Mercurial > repos > ziru-zhou > spp_phantompeak
diff run_spp.R @ 0:89c62e3b0f85 draft
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| author | ziru-zhou |
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| date | Tue, 18 Dec 2012 10:44:50 -0500 |
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--- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/run_spp.R Tue Dec 18 10:44:50 2012 -0500 @@ -0,0 +1,885 @@ +# run_spp.R +# ============= +# Author: Anshul Kundaje, Computer Science Dept., Stanford University +# Email: akundaje@stanford.edu +# Last updated: Oct 8, 2010 +# ============= +# MANDATORY ARGUMENTS +# -c=<ChIP_tagAlign/BAMFile>, full path and name of tagAlign/BAM file (can be gzipped) (FILE EXTENSION MUST BE tagAlign.gz, tagAlign, bam or bam.gz) +# MANDATORY ARGUMENT FOR PEAK CALLING +# -i=<Input_tagAlign/BAMFile>, full path and name of tagAlign/BAM file (can be gzipped) (FILE EXTENSION MUST BE tagAlign.gz, tagAlign, bam or bam.gz) +# OPTIONAL ARGUMENTS +# -s=<min>:<step>:<max> , strand shifts at which cross-correlation is evaluated, default=-100:5:600 +# -speak=<strPeak>, user-defined cross-correlation peak strandshift +# -x=<min>:<max>, strand shifts to exclude (This is mainly to avoid phantom peaks) default=10:(readlen+10) +# -p=<nodes> , number of parallel processing nodes, default=NULL +# -fdr=<falseDisoveryRate> , false discovery rate threshold for peak calling +# -npeak=<numPeaks>, threshold on number of peaks to call +# -tmpdir=<tempdir> , Temporary directory (if not specified R function tempdir() is used) +# -filtchr=<chrnamePattern> , Pattern to use to remove tags that map to specific chromosomes e.g. _ will remove all tags that map to chromosomes with _ in their name +# OUTPUT PARAMETERS +# -odir=<outputDirectory> name of output directory (If not set same as ChIP file directory is used) +# -savn=<narrowpeakfilename> OR -savn NarrowPeak file name +# -savr=<regionpeakfilename> OR -savr RegionPeak file name +# -savd=<rdatafile> OR -savd , save Rdata file +# -savp=<plotdatafile> OR -savp , save cross-correlation plot +# -out=<resultfile>, append peakshift result to a file +# format:Filename<tab>numReads<tab>estFragLen<tab>corr_estFragLen<tab>PhantomPeak<tab>corr_phantomPeak<tab>argmin_corr<tab>min_corr<tab>phantomPeakCoef<tab>relPhantomPeakCoef<tab>QualityTag +# -rf , if plot or rdata or narrowPeak file exists replace it. If not used then the run is aborted if the plot or Rdata or narrowPeak file exists +# -clean, if present will remove the original chip and control files after reading them in. CAUTION: Use only if the script calling run_spp.R is creating temporary files + +args <- commandArgs(trailingOnly=TRUE); # Read Arguments from command line +nargs = length(args); # number of arguments + +# ########################################################################### +# AUXILIARY FUNCTIONS +# ########################################################################### + +print.usage <- function() { +# =================================== +# Function will print function usage +# =================================== + cat('Usage: Rscript run_spp.R <options>\n',file=stderr()) + cat('MANDATORY ARGUMENTS\n',file=stderr()) + cat('-c=<ChIP_alignFile>, full path and name (or URL) of tagAlign/BAM file (can be gzipped)(FILE EXTENSION MUST BE tagAlign.gz, tagAlign, bam or bam.gz) \n',file=stderr()) + cat('MANDATORY ARGUMENTS FOR PEAK CALLING\n',file=stderr()) + cat('-i=<Input_alignFile>, full path and name (or URL) of tagAlign/BAM file (can be gzipped) (FILE EXTENSION MUST BE tagAlign.gz, tagAlign, bam or bam.gz) \n',file=stderr()) + cat('OPTIONAL ARGUMENTS\n',file=stderr()) + cat('-s=<min>:<step>:<max> , strand shifts at which cross-correlation is evaluated, default=-100:5:600\n',file=stderr()) + cat('-speak=<strPeak>, user-defined cross-correlation peak strandshift\n',file=stderr()) + cat('-x=<min>:<max>, strand shifts to exclude (This is mainly to avoid region around phantom peak) default=10:(readlen+10)\n',file=stderr()) + cat('-p=<nodes> , number of parallel processing nodes, default=0\n',file=stderr()) + cat('-fdr=<falseDisoveryRate> , false discovery rate threshold for peak calling\n',file=stderr()) + cat('-npeak=<numPeaks>, threshold on number of peaks to call\n',file=stderr()) + cat('-tmpdir=<tempdir> , Temporary directory (if not specified R function tempdir() is used)\n',file=stderr()) + cat('-filtchr=<chrnamePattern> , Pattern to use to remove tags that map to specific chromosomes e.g. _ will remove all tags that map to chromosomes with _ in their name\n',file=stderr()) + cat('OUTPUT