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#!/usr/bin/env perl
use strict;
use warnings;
use lib("/usr/lib/trinityrnaseq/PerlLib");
use Pipeliner;
use File::Basename;
use Cwd;
use Carp;
use Getopt::Long qw(:config no_ignore_case bundling pass_through);
our $HISAT_HOME;
BEGIN {
if ($ENV{HISAT_HOME}) {
$HISAT_HOME = $ENV{HISAT_HOME};
}
else {
my $hisat_prog = `which hisat`;
if ($hisat_prog) {
chomp $hisat_prog;
$HISAT_HOME = dirname($hisat_prog);
}
else {
die "Error, cannot find hisat in PATH setting";
}
}
}
my $usage = <<__EOUSAGE__;
######################################################################
#
# Required:
# --genome <string> target genome to align to
# --reads <string> fastq files. If pairs, indicate both in quotes, ie. "left.fq right.fq"
#
# Optional:
# -N <int> max number of alignments to report. (default: 1)
# -G <string> GTF file for incorporating reference splice site info.
# --CPU <int> number of threads (default: 2)
# --out_prefix <string> output prefix (default: hisat)
# --run_as_single_reads if paired, run as single reads
#
#######################################################################
Include additional hisat arguments as additional command line parameters.
########################################################################
__EOUSAGE__
;
my ($genome, $reads);
my $CPU = 2;
my $help_flag;
my $out_prefix = "hisat";
my $gtf_file;
my $run_as_single_flag = 0;
my $num_top_hits = 1;
&GetOptions( 'h' => \$help_flag,
'genome=s' => \$genome,
'reads=s' => \$reads,
'CPU=i' => \$CPU,
'out_prefix=s' => \$out_prefix,
'G=s' => \$gtf_file,
'run_as_single_reads' => \$run_as_single_flag,
'N=i' => \$num_top_hits,
);
if ($help_flag) {
die $usage;
}
unless ($genome && $reads) {
die $usage;
}
main: {
my $hisat_index = "$genome.hisat.idx";
if (! -s "$hisat_index.1.bt2") {
## build hisat index
my $cmd = "$HISAT_HOME/hisat-build $genome $hisat_index";
&process_cmd($cmd);
}
my $splice_incl = "";
if ($gtf_file) {
my $gtf_splice = "$gtf_file.hisat.splice";
unless (-s $gtf_splice) {
my $cmd = "$HISAT_HOME/extract_splice_sites.py $gtf_file > $gtf_file.hisat.splice";
&process_cmd($cmd);
}
$splice_incl = " --known-splicesite-infile $gtf_splice ";
}
## run HISAT
$reads = &add_zcat_fifo_and_add_hisat_params($reads);
my $top_hits_count = "";
if ($num_top_hits > 1) {
$top_hits_count = " -k $num_top_hits ";
}
my @tmpfiles;
my $pipeliner = new Pipeliner(-verbose => 1);
my $cmd = "bash -c \"set -o pipefail && $HISAT_HOME/hisat -x $hisat_index -q $reads $splice_incl -p $CPU $top_hits_count @ARGV | gzip -c > $out_prefix.sam.gz\" ";
$pipeliner->add_commands( new Command($cmd, "$out_prefix.sam.gz.ok") );
push (@tmpfiles, "$out_prefix.sam.gz");
$cmd = "bash -c \"set -o pipefail && gunzip -c $out_prefix.sam.gz | samtools view -@ $CPU -F 4 -Sb -o $out_prefix.bam \"";
$pipeliner->add_commands( new Command($cmd, "$out_prefix.bam.ok") );
push (@tmpfiles, "$out_prefix.bam");
$cmd = "samtools sort -@ $CPU $out_prefix.bam -o $out_prefix.cSorted.bam";
$pipeliner->add_commands( new Command($cmd, "$out_prefix.cSorted.bam.ok") );
$pipeliner->run();
if (-s "$out_prefix.cSorted.bam") {
$cmd = "samtools index $out_prefix.cSorted.bam";
&process_cmd($cmd);
}
unlink(@tmpfiles);
exit(0);
}
####
sub add_zcat_fifo_and_add_hisat_params {
my ($reads) = @_;
$reads =~ s/^\s+|\s+$//g;
my @adj_reads_list;
my $counter = 0;
my @read_files = split(/\s+/, $reads);
my @updated_read_filenames;
foreach my $reads_file (@read_files) {
$counter++;
if ($reads_file =~ /\.gz$/) {
$reads_file = "<(zcat $reads_file)";
}
push (@updated_read_filenames, $reads_file);
# add decoration
$reads_file = (scalar(@read_files) == 2) ? "-$counter $reads_file" : "-U $reads_file";
push (@adj_reads_list, $reads_file);
}
if ($run_as_single_flag) {
return("-U " . join(",", @updated_read_filenames));
}
else {
my $adj_reads = join(" ", @adj_reads_list);
return($adj_reads);
}
}
####
sub process_cmd {
my ($cmd) = @_;
print STDERR "CMD: $cmd\n";
#return;
my $ret = system($cmd);
if ($ret) {
die "Error, cmd: $cmd died with ret ($ret)";
}
return;
}
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