr; + } + if($k_mutation eq "T>C") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+9, $row_SeqContext12Percent, \$tc_NonTr, \$tc_Tr, $percent_NonTr, $percent_Tr); + + $percent_tc_NonTr += $percent_NonTr; + $percent_tc_Tr += $percent_Tr; + } + if($k_mutation eq "T>G") + { + triNtByMutCoding($refH_file, $sample, $k_context, $k_mutation, $ws, $row_SeqContext12, $col+11, $row_SeqContext12Percent, \$tg_NonTr, \$tg_Tr, $percent_NonTr, $percent_Tr); + + $percent_tg_NonTr += $percent_NonTr; + $percent_tg_Tr += $percent_Tr; + } + + # For checking if the total number of SBS is correct + $total_SBS_coding += $refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'NonTr'} + $refH_file->{$sample}{'SeqContextC'}{$k_context}{$k_mutation}{'Tr'}; + } + $row_SeqContext12++; $row_SeqContext12Percent++; + } + close COUNT; close PERCENT; + + + ## Write the total of each mutation types: 12 mut type on coding strand + $ws->write($row_SeqContext12, $col+1, $ca_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+2, $ca_Tr, $formatT_bottomHeader2); + $ws->write($row_SeqContext12, $col+3, $cg_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+4, $cg_Tr, $formatT_bottomHeader2); + $ws->write($row_SeqContext12, $col+5, $ct_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+6, $ct_Tr, $formatT_bottomHeader2); + $ws->write($row_SeqContext12, $col+7, $ta_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+8, $ta_Tr, $formatT_bottomHeader2); + $ws->write($row_SeqContext12, $col+9, $tc_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+10, $tc_Tr, $formatT_bottomHeader2); + $ws->write($row_SeqContext12, $col+11, $tg_NonTr, $formatT_bottomHeader2); $ws->write($row_SeqContext12, $col+12, $tg_Tr, $formatT_bottomHeader2); + # Write the total percentages of each mutation types: 12 mut type on coding strand + $ws->write($row_SeqContext12Percent, $col+1, $percent_ca_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+2, $percent_ca_Tr, $formatT_bottomHeader); + $ws->write($row_SeqContext12Percent, $col+3, $percent_cg_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+4, $percent_cg_Tr, $formatT_bottomHeader); + $ws->write($row_SeqContext12Percent, $col+5, $percent_ct_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+6, $percent_ct_Tr, $formatT_bottomHeader); + $ws->write($row_SeqContext12Percent, $col+7, $percent_ta_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+8, $percent_ta_Tr, $formatT_bottomHeader); + $ws->write($row_SeqContext12Percent, $col+9, $percent_tc_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+10, $percent_tc_Tr, $formatT_bottomHeader); + $ws->write($row_SeqContext12Percent, $col+11, $percent_tg_NonTr, $formatT_bottomHeader); $ws->write($row_SeqContext12Percent, $col+12, $percent_tg_Tr, $formatT_bottomHeader); + + if($total_SBS_coding == $refH_file->{$sample}{'TotalSBSCoding'}) + { + $ws->write($row_SeqContext12, $col+13, $refH_file->{$sample}{'TotalSBSCoding'}, $formatT_bottomHeader2) + } + else + { + print STDERR "Error: in the calculation of the total number of SBS on the coding strand!!!!\n"; + print STDERR "From hash table $refH_file->{$sample}{'TotalSBSCoding'}\tVS\t$total_SBS_coding\n"; + exit; + } + + my $totalP_SBS_coding = $percent_ca_NonTr + $percent_ca_Tr + $percent_cg_NonTr + $percent_cg_Tr + $percent_ct_NonTr + $percent_ct_Tr + $percent_ta_NonTr + $percent_ta_Tr + $percent_tc_NonTr + $percent_tc_Tr + $percent_tg_NonTr + $percent_tg_Tr; + $totalP_SBS_coding = sprintf("%.0f", $totalP_SBS_coding); + + if($totalP_SBS_coding != 100) + { + print STDERR "Error: The percentages for the trinucleotide sequence context on the coding strand for 12 mutation types is not equal to 100!!!