ARGUMENTS\n',file=stderr()) + cat('-odir=<outputDirectory> name of output directory (If not set same as ChIP file directory is used)\n',file=stderr()) + cat('-savn=<narrowpeakfilename> OR -savn NarrowPeak file name (fixed width peaks)\n',file=stderr()) + cat('-savr=<regionpeakfilename> OR -savr RegionPeak file name (variable width peaks with regions of enrichment)\n',file=stderr()) + cat('-savd=<rdatafile> OR -savd, save Rdata file\n',file=stderr()) + cat('-savp=<plotdatafile> OR -savp, save cross-correlation plot\n',file=stderr()) + cat('-out=<resultfile>, append peakshift/phantomPeak results to a file\n',file=stderr()) + cat(' format:Filename<tab>numReads<tab>estFragLen<tab>corr_estFragLen<tab>PhantomPeak<tab>corr_phantomPeak<tab>argmin_corr<tab>min_corr<tab>phantomPeakCoef<tab>relPhantomPeakCoef<tab>QualityTag)\n',file=stderr()) + cat('-rf, if plot or rdata or narrowPeak file exists replace it. If not used then the run is aborted if the plot or Rdata or narrowPeak file exists\n',file=stderr()) + cat('-clean, if present will remove the original chip and control files after reading them in. CAUTION: Use only if the script calling run_spp.R is creating temporary files\n',file=stderr()) +} # end: print.usage() + +get.file.parts <- function(file.fullpath) { +# =================================== +# Function will take a file name with path and split the file name into +# path, fullname, name and ext +# =================================== + if (! is.character(file.fullpath)) { + stop('File name must be a string') + } + + file.parts <- strsplit(as.character(file.fullpath), .Platform$file.sep, fixed=TRUE)[[1]] # split on file separator + + if (length(file.parts) == 0) { # if empty file name + return(list(path='', + fullname='', + name='', + ext='') + ) + } else { + if (length(file.parts) == 1) { # if no path then just the file name itself + file.path <- '.' + file.fullname <- file.parts + } else { + file.path <- paste(file.parts[1:(length(file.parts)-1)], collapse=.Platform$file.sep) # 1:last-1 token is path + file.fullname <- file.parts[length(file.parts)] # last token is filename + } + file.fullname.parts <- strsplit(file.fullname,'.',fixed=TRUE)[[1]] # split on . + if (length(file.fullname.parts) == 1) { # if no extension + file.ext <- '' + file.name <- file.fullname.parts + } else { + file.ext <- paste('.', file.fullname.parts[length(file.fullname.parts)], sep="") # add the . to the last token + file.name <- paste(file.fullname.parts[1:(length(file.fullname.parts)-1)], collapse=".") + } + return(list(path=file.path, + fullname=file.fullname, + name=file.name, + ext=file.ext)) + } +} # end: get.file.parts() + +parse.arguments <- function(args) { +# =================================== +# Function will parse arguments +# =================================== + # Set arguments to default values + chip.file <- NA # main ChIP tagAlign/BAM file name + isurl.chip.file <- FALSE # flag indicating whether ChIP file is a URL + control.file <- NA # control tagAlign/BAM file name + isurl.control.file <- FALSE # flag indicating whether control file is a URL + sep.min <- -100 # min strand shift + sep.max <- 600 # max strand shift + sep.bin <- 5 # increment for strand shift + sep.peak <- NA # user-defined peak shift + exclude.min <- 10 # lowerbound of strand shift exclusion region + exclude.max <- NaN # upperbound of strand shift exclusion region + n.nodes <- NA # number of parallel processing nodes + fdr <- 0.01 # false discovery rate threshold for peak calling + npeak <- NA # threshold on number of peaks to call + temp.dir <- tempdir() # temporary directory + chrname.rm.pattern <- NA # chromosome name pattern used to remove tags + output.odir <- NA # Output directory name + output.npeak.file <- NA # Output narrowPeak file name + output.rpeak.file <- NA # Output regionPeak file name + output.rdata.file <- NA # Rdata file + output.plot.file <- NA # cross correlation plot file + output.result.file <- NA # result file + replace.flag <- FALSE # replace file flag + clean.files.flag <- FALSE # file deletion flag + + # Parse arguments + for (each.arg in args) { + + if (grepl('^-c=',each.arg)) { #-c=<chip.file> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + chip.file <- arg.split[2] # second part is chip.file + } else { + stop('No tagAlign/BAM file name provided for parameter -c=') + } + + } else if (grepl('^-i=',each.arg)) { #-i=<control.file> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + control.file <- arg.split[2] # second part is control.file + } else { + stop('No tagAlign/BAM file name provided for parameter -i=') + } + + } else if (grepl('^-s=',each.arg)) { #-s=<sep.min>:<sep.bin>:<sep.max> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + sep.vals <- arg.split[2] # second part is sepmin:sepbin:sepmax + sep.vals.split <- strsplit(sep.vals,':',fixed=TRUE)[[1]] # split on : + if (length(sep.vals.split) != 3) { # must have 3 parts + stop('Strand shift limits must be specified as -s=sepmin:sepbin:sepmax') + } else { + if (any(is.na(as.numeric(sep.vals.split)))) { # check that sep vals are numeric + stop('Strand shift limits must be numeric values') + } + sep.min <- round(as.numeric(sep.vals.split[1])) + sep.bin <- round(as.numeric(sep.vals.split[2])) + sep.max <- round(as.numeric(sep.vals.split[3])) + if ((sep.min > sep.max) || (sep.bin > (sep.max - sep.min)) || (sep.bin < 0)) { + stop('Illegal separation values -s=sepmin:sepbin:sepmax') + } + } + } else { + stop('Strand shift limits must be specified as -s=sepmin:sepbin:sepmax') + } + + } else if (grepl('^-speak=',each.arg)) { #-speak=<sep.peak> , user-defined cross-correlation peak strandshift + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + sep.peak <- arg.split[2] # second part is <sep.peak> + if (is.na(as.numeric(sep.peak))) { # check that sep.peak is numeric + stop('-speak=<sep.peak>: User defined peak shift must be numeric') + } + sep.peak <- as.numeric(sep.peak) + } else { + stop('User defined peak shift must be provided as -speak=<sep.peak>') + } + + } else if (grepl('^-x=',each.arg)) { #-x=<exclude.min>:<exclude.max> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + exclude.vals <- arg.split[2] # second part is excludemin:excludemax + exclude.vals.split <- strsplit(exclude.vals,':',fixed=TRUE)[[1]] # split on : + if (length(exclude.vals.split) != 2) { # must have 2 parts + stop('Exclusion limits must be specified as -x=excludemin:excludemax') + } else { + if (any(is.na(as.numeric(exclude.vals.split)))) { # check that exclude vals are numeric + stop('Exclusion limits must be numeric values') + } + exclude.min <- round(as.numeric(exclude.vals.split[1])) + exclude.max <- round(as.numeric(exclude.vals.split[2])) + if (exclude.min > exclude.max) { + stop('Illegal exclusion limits -x=excludemin:excludemax') + } + } + } else { + stop('Exclusion limits must be specified as -x=excludemin:excludemax') + } + + } else if (grepl('^-p=',each.arg)) { #-p=<n.nodes> , number of parallel processing nodes, default=NULL + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + n.nodes <- arg.split[2] # second part is numnodes + if (is.na(as.numeric(n.nodes))) { # check that n.nodes is numeric + stop('-p=<numnodes>: numnodes must be numeric') + } + n.nodes <- round(as.numeric(n.nodes)) + } else { + stop('Number of parallel nodes must be provided as -p=<numnodes>') + } + + } else if (grepl('^-fdr=',each.arg)) { #-fdr=<fdr> , false discovery rate, default=0.01 + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + fdr <- arg.split[2] # second part is fdr + if (is.na(as.numeric(fdr))) { # check that fdr is numeric + stop('-fdr=<falseDiscoveryRate>: false discovery rate must be numeric') + } + fdr <- as.numeric(fdr) + } else { + stop('False discovery rate must be provided as -fdr=<fdr>') + } + + } else if (grepl('^-npeak=',each.arg)) { #-npeak=<numPeaks> , number of peaks threshold, default=NA + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + npeak <- arg.split[2] # second part is npeak + if (is.na(as.numeric(npeak))) { # check that npeak is numeric + stop('-npeak=<numPeaks>: threshold on number of peaks must be numeric') + } + npeak <- round(as.numeric(npeak)) + } else { + stop('Threshold on number of peaks must be provided as -npeak=<numPeaks>') + } + + } else if (grepl('^-tmpdir=',each.arg)) { #-tmpdir=<temp.dir> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + temp.dir <- arg.split[2] # second part is temp.dir + } else { + stop('No temporary directory provided for parameter -tmpdir=') + } + + } else if (grepl('^-filtchr=',each.arg)) { #-filtchr=<chrname.rm.pattern> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + chrname.rm.pattern <- arg.split[2] # second part is chrname.rm.pattern + } else { + stop('No pattern provided for parameter -filtchr=') + } + + } else if (grepl('^-odir=',each.arg)) { #-odir=<output.odir> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + output.odir <- arg.split[2] # second part is output.odir + } else { + stop('No output directory provided for parameter -odir=') + } + + } else if (grepl('^-savn',each.arg)) { # -savn=<output.npeak.file> OR -savn , save narrowpeak + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2])) { + output.npeak.file <- arg.split[2] #-savn= + } else if (each.arg=='-savn') { + output.npeak.file <- NULL # NULL indicates get the name from the main file name + } else { + stop('Argument for saving narrowPeak file must be -savn or -savn=<filename>') + } + + } else if (grepl('^-savr',each.arg)) { # -savr=<output.rpeak.file> OR -savr , save regionpeak + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2])) { + output.rpeak.file <- arg.split[2] #-savr= + } else if (each.arg=='-savr') { + output.rpeak.file <- NULL # NULL indicates get the name from the main file name + } else { + stop('Argument for saving regionPeak file must be -savr or -savr=<filename>') + } + + } else if (grepl('^-savd',each.arg)) { # -savd=<output.rdata.file> OR -savd , save Rdata file + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2])) { + output.rdata.file <- arg.split[2] #-savd= + } else if (each.arg=='-savd') { + output.rdata.file <- NULL # NULL indicates get the name from the main file name + } else { + stop('Argument for saving Rdata file must be -savd or -savd=<filename>') + } + + } else if (grepl('^-savp',each.arg)) { # -savp=<output.plot.file> OR -savp , save cross-correlation plot + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2])) { + output.plot.file <- arg.split[2] #-savp= + } else if (each.arg=='-savp') { + output.plot.file <- NULL # NULL indicates get the name from the main file name + } else { + stop('Argument for saving Rdata file must be -savp or -savp=<filename>') + } + + } else if (grepl('^-out=',each.arg)) { #-out=<output.result.file> + + arg.split <- strsplit(each.arg,'=',fixed=TRUE)[[1]] # split on = + if (! is.na(arg.split[2]) ) { + output.result.file <- arg.split[2] # second part is output.result.file + } else { + stop('No result file provided for parameter -out=') + } + + } else if (each.arg == '-rf') { + + replace.flag <- TRUE + + } else if (each.arg == '-clean') { + + clean.files.flag <- TRUE + + } else { + + stop('Illegal argument ',each.arg) + } + } + # End: for loop + + # Check mandatory arguments + if (is.na(chip.file)) { + stop('-c=<tagAlign/BAMFileName> is a mandatory argument') + } + + if (is.na(control.file) && ! is.na(output.npeak.file)) { + stop('-i=<tagAlign/BAMFileName> is required for peak calling') + } + + # Check if ChIP and control files are URLs + if (grepl('^http://',chip.file)) { + isurl.chip.file <- TRUE + } + if (grepl('^http://',control.file)) { + isurl.control.file <- TRUE + } + + # If ChIP file is a URL output.odir MUST be specified + if (isurl.chip.file && is.na(output.odir)) { + stop('If ChIP file is a URL, then output directory MUST be specified') + } + + # Check that ChIP and control files exist + if (isurl.chip.file) { + if (system(paste('wget -q --spider',chip.file)) != 0) { + stop('ChIP file URL not valid: ',chip.file) + } + } else if (!file.exists(chip.file)) { + stop('ChIP File:',chip.file,' does not exist') + } + + if (!is.na(control.file)) { + if (isurl.control.file) { + if (system(paste('wget -q --spider',control.file)) != 0) { + stop('Control file URL not valid: ',control.file) + } + } else if (!file.exists(control.file)) { + stop('Control File:',control.file,' does not exist') + } + } + + # Correct other arguments + if (is.na(output.odir)) { # Reconstruct output.odir if not provided + output.odir <- get.file.parts(chip.file)$path + } + + if (is.null(output.npeak.file)) { # Reconstruct output.npeak.file if NULL + output.npeak.file <- file.path(output.odir, paste(get.file.parts(chip.file)$name, '_VS_', get.file.parts(control.file)$name,'.narrowPeak', sep="")) + } + + if (is.null(output.rpeak.file)) { # Reconstruct output.rpeak.file if NULL + output.rpeak.file <- file.path(output.odir, paste(get.file.parts(chip.file)$name, '_VS_', get.file.parts(control.file)$name,'.regionPeak', sep="")) + } + + if (is.null(output.rdata.file)) { # Reconstruct output.rdata.file if NULL + output.rdata.file <- file.path(output.odir, paste(get.file.parts(chip.file)$name, '.Rdata', sep="")) + } + + if (is.null(output.plot.file)) { # Reconstruct output.plot.file if NULL + output.plot.file <- file.path(output.odir, paste(get.file.parts(chip.file)$name, '.pdf', sep="")) + } + + return(list(chip.file=chip.file, + isurl.chip.file=isurl.chip.file, + control.file=control.file, + isurl.control.file=isurl.control.file, + sep.range=c(sep.min,sep.bin,sep.max), + sep.peak=sep.peak, + ex.range=c(exclude.min,exclude.max), + n.nodes=n.nodes, + fdr=fdr, + npeak=npeak, + temp.dir=temp.dir, + chrname.rm.pattern=chrname.rm.pattern, + output.odir=output.odir, + output.npeak.file=output.npeak.file, + output.rpeak.file=output.rpeak.file, + output.rdata.file=output.rdata.file, + output.plot.file=output.plot.file, + output.result.file=output.result.file, + replace.flag=replace.flag, + clean.files.flag=clean.files.flag)) +} # end: parse.arguments() + +read.align <- function(align.filename) { +# =================================== +# Function will read a tagAlign or BAM file +# =================================== + if (grepl('(\\.bam)?.