THIS IS A PRIMARY OUTPUT THAT SHOULD UNDERGO FURTHER QUALITY FILTERING TO GET RID OFF POTENTIAL FALSE POSITIVE DOMAINS

WHAT IT DOES

This tool uses external aligning programme LAST and RepeatExplorer database of TE protein domains(REXdb) (Viridiplantae and Metazoa)

Lastal runs similarity search to find hits between query DNA sequence and our database of protein domains from all Viridiplantae repetitive elements. Hits with overlapping positions in the sequence (even through other hits) forms a cluster which represents one potential protein domain. Strand orientation is taken into consideration when forming the clusters which means each cluster is built from forward or reverse stranded hits exclusively. The clusters are subsequently processed separately; within one cluster positions are scanned base-by-base and classification strings are assigned for each of them based on the database sequences which were mapped on that place. These asigned classification strings consist of a domain type as well as class and lineage of the repetitive element where the database protein comes from. Different classification levels are separated by "|" character. Every hit is scored according to the scoring matrix used for DNA-protein alignment (BLOSUM80). For single position only the hits reaching certain percentage (80% by default) of the overall best score within the whole cluster are reported. One cluster of overlapping hits represents one domain region and is recorded as one line in the resulting GFF3 file. Regarding the classition strings assigned to one region (cluster) there are three situations that can occur:

1. There is a single classification string assigned to each position as well as classifications along all the positions in the region are mutually uniform, in this case domain's final classification is equivalent to this unique classification.
2. There are multiple classification strings assigned to one cluster, i.e. one domain, which leads to classification to the common (less specific) level of all the strings
3. There is a conflict at the domain type level, domains are reported with slash (e.g. RT/INT) and the classification is in this case ambiguous

There are 2 outputs produced by this tool:

1. GFF3 file of all proteins domains built from all hits found by LAST. Domains are reported per line as regions (start - end) on the original DNA sequence including the seq ID, alignment score and strand orientation. The last "Attributes" column contains several semicolon-separated information related to annotation, repetitive classification, alignment and its quality. This file can undergo further filtering using Protein Domain Filter tool

• Attributes reported always:

  Name

  type of domain; if ambiguous reported with slash

  Final_classification

  definite classification based on all partial classifications of Region_hits_classifications attribute or "Ambiguous_domain" when there is an ambiguous domain type

  Region_Hits_Classifications

  all hits classifications (comma separated) from a certain domain region that reach the set score threshold; in case of multiple annotations the square brackets indicate the number of bases having this particular classification

• Attributes only reported in case of unambiguous domain type (all the attributes including quality information are related to the Best_Hit of the region):

  Best_hit

  classification and position of the best alignment with the highest score within the cluster; in the square brackets is the percentage of the whole cluster range that this best hit covers

  Best_Hit_DB_Pos

  showing which part of the original datatabase domain corresponding to the Best Hit was aligned on query DNA (e.g. Best_Hit_DB_Pos=17:75of79 means the Best Hit reported in GFF represents region from 17th to 75th of total 79 aminoacids in the original domain from the database)

  DB_Seq

  database protein sequence of the best hit mapped to the query DNA

  Query_Seq

  alignment sequence of the query DNA for the best hit

  Identity

  ratio of identical amino acids in alignment sequence to the length of alignment

  Similarity

  ratio of alignment positions with positive score (according to the scoring matrix) to the length of alignment

  Relat_Length

  ratio of gapless length of the aligned protein sequence to the whole length of the database protein

  Relat_Interruptions

  number of the interruptions (frameshifts + stop codons) in aligned translated query sequence per each starting 100 AA

  Hit_to_DB_Length

  proportion of alignment length to the original length of the protein domain from database

!NOTE: Tool can in average process 0.5 Gbps of the DNA sequence per day. This is only a rough estimate and it is highly dependent on input data (repetive elements occurence) as well as computing resources. Maximum running time of the tool is 7 days.