\n$totalP_SBS_coding\n"; + exit; + } +} +# Trinucleotide count and percentage by mutation type on Coding strand (Sub function of writeTriNtCoding) +sub triNtByMutCoding +{ + my ($refH_file, $sample, $context, $mutation, $ws, $row, $col, $rowP, $refNonTr, $refTr, $percent_NonTr, $percent_Tr) = @_; + + $$refNonTr += $refH_file->{$sample}{'SeqContextC'}{$context}{$mutation}{'NonTr'}; + $$refTr += $refH_file->{$sample}{'SeqContextC'}{$context}{$mutation}{'Tr'}; + + # COUNT : 12 mutation type (stranded bar graph) + $ws->write($row, $col, $refH_file->{$sample}{'SeqContextC'}{$context}{$mutation}{'NonTr'}, $format_A10); + $ws->write($row, $col+1, $refH_file->{$sample}{'SeqContextC'}{$context}{$mutation}{'Tr'}, $format_A10); + + + ## PERCENT : 12 mutation type (stranded bar graph) + $ws->write($rowP, $col, $percent_NonTr, $format_A10); + $ws->write($rowP, $col+1, $percent_Tr, $format_A10); +} + + +############################################################################################################ +# Create and write the figures on the Excel report +sub createWriteFigs +{ + my ($ws, $row, $col, $folderFigure, $sample, $c_ca6_g, $c_cg6_g, $c_ct6_g, $c_ta6_g, $c_tc6_g, $c_tg6_g) = @_; + + ######## Create figures + # Bar char for SBS distribution (Figure 1) + # Pie char for Impact on protein sequence (Figure 2) + # Stranded distribution of SBS (Figure 3) + # Heatmaps for trinucleotide context + `Rscript $pathRScriptFigs --folderFigure $folderFigure --folderTemp $folder_temp --filename $sample`; + + # Bar chart for trinucleotide context on coding strand + `Rscript $pathRScriptTxnSB $folderFigure/Stranded_Analysis/$sample/$sample-StrandedSignatureCount.txt $folderFigure/Stranded_Analysis/$sample/$sample-StrandedSignatureCount $folder_temp/$sample-StrandedSignatureCount Count`; + + `Rscript $pathRScriptTxnSB $folderFigure/Stranded_Analysis/$sample/$sample-StrandedSignaturePercent.txt $folderFigure/Stranded_Analysis/$sample/$sample-StrandedSignaturePercent $folder_temp/$sample-StrandedSignaturePercent Percent`; + + # Bar plot for representing the sequence context (NMF like style) + `Rscript $pathRScriptMutSpectrum $folderFigure/Trinucleotide_Sequence_Context/$sample/$sample-MutationSpectraPercent-Genomic.txt $sample $folderFigure/Trinucleotide_Sequence_Context/$sample $folder_temp $c_ca6_g $c_cg6_g $c_ct6_g $c_ta6_g $c_tc6_g $c_tg6_g`; + + + ######## Write the figures in the Excel report + # Bar char for SBS distribution (Figure 1) + $ws->insert_image(1, 0, "$folder_temp/$sample-SBS_distribution-Report.png", 0, 0, .2, .2); + + # Impact of the SBS on the protein (Figure 2) + $ws->write(0, 6, "Graph 2. Impact on protein sequence", $formatT_graphTitle); + $ws->insert_image(1, 6, "$folder_temp/$sample-DistributionExoFunc-Report.png", 0, 0, .2, .2); + + # Stranded distribution of SBS (Figure 3) + $ws->write(0, 11, "Graph 3. Stranded distribution of SBS", $formatT_graphTitle); + $ws->insert_image(1, 11, "$folder_temp/$sample-StrandBias-Report.png", 0, 0, .2, .2); + + ## Trinucleotide context on coding strand (Scale the inserted image: width x 0.7, height x 0.8) + $ws->insert_image($row+3, $col+15, "$folder_temp/$sample-StrandedSignatureCount-Report.png", 0, 0, .16, .16); + $ws->insert_image($row+24, $col+15, "$folder_temp/$sample-StrandedSignaturePercent-Report.png", 0, 0, .16, .16); + + # Heatamp for the sequence context on the genomic strand (6 mutation types) + $ws->insert_image(4, $col, "$folder_temp/$sample-HeatmapCount-Genomic-Report.png"); + $ws->insert_image(4, $col+10, "$folder_temp/$sample-HeatmapPercent-Genomic-Report.png"); + + # Bar plot for the sequence context on the genomic strand (6 mutation types) + $ws->insert_image(27, $col+3, "$folder_temp/$sample-MutationSpectraPercent-Genomic-Report.png"); +} + + +############################################################################################################ +# Write