*(\\.tagAlign)',align.filename)) { # if tagalign file + chip.data <- read.tagalign.tags(align.filename) + # get readlength info + tmpDataRows <- read.table(align.filename,nrows=500) + chip.data$read.length <- round(median(tmpDataRows$V3 - tmpDataRows$V2)) + } else if (grepl('(\\.tagAlign)?.*(\\.bam)',align.filename)) { # if bam file + # create BAM file name + bam2align.filename <- sub('\\.bam','.tagAlign',align.filename) + # generate command to convert bam to tagalign + command <- vector(length=2) + command[1] <- sprintf("samtools view -F 0x0204 -o - %s",align.filename) + command[2] <- paste("awk 'BEGIN{FS=" , '"\t"' , ";OFS=", '"\t"} {if (and($2,16) > 0) {print $3,($4-1),($4-1+length($10)),"N","1000","-"} else {print $3,($4-1),($4-1+length($10)),"N","1000","+"}}', "' 1> ", bam2align.filename, sep="") + # command[2] <- paste("awk 'BEGIN{OFS=", '"\t"} {if (and($2,16) > 0) {print $3,($4-1),($4-1+length($10)),"N","1000","-"} else {print $3,($4-1),($4-1+length($10)),"N","1000","+"}}', "' 1> ", bam2align.filename, sep="") + command <- paste(command,collapse=" | ") + # Run command + status <- system(command,intern=FALSE,ignore.stderr=FALSE) + if ((status != 0) || !file.exists(bam2align.filename)) { + cat(sprintf("Error converting BAM to tagalign file: %s\n",align.filename),file=stderr()) + q(save="no",status=1) + } + # read converted BAM file + chip.data <- read.tagalign.tags(bam2align.filename) + # get readlength info + tmpDataRows <- read.table(bam2align.filename,nrows=500) + chip.data$read.length <- round(median(tmpDataRows$V3 - tmpDataRows$V2)) + # delete temporary tagalign file + file.remove(bam2align.filename) + } else { + cat(sprintf("Error:Unknown file format for file:%s\n",align.fname),file=stderr()) + q(save="no",status=1) + } + return(chip.data) +} # end: read.align() + +print.run.params <- function(params){ +# =================================== +# Output run parameters +# =================================== + cat('################\n',file=stdout()) + cat(iparams$chip.file, + iparams$control.file, + iparams$sep.range, + iparams$sep.peak, + iparams$ex.range, + iparams$n.nodes, + iparams$fdr, + iparams$npeak, + iparams$output.odir, + iparams$output.npeak.file, + iparams$output.rpeak.file, + iparams$output.rdata.file, + iparams$output.plot.file, + iparams$output.result.file, + iparams$replace.flag, + labels=c('ChIP data:','Control data:', 'strandshift(min):','strandshift(step):','strandshift(max)','user-defined peak shift', + 'exclusion(min):','exclusion(max):','num parallel nodes:','FDR threshold:','NumPeaks Threshold:','Output Directory:', + 'narrowPeak output file name:', 'regionPeak output file name:', 'Rdata filename:', + 'plot pdf filename:','result filename:','Overwrite files?:'), + fill=18, + file=stdout()) + cat('\n',file=stdout()) +} # end: print.run.parameters() + +check.replace.flag <- function(params){ +# =================================== +# Check if files exist +# =================================== +# If replace.flag is NOT set, check if output files exist and abort if necessary + if (! iparams$replace.flag) { + if (! is.na(iparams$output.npeak.file)) { + if (file.exists(iparams$output.npeak.file)) { + cat('narrowPeak file already exists. Aborting Run. Use -rf if you want to overwrite\n',file=stderr()) + q(save="no",status=1) + } + } + if (! is.na(iparams$output.rpeak.file)) { + if (file.exists(iparams$output.rpeak.file)) { + cat('regionPeak file already exists. Aborting Run. Use -rf if you want to overwrite\n',file=stderr()) + q(save="no",status=1) + } + } + if (! is.na(iparams$output.plot.file)) { + if (file.exists(iparams$output.plot.file)) { + cat('Plot file already exists. Aborting Run. Use -rf if you want to overwrite\n',file=stderr()) + q(save="no",status=1) + } + } + if (! is.na(iparams$output.rdata.file)) { + if (file.exists(iparams$output.rdata.file)) { + cat('Rdata file already exists. Aborting Run. Use -rf if you want to overwrite\n',file=stderr()) + q(save="no",status=1) + } + } + } +} + +# ############################################################################# +# MAIN FUNCTION +# ############################################################################# + +# Check number of arguments +minargs = 1; +maxargs = 17; +if (nargs < minargs | nargs > maxargs) { + print.usage() + q(save="no",status=1) +} + +# Parse arguments +# iparams$chip.file +# iparams$isurl.chip.file +# iparams$control.file +# iparams$isurl.control.file +# iparams$sep.range +# iparams$sep.peak +# iparams$ex.range +# iparams$n.nodes +# iparams$fdr +# iparams$npeak +# iparams$temp.dir +# iparams$output.odir +# iparams$output.npeak.file +# iparams$output.rpeak.file +# iparams$output.rdata.file +# iparams$output.plot.file +# iparams$output.result.file +# iparams$replace.flag +# iparams$clean.files.flag +iparams <- parse.arguments(args) + +# Print run parameters +print.run.params(iparams) + +# Check if output files exist +check.replace.flag(iparams) + +# curr.chip.file and curr.control.file always point to the original ChIP and control files on disk +# ta.chip.filename & ta.control.filename always point to the final but temporary versions of the ChIP and control files that will be passed to read.align + +# Download ChIP and control files if necessary to temp.dir +if (iparams$isurl.chip.file) { + curr.chip.file <- file.path(iparams$temp.dir, get.file.parts(iparams$chip.file)$fullname) # file is downloaded to temp.dir. Has same name as URL suffix + cat('Downloading ChIP file:',iparams$chip.file,"\n",file=stdout()) + if (system(paste('wget -N -q -P',iparams$temp.dir,iparams$chip.file)) != 0) { + stop('Error downloading ChIP file:',iparams$chip.file) + } +} else { + curr.chip.file <- iparams$chip.file # file is in original directory +} + +if (iparams$isurl.control.file) { + curr.control.file <- file.path(iparams$temp.dir, get.file.parts(iparams$control.file)$fullname) # file is downloaded to temp.dir. Has same name as URL suffix + cat('Downloading control file:',iparams$control.file,"\n",file=stdout()) + if (system(paste('wget -N -q -P',iparams$temp.dir,iparams$control.file)) != 0) { + stop('Error downloading Control file:',iparams$control.file) + } +} else { + curr.control.file <- iparams$control.file # file is in original directory +} + +# unzip ChIP and input files if required AND copy to temp directory +if (get.file.parts(curr.chip.file)$ext == '.gz') { + ta.chip.filename <- tempfile(get.file.parts(curr.chip.file)$name, tmpdir=iparams$temp.dir) # unzip file to temp.dir/[filename with .gz removed][randsuffix] + cat('Decompressing ChIP file\n',file=stdout()) + if (system(paste("gunzip -c",curr.chip.file,">",ta.chip.filename)) != 0) { + stop('Unable to decompress file:', iparams$chip.file) + } + if (iparams$clean.files.flag) { # Remove original file if clean.files.flag is set + file.remove(curr.chip.file) + } +} else { + ta.chip.filename <- tempfile(get.file.parts(curr.chip.file)$fullname, tmpdir=iparams$temp.dir) + if (iparams$clean.files.flag) { + file.rename(curr.chip.file,ta.chip.filename) # move file to temp.dir/[filename][randsuffix] + } else { + file.copy(curr.chip.file,ta.chip.filename) # copy file to temp.dir/[filename][randsuffix] + } +} + +if (! is.na(iparams$control.file)) { + if (get.file.parts(curr.control.file)$ext == '.gz') { + ta.control.filename <- tempfile(get.file.parts(curr.control.file)$name, tmpdir=iparams$temp.dir) # unzip file to temp.dir/[filename with .gz removed][randsuffix] + cat('Decompressing control file\n',file=stdout()) + if (system(paste("gunzip -c",curr.control.file,">",ta.control.filename)) != 0) { + stop('Unable to decompress file:', iparams$control.file) + } + if (iparams$clean.files.flag) { # Remove original file if clean.files.flag is set + file.remove(curr.control.file) + } + } else { + ta.control.filename <- tempfile(get.file.parts(curr.control.file)$fullname, tmpdir=iparams$temp.dir) # copy file to temp.dir/[filename][randsuffix] + + if (iparams$clean.files.flag) { + file.rename(curr.control.file,ta.control.filename) # move file to temp.dir/[filename][randsuffix] + } else { + file.copy(curr.control.file,ta.control.filename) # copy file to temp.dir/[filename][randsuffix] + } + } +} + +# Remove downloaded files +if (iparams$isurl.chip.file & file.exists(curr.chip.file)) { + file.remove(curr.chip.file) +} + +if (! is.na(iparams$control.file)) { + if (iparams$isurl.control.file & file.exists(curr.control.file)) { + file.remove(curr.control.file) + } +} + +# Load SPP library +library(spp) + +# Read ChIP tagAlign/BAM files +cat("Reading ChIP tagAlign/BAM file",iparams$chip.file,"\n",file=stdout()) +chip.data <- read.align(ta.chip.filename) +cat("ChIP data read length",chip.data$read.length,"\n",file=stdout()) +file.remove(ta.chip.filename) # Delete temporary file +if (length(chip.data$tags)==0) { + stop('Error in ChIP file format:', iparams$chip.file) +} +# Remove illegal chromosome names +if (! is.na(iparams$chrname.rm.pattern)) { + selectidx <- which(grepl(iparams$chrname.rm.pattern,names(chip.data$tags))==FALSE) + chip.data$tags <- chip.data$tags[selectidx] + chip.data$quality <- chip.data$quality[selectidx] +} +chip.data$num.tags <- sum(unlist(lapply(chip.data$tags,function(d) length(d)))) + +# Read Control tagAlign/BAM files +if (! is.na(iparams$control.file)) { + cat("Reading Control tagAlign/BAM file",iparams$control.file,"\n",file=stdout()) + control.data <- read.align(ta.control.filename) + file.remove(ta.control.filename) # Delete temporary file + if (length(control.data$tags)==0) { + stop('Error in control file format:', iparams$chip.file) + } + cat("Control data read length",control.data$read.length,"\n",file=stdout()) + # Remove illegal chromosome names + if (! is.na(iparams$chrname.rm.pattern)) { + selectidx <- which(grepl(iparams$chrname.rm.pattern,names(control.data$tags))==FALSE) + control.data$tags <- control.data$tags[selectidx] + control.data$quality <- control.data$quality[selectidx] + } + control.data$num.tags <- sum(unlist(lapply(control.data$tags,function(d) length(d)))) +} + +# Open multiple processes if required +if (is.na(iparams$n.nodes)) { + cluster.nodes <- NULL +} else { + library(snow) + cluster.nodes <- makeCluster(iparams$n.nodes) +} + +# ################################# +# Calculate cross-correlation for various strand shifts +# ################################# +cat("Calculating peak characteristics\n",file=stdout()) +# crosscorr +# $cross.correlation : Cross-correlation profile as an $x/$y data.frame +# $peak : Position ($x) and height ($y) of automatically detected cross-correlation peak. +# $whs: Optimized window half-size for binding detection (based on the width of the cross-correlation peak) +crosscorr <- get.binding.characteristics(chip.data, + srange=iparams$sep.range[c(1,3)], + bin=iparams$sep.range[2], + accept.all.tags=T, + cluster=cluster.nodes) +if (!is.na(iparams$n.nodes)) { + stopCluster(cluster.nodes) +} + +# Smooth the cross-correlation curve if required +cc <- crosscorr$cross.correlation +crosscorr$min.cc <- crosscorr$cross.correlation[ which.min(crosscorr$cross.correlation$y) , ] # minimum value and shift of cross-correlation +cat("Minimum cross-correlation value", crosscorr$min.cc$y,"\n",file=stdout()) +cat("Minimum cross-correlation shift", crosscorr$min.cc$x,"\n",file=stdout()) +sbw <- 2*floor(ceiling(5/iparams$sep.range[2]) / 2) + 1 # smoothing bandwidth +cc$y <- runmean(cc$y,sbw,alg="fast") + +# Compute cross-correlation peak +bw <- ceiling(2/iparams$sep.range[2]) # crosscorr[i] is compared to crosscorr[i+/-bw] to find peaks +peakidx <- (diff(cc$y,bw)>=0) # cc[i] > cc[i-bw] +peakidx <- diff(peakidx,bw) +peakidx <- which(peakidx==-1) + bw + +# exclude peaks from the excluded region +if ( is.nan(iparams$ex.range[2]) ) { + iparams$ex.range[2] <- chip.data$read.length+10 +} +peakidx <- peakidx[(cc$x[peakidx] < iparams$ex.range[1]) | (cc$x[peakidx] > iparams$ex.range[2])] +cc <- cc[peakidx,] + +# Find max peak position and other peaks within 0.9*max_peakvalue that are further away from maxpeakposition +maxpeakidx <- which.max(cc$y) +maxpeakshift <- cc$x[maxpeakidx] +maxpeakval <- cc$y[maxpeakidx] +peakidx <-which((cc$y >= 0.9*maxpeakval) & (cc$x >= maxpeakshift)) +cc <- cc[peakidx,] + +# sort the peaks and get the top 3 +sortidx <- order(cc$y,decreasing=TRUE) +sortidx <- sortidx[c(1:min(3,length(sortidx)))] +cc.peak <- cc[sortidx,] + +# Override peak shift if user supplies peak shift +if (! is.na(iparams$sep.peak)) { + cc.peak <- approx(crosscorr$cross.correlation$x,crosscorr$cross.correlation$y,iparams$sep.peak,rule=2) +} +cat("Peak cross-correlation value", paste(cc.peak$y,collapse=","),"\n",file=stdout()) +cat("Peak strand shift",paste(cc.peak$x,collapse=","),"\n",file=stdout()) + +# Reset values in crosscorr +crosscorr$peak$x <- cc.peak$x[1] +crosscorr$peak$y <- cc.peak$y[1] + +# Compute window half size +whs.thresh <- crosscorr$min.cc$y + (crosscorr$peak$y - crosscorr$min.cc$y)/3 +crosscorr$whs <- max(crosscorr$cross.correlation$x[crosscorr$cross.correlation$y >= whs.thresh]) +cat("Window half size",crosscorr$whs,"\n",file=stdout()) + +# Compute phantom peak coefficient +ph.peakidx <- which( ( crosscorr$cross.correlation$x >= ( chip.data$read.length - round(2*iparams$sep.range[2]) ) ) & + ( crosscorr$cross.correlation$x <= ( chip.data$read.length + round(1.5*iparams$sep.range[2]) ) ) ) +ph.peakidx <- ph.peakidx[ which.max(crosscorr$cross.correlation$y[ph.peakidx]) ] +crosscorr$phantom.cc <- crosscorr$cross.correlation[ph.peakidx,] +cat("Phantom peak