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Copy pathTransDecoder.Predict
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executable file
·452 lines (310 loc) · 12.8 KB
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#!/usr/bin/env perl
=pod
=head1 NAME
L<Transdecoder|http://transdecoder.sourceforge.net> - Transcriptome Protein Prediction
=head1 USAGE
Required:
-t <string> transcripts.fasta
Common options:
--retain_long_orfs <int> retain all ORFs found that are equal or longer than these many nucleotides even if no other evidence
marks it as coding (default: 900 bp => 300aa)
--retain_pfam_hits <string> domain table output file from running hmmscan to search Pfam (see transdecoder.github.io for info)
Any ORF with a pfam domain hit will be retained in the final output.
--retain_blastp_hits <string> blastp output in '-outfmt 6' format.
Any ORF with a blast match will be retained in the final output.
--single_best_orf Retain only the single best ORF per transcript.
(Best is defined as having (optionally pfam and/or blast support) and longest orf)
--cpu <int> Use multiple cores for cd-hit-est. (default=1)
-G <string> genetic code (default: universal; see PerlDoc; options: Euplotes, Tetrahymena, Candida, Acetabularia, ...)
Advanced options
--train <string> FASTA file with ORFs to train Markov Mod for protein identification; otherwise
longest non-redundant ORFs used
-T <int> If no --train, top longest ORFs to train Markov Model (hexamer stats) (default: 500)
Note, 10x this value are first selected for use with cd-hit to remove redundancies,
and then this -T value of longest ORFs are selected from the non-redundant set.
=cut
=head1 Genetic Codes
See L<http://golgi.harvard.edu/biolinks/gencode.html>. These are currently supported:
universal (default)
Euplotes
Tetrahymena
Candida
Acetabularia
Mitochondrial-Canonical
Mitochondrial-Vertebrates
Mitochondrial-Arthropods
Mitochondrial-Echinoderms
Mitochondrial-Molluscs
Mitochondrial-Ascidians
Mitochondrial-Nematodes
Mitochondrial-Platyhelminths
Mitochondrial-Yeasts
Mitochondrial-Euascomycetes
Mitochondrial-Protozoans
=cut
use strict;
use warnings;
use FindBin;
use Pod::Usage;
use Getopt::Long qw(:config posix_default no_ignore_case bundling pass_through);
use Data::Dumper;
use List::Util qw (min max);
use File::Basename;
use lib ("$FindBin::RealBin/PerlLib");
use POSIX qw(ceil);
use Gene_obj;
use Nuc_translator;
use Fasta_reader;
use Longest_orf;
my $UTIL_DIR = "$FindBin::RealBin/util";
$ENV{PATH} = "$UTIL_DIR/bin:$ENV{PATH}";
my ($cd_hit_est_exec) = &check_program('cd-hit-est');
my ($transcripts_file,$train_file);
my $top_ORFs_train = 500;
my $help;
my $verbose;
my $search_pfam = "";
my ($reuse,$pfam_out);
my $RETAIN_LONG_ORFS = 900;
my $retain_pfam_hits_file;
my $retain_blastp_hits_file;
my $cpu = 1;
my $MPI_DEBUG = 1;
my $single_best_orf_flag = 0;
my $genetic_code = "";
&GetOptions( 't=s' => \$transcripts_file,
'train:s' => \$train_file,
'h' => \$help,
'v' => \$verbose,
'T=i' => \$top_ORFs_train,
'search_pfam=s' => \$search_pfam,
'reuse' => \$reuse,
'retain_long_orfs=i' => \$RETAIN_LONG_ORFS,
'debug' => \$MPI_DEBUG,
'retain_pfam_hits=s' => \$retain_pfam_hits_file,
'retain_blastp_hits=s' => \$retain_blastp_hits_file,
'cpu=i' => \$cpu,
'single_best_orf' => \$single_best_orf_flag,
'G=s' => \$genetic_code,
);
pod2usage(-verbose => 2, -output => \*STDERR) if ($help);
if (@ARGV) {
die "Error, don't understand options: @ARGV";
}
our $SEE = $verbose;
pod2usage(-verbose => 2, -output => \*STDERR, -message => "No transcript file (-t)\n") unless ($transcripts_file && -s $transcripts_file);
if ($genetic_code) {
$genetic_code = " --genetic_code $genetic_code";
}
main: {
my $workdir = basename($transcripts_file) . ".transdecoder_dir";
unless (-d $workdir) {
die "Error, cannot find directory: $workdir, be sure to first run TransDecoder.LongOrfs before TransDecoder.Predict\n\n";
}
my $prefix = "$workdir/longest_orfs";
my $cds_file = "$prefix.cds";
my $gff3_file = "$prefix.gff3";
my $pep_file = "$prefix.pep";
## Train a Markov model based on user-provided file or longest candidate CDS sequences, score all candidates, and select the final set.