NMF input for count and percentages in the Excel report +sub writeInputNMF +{ + my ($ws_inputNMF_count, $ws_inputNMF_percent, $outCount, $outPercent) = @_; + + + open(OUTINPUTNMFC, ">", $outCount) or die "$!: $outCount\n"; # with the count + open(OUTINPUTNMFP, ">", $outPercent) or die "$!: $outPercent\n"; # With the frequency un-normalized + + foreach my $k_sample (@{$h_inputNMF{'Sample'}}) + { + print OUTINPUTNMFC "\t$k_sample"; + print OUTINPUTNMFP "\t$k_sample"; + } + print OUTINPUTNMFC "\n"; print OUTINPUTNMFP "\n"; + + my $row_inputNMF = 1; + foreach my $k_context (sort keys %{$h_inputNMF{'Count'}}) + { + $k_context =~ /(\w)_(\w)/; my ($base5, $base3) = ($1, $2); + foreach my $k_mutation (sort keys %{$h_inputNMF{'Count'}{$k_context}}) + { + my ($col_inputNMF_Count, $col_inputNMF_Percent) = (1, 1); + my $contextNMF = $base5."[$k_mutation]".$base3; + + # Write the input in the Excel report, only when all the samples are in the same workbook + if($oneReportPerSample == 2) + { + $ws_inputNMF_count->write($row_inputNMF, 0, $contextNMF); + $ws_inputNMF_percent->write($row_inputNMF, 0, $contextNMF); + } + + print OUTINPUTNMFC $contextNMF,"\t"; print OUTINPUTNMFP $contextNMF,"\t"; + + foreach (@{$h_inputNMF{'Count'}{$k_context}{$k_mutation}}) { print OUTINPUTNMFC "$_\t"; } print OUTINPUTNMFC "\n"; + foreach (@{$h_inputNMF{'Percent'}{$k_context}{$k_mutation}}) { print OUTINPUTNMFP "$_\t"; } print OUTINPUTNMFP "\n"; + + foreach (@{$h_inputNMF{'Count'}{$k_context}{$k_mutation}}) + { + if($oneReportPerSample == 2) + { + $ws_inputNMF_count->write($row_inputNMF, $col_inputNMF_Count, $_); + } + $col_inputNMF_Count++; + } + foreach (@{$h_inputNMF{'Percent'}{$k_context}{$k_mutation}}) + { + if($oneReportPerSample == 2) + { + $ws_inputNMF_percent->write($row_inputNMF, $col_inputNMF_Percent, $_); + } + $col_inputNMF_Percent++; + } + $row_inputNMF++; + } + } + close OUTINPUTNMFP; close OUTINPUTNMFC; +} + + +###################################################################################################################################################### +# Define format and background colors for the Excel report # +###################################################################################################################################################### +# Font: Arial size 10 +sub Format_A10 +{ + my ($wb, $format) = @_; + $$format = $wb->add_format(font=>'Arial', size=>10); $$format->set_align('center'); +} +# Font: Arial size 11 bold and center +sub Format_A11Bold +{ + my ($wb, $format) = @_; + $$format = $wb->add_format(font=>'Arial', size=>11, bold=>1); $$format->set_align('center'); +} +# Font: Arial size 10 italic red and center +sub Format_A10ItalicRed +{ + my ($wb, $format) = @_; + $$format = $wb->add_format(font=>'Arial', size=>10, italic=>1, color => 'red'); $$format->set_align('center'); +} +# Format: Arialt size 11 bold and left +sub Format_A11BoldLeft +{ + my ($wb, $format) = @_; + $$format = $wb->add_format(valign =>'left', font=>'Arial', size=>11, bold=>1); +} +# Font: Arialt size 10 bold and left +sub Format_A10BoldLeft +{ + my ($wb, $format) = @_; + $$format = $wb->add_format(valign =>'left', font=>'Arial', size=>10, bold=>1); +} +# Define the format of the border of the section (for delimiting the different section of the report) +sub Format_section +{ + my ($wb, $format_topLeft, $format_topRight, $format_bottomLeft, $format_bottomRight, $format_top, $format_right, $format_bottom, $format_left) = @_; + + $$format_topLeft = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); + $$format_topLeft->set_top(2); $$format_topLeft->set_top_color('blue'); + $$format_topLeft->set_left(2); $$format_topLeft->set_left_color('blue'); + + $$format_topRight = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); + $$format_topRight->set_top(2); $$format_topRight->set_top_color('blue'); + $$format_topRight->set_right(2); $$format_topRight->set_right_color('blue'); + + $$format_bottomLeft = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); + $$format_bottomLeft->set_bottom(2); $$format_bottomLeft->set_bottom_color('blue'); + $$format_bottomLeft->set_left(2); $$format_bottomLeft->set_left_color('blue'); + + $$format_bottomRight = $wb->add_format(valign => 'left', bold => 1, font => 'Arial', size => 12); + $$format_bottomRight->set_bottom(2); $$format_bottomRight->set_bottom_color('blue'); + $$format_bottomRight->set_right(2); $$format_bottomRight->set_right_color('blue'); + + $$format_top = $wb->add_format(); $$format_top->set_top(2); $$format_top->set_top_color('blue'); + $$format_right = $wb->add_format(); $$format_right->set_right(2); $$format_right->set_right_color('blue'); + $$format_bottom = $wb->add_format(); $$format_bottom->set_bottom(2); $$format_bottom->set_bottom_color('blue'); + $$format_left = $wb->add_format(); $$format_left->set_left(2); $$format_left->set_left_color('blue'); +} +# Define the header +sub Format_Header +{ + my ($wb, $format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG, $format_TG2, $format_LeftHeader, $format_RightHeader, $format_LeftHeader2) = @_; + + my ($blue, $black, $red, $gray, $green, $pink); + Color($wb, \$blue, \$black, \$red, \$gray, \$green, \$pink); + + my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); + BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); + + + $$format_CA = $wb->add_format(bg_color => $blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CA->set_align('center'); $$format_CA->set_center_across(); + $$format_CG = $wb->add_format(bg_color => $black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CG->set_align('center'); $$format_CG->set_center_across(); + $$format_CT = $wb->add_format(bg_color => $red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CT->set_align('center'); $$format_CT->set_center_across(); + $$format_TA = $wb->add_format(bg_color => $gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TA->set_align('center'); $$format_TA->set_center_across(); + $$format_TC = $wb->add_format(bg_color => $green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TC->set_align('center'); $$format_TC->set_center_across(); + $$format_TG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG->set_align('center'); $$format_TG->set_center_across(); + $$format_TG->set_right(2); $$format_TG->set_right_color('blue'); + + $$format_TG2 = $wb->add_format(bg_color => $pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG2->set_align('center'); $$format_TG2->set_center_across(); + + $$format_LeftHeader = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_LeftHeader->set_align('center'); $$format_LeftHeader->set_left(2); $$format_LeftHeader->set_left_color('blue'); + $$format_LeftHeader2 = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_LeftHeader2->set_left(2); $$format_LeftHeader2->set_left_color('blue'); + $$format_RightHeader = $wb->add_format(bold=>1, font=>'Arial', size=>11); $$format_RightHeader->set_align('center'); $$format_RightHeader->set_right(2); $$format_RightHeader->set_right_color('blue'); +} +# Define the header for the part "Strand bias by segment" +sub Format_HeaderSBSDistrBySegAndFunc +{ + my ($wb, $format_LeftCA, $format_LeftCG, $format_LeftCT, $format_LeftTA, $format_LeftTC, $format_LeftTG, $format_RightCA, $format_RightCG, $format_RightCT, $format_RightTA, $format_RightTC, $format_RightTG) = @_; + + my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); + BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); + + $$format_LeftCA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCA->set_align('center'); $$format_LeftCA->set_left(2); $$format_LeftCA->set_left_color('blue'); + $$format_LeftCG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCG->set_align('center'); $$format_LeftCG->set_left(2); $$format_LeftCG->set_left_color('blue'); + $$format_LeftCT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftCT->set_align('center'); $$format_LeftCT->set_left(2); $$format_LeftCT->set_left_color('blue'); + $$format_LeftTA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTA->set_align('center'); $$format_LeftTA->set_left(2); $$format_LeftTA->set_left_color('blue'); + $$format_LeftTC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTC->set_align('center'); $$format_LeftTC->set_left(2); $$format_LeftTC->set_left_color('blue'); + $$format_LeftTG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_LeftTG->set_align('center'); $$format_LeftTG->set_left(2); $$format_LeftTG->set_left_color('blue'); + + + $$format_RightCA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCA->set_align('center'); $$format_RightCA->set_right(2); $$format_RightCA->set_right_color('blue'); + $$format_RightCG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCG->set_align('center'); $$format_RightCG->set_right(2); $$format_RightCG->set_right_color('blue'); + $$format_RightCT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightCT->set_align('center'); $$format_RightCT->set_right(2); $$format_RightCT->set_right_color('blue'); + $$format_RightTA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTA->set_align('center'); $$format_RightTA->set_right(2); $$format_RightTA->set_right_color('blue'); + $$format_RightTC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTC->set_align('center'); $$format_RightTC->set_right(2); $$format_RightTC->set_right_color('blue'); + $$format_RightTG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_RightTG->set_align('center'); $$format_RightTG->set_right(2); $$format_RightTG->set_right_color('blue'); +} +# Define the header for the part "Trinucleotide sequence context on the coding strand" +sub Format_Header12MutType +{ + my ($wb, $format_CA, $format_CG, $format_CT, $format_TA, $format_TC, $format_TG) = @_; + + my ($bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink); + BackgroundColor($wb, \$bgColor_blue, \$bgColor_black, \$bgColor_red, \$bgColor_gray, \$bgColor_green, \$bgColor_pink); + + $$format_CA = $wb->add_format(bg_color=>$bgColor_blue, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CA->set_align('center'); + $$format_CG = $wb->add_format(bg_color=>$bgColor_black, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CG->set_align('center'); + $$format_CT = $wb->add_format(bg_color=>$bgColor_red, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_CT->set_align('center'); + $$format_TA = $wb->add_format(bg_color=>$bgColor_gray, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TA->set_align('center'); + $$format_TC = $wb->add_format(bg_color=>$bgColor_green, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TC->set_align('center'); + $$format_TG = $wb->add_format(bg_color=>$bgColor_pink, font=>'Arial', bold=>1, size=>11, color=>'white'); $$format_TG->set_align('center'); +} +# Define the format for the text that needs a section border +sub Format_TextSection +{ + my ($wb, $formatT_left, $formatT_right, $formatT_bottomRight, $formatT_bottomLeft, $formatT_bottom, $formatT_bottomHeader, $formatT_bottomRightHeader, $formatT_bottomHeader2, $formatT_rightHeader) = @_; + + $$formatT_left = $wb->add_format(valign=>'center', font=>'Arial', size=>10); + $$formatT_left->set_left(2); $$formatT_left->set_left_color('blue'); + + $$formatT_right = $wb->add_format(valign=>'center', font=>'Arial', size=>10); + $$formatT_right->set_right(2); $$formatT_right->set_right_color('blue'); + + $$formatT_bottomRight = $wb->add_format(valign=>'center', font=>'Arial', size=>10); + $$formatT_bottomRight->set_bottom(2); $$formatT_bottomRight->set_bottom_color('blue'); + $$formatT_bottomRight->set_right(2); $$formatT_bottomRight->set_right_color('blue'); + + $$formatT_bottomLeft = $wb->add_format(valign=>'center', font=>'Arial', size=>10); + $$formatT_bottomLeft->set_bottom(2); $$formatT_bottomLeft->set_bottom_color('blue'); + $$formatT_bottomLeft->set_left(2); $$formatT_bottomLeft->set_left_color('blue'); + + $$formatT_bottom = $wb->add_format(valign=>'center', font=>'Arial', size=>10); + $$formatT_bottom->set_bottom(2); $$formatT_bottom->set_bottom_color('blue'); + + my $bgColor_totallighGray = $wb->set_custom_color(54, 230, 230, 230); + $$formatT_bottomHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomHeader->set_align('center'); + $$formatT_bottomHeader->set_bottom(2); $$formatT_bottomHeader->set_bottom_color('blue'); + + $$formatT_bottomRightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomRightHeader->set_align('center'); + $$formatT_bottomRightHeader->set_bottom(2); $$formatT_bottomRightHeader->set_bottom_color('blue'); + $$formatT_bottomRightHeader->set_right(2); $$formatT_bottomRightHeader->set_right_color('blue'); + + $$formatT_bottomHeader2 = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_bottomHeader2->set_align('center'); + + $$formatT_rightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>11); $$formatT_rightHeader->set_align('center'); + $$formatT_rightHeader->set_right(2); $$formatT_rightHeader->set_right_color('blue'); +} +# Define the format for the graphs titles +sub Format_GraphTitle +{ + my ($wb, $formatT_graphTitle) = @_; + + $$formatT_graphTitle = $wb->add_format(font=>'Arial', size=>12, bold=>1); +} +# Define the format of the border of the tables +sub Format_Table +{ + my ($wb, $table_topleft, $table_topRight, $table_bottomleft, $table_bottomRight, $table_top, $table_right, $table_bottom, $table_bottomItalicRed, $table_left, $table_bottomrightHeader, $table_left2, $table_middleHeader, $table_middleHeader2) = @_; + + $$table_topleft = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_topleft->set_top(1); $$table_topleft->set_left(1); + $$table_topRight = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_topRight->set_top(1); $$table_topRight->set_right(1); + $$table_bottomleft = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_bottomleft->set_bottom(1); $$table_bottomleft->set_left(1); + $$table_bottomRight = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_bottomRight->set_bottom(1); $$table_bottomRight->set_right(1); + + $$table_top = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_top->set_top(1); + $$table_right = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_right->set_right(1); + $$table_bottom = $wb->add_format(valign=>'center', font=>'Arial', size=>10); $$table_bottom->set_bottom(1); + $$table_bottomItalicRed = $wb->add_format(valign=>'center', font=>'Arial', size=>10, italic=>1, color => 'red'); $$table_bottomItalicRed->set_bottom(1); + $$table_left = $wb->add_format(valign=>'center', bold=>1, font=>'Arial', size=>10); $$table_left->set_left(1); + + my $bgColor_totallighGray = $wb->set_custom_color(54, 230, 230, 230); + $$table_bottomrightHeader = $wb->add_format(bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); $$table_bottomrightHeader->set_bottom(1); $$table_bottomrightHeader->set_right(1); + + $$table_left2 = $wb->add_format(valign=>'left', font=>'Arial', size=>10); $$table_left2->set_left(1); + + $$table_middleHeader = $wb->add_format(valign=>'center', bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); + $$table_middleHeader2 = $wb->add_format(valign=>'center', bg_color=>$bgColor_totallighGray, font=>'Arial', bold=>1, size=>10); $$table_middleHeader2->set_bottom(1); +} +# Define the color +sub Color +{ + my ($wb, $blue, $black, $red, $gray, $green, $pink) = @_; + + $$blue = $wb->set_custom_color(40, 0, 0, 204);# C:G>A:T in blue + $$black = $wb->set_custom_color(41, 0, 0, 0);# C:G>G:C in black + $$red = $wb->set_custom_color(42, 255, 0, 0);# C:G>T:A in red + $$gray = $wb->set_custom_color(43, 205, 205, 205); # T:A>A:T in light gray + $$green = $wb->set_custom_color(44, 0, 204, 51);# T:A>C:G in green + $$pink = $wb->set_custom_color(45, 255, 192, 203);# T:A>G:C in pink +} +sub BackgroundColor +{ + my ($wb, $bgColor_blue, $bgColor_black, $bgColor_red, $bgColor_gray, $bgColor_green, $bgColor_pink) = @_; + + $$bgColor_blue = $wb->set_custom_color(48, 0, 0, 204); + $$bgColor_black = $wb->set_custom_color(49, 0, 0, 0); + $$bgColor_red = $wb->set_custom_color(50, 255, 0, 0); + $$bgColor_gray = $wb->set_custom_color(51, 205, 205, 205); + $$bgColor_green = $wb->set_custom_color(52, 0, 204, 51); + $$bgColor_pink = $wb->set_custom_color(53, 255, 192, 203); +} + + + + +=head1 NAME + +mutSpec-Stat + +=head1 SYNOPSIS + + mutSpecstat.pl [arguments] <query-file> + + <query-file> a folder with one or multiple VCFs + + Arguments: + -h, --help print help message + -m, --man print complete documentation + -v, --verbose use verbose output + --refGenome the reference genome to use (human, mouse or rat genomes) + -o, --outfile <string> output directory for the result. If none is specify the result will be write in the same directory as the input file + --temp <string> the path for saving the temporary files + --pathSeqRefGenome the path to the fasta reference sequences + --poolData generate the pool of all the samples (optional) + --reportSample generate a report for each sample (optional) + + +Function: automatically run a pipeline and calculate various statistics on mutations + + Example: mutSpecstat.pl --refGenome hg19 --outfile output_directory --temp path_to_temporary_directory --pathRscript path_to_R_scripts --pathSeqRefGenome path_fasta_ref_seq --poolData --reportSample inputFolder + + Version: 02-2017 (February 2016) + + +=head1 OPTIONS + +=over 8 + +=item B<--help> + +print a brief usage message and detailed explanation of options. + +=item B<--man> + +print the complete manual of the program. + +=item B<--verbose> + +use verbose output. + +=item B<--refGenome> + +the reference genome to use, could be human, mouse or rat genomes. + +=item B<--outfile> + +the directory of output file names. If it is nor specify the same directory as the input file is used. + +=item B<--temp> + +the path for saving temporary files generated by the script. +If any is specify a temporary folder is created in the same directory where the script is running. +Deleted when the script is finish + +=item B<--pathSeqRefGenome> + +The path to the fasta reference sequences + +=item B<--poolData only for the report> + +calculate the statistics on the pool of all the data pass in input + +=item B<--reportSample only for the report> + +generate a report for each samples + +=head1 DESCRIPTION + +mutSpecstat is a perl script for calculated various statistics on mutations +An Excel report containing the mutation type distribution per functional region, the strand bias and the sequence context on genomic and coding sequence is created. +The different statistics are illustrated using ggplot2. + +=cut