location",crosscorr$phantom.cc$x,"\n",file=stdout()) +cat("Phantom peak Correlation",crosscorr$phantom.cc$y,"\n",file=stdout()) +crosscorr$phantom.coeff <- crosscorr$peak$y / crosscorr$phantom.cc$y +crosscorr$phantom.coeff <- crosscorr$peak$y / crosscorr$min.cc$y +cat("Normalized cross-correlation coefficient (NCCC)",crosscorr$phantom.coeff,"\n",file=stdout()) +crosscorr$rel.phantom.coeff <- (crosscorr$peak$y - crosscorr$min.cc$y) / (crosscorr$phantom.cc$y - crosscorr$min.cc$y) +cat("Relative Cross correlation Coefficient (RCCC)",crosscorr$rel.phantom.coeff,"\n",file=stdout()) +crosscorr$phantom.quality.tag <- NA +if ( (crosscorr$rel.phantom.coeff >= 0) & (crosscorr$rel.phantom.coeff < 0.25) ) { + crosscorr$phantom.quality.tag <- -2 +} else if ( (crosscorr$rel.phantom.coeff >= 0.25) & (crosscorr$rel.phantom.coeff < 0.5) ) { + crosscorr$phantom.quality.tag <- -1 +} else if ( (crosscorr$rel.phantom.coeff >= 0.5) & (crosscorr$rel.phantom.coeff < 1) ) { + crosscorr$phantom.quality.tag <- 0 +} else if ( (crosscorr$rel.phantom.coeff >= 1) & (crosscorr$rel.phantom.coeff < 1.5) ) { + crosscorr$phantom.quality.tag <- 1 +} else if ( (crosscorr$rel.phantom.coeff >= 1.5) ) { + crosscorr$phantom.quality.tag <- 2 +} +cat("Phantom Peak Quality Tag",crosscorr$phantom.quality.tag,"\n",file=stdout()) + +# Output result to result file if required +#Filename\tnumReads\tPeak_shift\tPeak_Correlation\tRead_length\tPhantomPeak_Correlation\tMin_Correlation_Shift\tMin_Correlation\tNormalized_CrossCorrelation_Coefficient\tRelative_CrossCorrelation_Coefficient\tQualityTag) +if (! is.na(iparams$output.result.file)) { + cat(get.file.parts(iparams$chip.file)$fullname, + chip.data$num.tags, + paste(cc.peak$x,collapse=","), + paste(cc.peak$y,collapse=","), + crosscorr$phantom.cc$x, + crosscorr$phantom.cc$y, + crosscorr$min.cc$x, + crosscorr$min.cc$y, + crosscorr$phantom.coeff, + crosscorr$rel.phantom.coeff, + crosscorr$phantom.quality.tag, + sep="\t", + file=iparams$output.result.file, + append=TRUE) + cat("\n", + file=iparams$output.result.file, + append=TRUE) +} + +# Save figure if required +if (! is.na(iparams$output.plot.file)) { + pdf(file=iparams$output.plot.file,width=5,height=5) + par(mar = c(4,3.5,2,0.5), mgp = c(1.5,0.5,0), cex = 0.8); + plot(crosscorr$cross.correlation, + type='l', + xlab=sprintf("strand-shift (%s)",paste(cc.peak$x,collapse=",")), + ylab="cross-correlation") + abline(v=cc.peak$x,lty=2,col=2) + abline(v=crosscorr$phantom.cc$x,lty=2,col=4) + title(main=get.file.parts(iparams$chip.file)$fullname, + sub=sprintf("NSC=%g,RSC=%g,Qtag=%d",crosscorr$phantom.coeff,crosscorr$rel.phantom.coeff,crosscorr$phantom.quality.tag)) + dev.off(); +} + +# Save RData file if required +if (! is.na(iparams$output.rdata.file)) { + save(iparams, + crosscorr, + cc.peak, + file=iparams$output.rdata.file); +} + +# ################################# +# Call peaks +# ################################# + +if ( !is.na(iparams$output.npeak.file) || !is.na(iparams$output.rpeak.file) ) { + + # Remove local tag anomalies + cat('Removing read stacks\n',file=stdout()) + chip.data <- remove.local.tag.anomalies(chip.data$tags) + control.data <- remove.local.tag.anomalies(control.data$tags) + + # Open multiple processes if required + if (is.na(iparams$n.nodes)) { + cluster.nodes <- NULL + } else { + cluster.nodes <- makeCluster(iparams$n.nodes) + } + + # Find peaks + cat('Finding peaks\n',file=stdout()) + if (!is.na(iparams$npeak)) { + iparams$fdr <- 0.96 + } + narrow.peaks <- find.binding.positions(signal.data=chip.data,control.data=control.data,fdr=iparams$fdr,method=tag.lwcc,whs=crosscorr$whs,cluster=cluster.nodes) + if (!is.na(iparams$n.nodes)) { + stopCluster(cluster.nodes) + } + cat(paste("Detected",sum(unlist(lapply(narrow.peaks$npl,function(d) length(d$x)))),"peaks"),"\n",file=stdout()) + + # Write to narrowPeak file + if (!is.na(iparams$output.npeak.file)) { + write.narrowpeak.binding(narrow.peaks,iparams$output.npeak.file,margin=round(crosscorr$whs/2),npeaks=iparams$npeak) + system(paste('gzip -f ',iparams$output.npeak.file)) + } + + # Compute and write regionPeak file + if (!is.na(iparams$output.rpeak.file)) { + region.peaks <- add.broad.peak.regions(chip.data,control.data,narrow.peaks,window.size=max(50,round(crosscorr$whs/4)),z.thr=10) + write.narrowpeak.binding(region.peaks,iparams$output.rpeak.file,margin=round(crosscorr$whs/2),npeaks=iparams$npeak) + system(paste('gzip -f ',iparams$output.rpeak.file)) + } + + # Save Rdata file + if (! is.na(iparams$output.rdata.file)) { + save(iparams, + crosscorr, + cc.peak, + narrow.peaks, + region.peaks, + file=iparams$output.rdata.file); + } + +} + +