my $top_cds_file;
if ($train_file) {
if (! -s $train_file) {
die "Error, cannot locate train file: $train_file";
}
$top_cds_file = $train_file;
}
else {
$top_cds_file = "$cds_file.top_${top_ORFs_train}_longest";
my $checkpoint = "$top_cds_file.ok";
if (! -e $checkpoint) {
# to speed things up only check for redundancy up to x the number of entries we want
my $red_num = $top_ORFs_train * 10;
my $red_num_cds_longest_file = "$cds_file.top_longest_${red_num}";
&process_cmd("$UTIL_DIR/get_top_longest_fasta_entries.pl $cds_file $red_num > $red_num_cds_longest_file");
&process_cmd("$cd_hit_est_exec -r 1 -i $red_num_cds_longest_file -T $cpu -c 0.80 -o $red_num_cds_longest_file.nr80 -M 0 ");
&process_cmd("$UTIL_DIR/get_top_longest_fasta_entries.pl $red_num_cds_longest_file.nr80 $top_ORFs_train > $top_cds_file");
&process_cmd("touch $checkpoint");
}
}
# get hexamer scores
my $hexamer_scores_file = "$workdir/hexamer.scores";
my $hexamer_checkpoint = "$hexamer_scores_file.ok";
if (! -e $hexamer_checkpoint) {
my $base_freqs_file = "$workdir/base_freqs.dat";
my $cmd = "$UTIL_DIR/seq_n_baseprobs_to_logliklihood_vals.pl $top_cds_file $base_freqs_file > $hexamer_scores_file";
&process_cmd($cmd);
&process_cmd("touch $hexamer_checkpoint");
}
# score all cds entries
my $cds_scores_file = "$cds_file.scores";
my $cds_scores_checkpoint = "$cds_scores_file.ok";
if (! -e $cds_scores_checkpoint) {
my $cmd = "$UTIL_DIR/score_CDS_liklihood_all_6_frames.pl $cds_file $hexamer_scores_file > $cds_scores_file";
&process_cmd($cmd);
&process_cmd("touch $cds_scores_checkpoint");
}
## Retain those that have pfam matches
my %has_pfam_hit;
if ($retain_pfam_hits_file) {
%has_pfam_hit = &parse_pfam_hits($retain_pfam_hits_file);
}
my %has_blastp_hit;
if ($retain_blastp_hits_file) {
%has_blastp_hit = &parse_blastp_hits_file($retain_blastp_hits_file);
}
# get accs for best entries
my $acc_file = "$cds_file.scores.selected";
{
my %att_counter;
open (my $ofh, ">$acc_file") or die "Error, cannot write to $acc_file";
open (my $ifh, "$cds_file.scores") or die "Error, cannot open file $cds_file.scores";
while (<$ifh>) {
chomp;
my ($acc, $orf_length, @scores) = split(/\t/);
my @ATTS;
my $score_1 = shift @scores;
my $max_score_other_frame = max(@scores);
my $keep_flag = 0;
if ($has_pfam_hit{$acc}) {
$keep_flag = 1;
push (@ATTS, "PFAM");
print STDERR "-$acc flagged as having a pfam domain.\n" if $verbose;
}
if ($has_blastp_hit{$acc}) {
$keep_flag = 1;
push (@ATTS, "BLASTP");
print STDERR "-$acc flagged as having a blastp match.\n" if $verbose;
}
if ($orf_length >= $RETAIN_LONG_ORFS) {
$keep_flag = 1;
push (@ATTS, "LONGORF");
}
if ($score_1 > 0 && $score_1 > $max_score_other_frame) {
$keep_flag = 1;
push (@ATTS, "FRAMESCORE");
}
if ($keep_flag) {
print $ofh "$acc\n";
my $att_string = join("|", sort @ATTS);
$att_counter{$att_string}++;
}
}
close $ifh;
close $ofh;
# report on the categories of the selected ORFs
print STDERR "\n#####################\nCounts of kept entries according to attributes:\n";
foreach my $att (reverse sort {$att_counter{$a}<=>$att_counter{$b}} keys %att_counter) {
my $count = $att_counter{$att};
print STDERR join("\t", $att, $count) . "\n";
}
print STDERR "########################\n\n\n";
}
# index the current gff file:
my $cmd = "$UTIL_DIR/index_gff3_files_by_isoform.pl $gff3_file";
&process_cmd($cmd);
# retrieve the best entries:
$cmd = "$UTIL_DIR/gene_list_to_gff.pl $acc_file $gff3_file.inx > $cds_file.best_candidates.gff3";
&process_cmd($cmd);
##############################
## Generate the final outputs.
##############################
my $final_output_prefix = basename($transcripts_file) . ".transdecoder";
{
# exclude shadow orfs (smaller orfs in different reading frame that are eclipsed by longer orfs)
$cmd = "$UTIL_DIR/remove_eclipsed_ORFs.pl $cds_file.best_candidates.gff3 > $cds_file.eclipsed_removed.gff3";
my $target_final_file = "$cds_file.eclipsed_removed.gff3";
&process_cmd($cmd);
if ($single_best_orf_flag) {
$cmd = "$UTIL_DIR/single_best_ORF_per_transcript.pl ";
if ($retain_blastp_hits_file) {
$cmd .= " --blast_hits $retain_blastp_hits_file ";
}
if ($retain_pfam_hits_file) {
$cmd .= " --pfam_hits $retain_pfam_hits_file ";
}
$cmd .= " --gff3_file $cds_file.eclipsed_removed.gff3 > $cds_file.single_best_orf.gff3";
&process_cmd($cmd);
$target_final_file = "$cds_file.single_best_orf.gff3";
}
&process_cmd("cp $target_final_file $final_output_prefix.gff3");
## write final outputs:
## make a BED file for viewing in IGV
my $gff3_file = "$final_output_prefix.gff3";
my $bed_file = $gff3_file;
$bed_file =~ s/\.gff3$/\.bed/;
$cmd = "$UTIL_DIR/gff3_file_to_bed.pl $gff3_file > $bed_file";
&process_cmd($cmd);
# make a peptide file:
my $best_pep_file = $gff3_file;
$best_pep_file =~ s/\.gff3$/\.pep/;
$cmd = "$UTIL_DIR/gff3_file_to_proteins.pl --gff3 $gff3_file --fasta $transcripts_file $genetic_code > $best_pep_file";
&process_cmd($cmd);
# make a CDS file:
my $best_cds_file = $best_pep_file;
$best_cds_file =~ s/\.pep$/\.cds/;
$cmd = "$UTIL_DIR/gff3_file_to_proteins.pl --gff3 $gff3_file --fasta $transcripts_file --seqType CDS $genetic_code > $best_cds_file";
&process_cmd($cmd);
}
print STDERR "transdecoder is finished. See output files $final_output_prefix.\*\n\n\n";
exit(0);
}
####
sub process_cmd {
my ($cmd) = @_;
print "CMD: $cmd\n";
my $ret = system($cmd);
if ($ret) {
die "Error, cmd: $cmd died with ret $ret";
}
return;
}
####
sub parse_pfam_hits {
my ($pfam_hits_file) = @_;
my %has_pfam_hit;
if (! -e $pfam_hits_file) {
die "Error, cannot find pfam hits file: $pfam_hits_file";
}
print "PFAM output found and processing...\n";
# capture those proteins having pfam hits
open (my $fh, $pfam_hits_file) or die "Error, cannot open file: $pfam_hits_file";
while (my $ln=<$fh>) {
next if $ln=~/^\#/;
my @x = split(/\s+/,$ln);
next unless $x[3]; # domtbl
my $orf_acc = $x[3];
$has_pfam_hit{$orf_acc} = 1;
}
close $fh;
return(%has_pfam_hit);
}
####
sub parse_blastp_hits_file {
my ($blastp_file) = @_;
unless (-e $blastp_file) {
die "Error, cannot find file $blastp_file";
}
my %blastp_hits;
open (my $fh, $blastp_file) or die "Error, cannot open file $blastp_file";
while (<$fh>) {
chomp;
my @x = split(/\t/);
my $id = $x[0];
$blastp_hits{$id} = 1;
}
close $fh;
return(%blastp_hits);
}
sub check_program() {
my @paths;
foreach my $prog (@_) {
my $path = `which $prog`;
unless ($path =~ /\w/) {
die "Error, path to a required program ($prog) cannot be found\n\n"
}
chomp($path);
$path = readlink($path) if -l $path;
push( @paths, $path );
}
return @paths;
}