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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);
my $usage = <<__EOUSAGE__;
######################################################################
#
# Required:
# --genome <string> target genome to align to
# --gtf|G <string> GTF file for incorporating reference splice site info. (recommended)
#
# --reads <string> fastq files. If pairs, indicate both in quotes, ie. "left.fq right.fq"
#
# Optional:
# --CPU <int> number of threads (default: 2)
# --out_prefix <string> output prefix (default: star)
# --out_dir <string> output directory (default: current working directory)
# --star_path <string> full path to the STAR program to use.
# --patch <string> genomic targets to patch the genome fasta with.
# --chim_search include Chimeric.junction outputs
#
#######################################################################
__EOUSAGE__
;
my ($genome, $reads);
my $CPU = 2;
my $help_flag;
my $out_prefix = "star";
my $gtf_file;
my $out_dir;
my $ADV = 0;
my $star_path = "STAR";
my $patch;
my $chim_search;
&GetOptions( 'h' => \$help_flag,
'genome=s' => \$genome,
'reads=s' => \$reads,
'CPU=i' => \$CPU,
'out_prefix=s' => \$out_prefix,
'gtf|G=s' => \$gtf_file,
'out_dir=s' => \$out_dir,
'ADV' => \$ADV,
'star_path=s' => \$star_path,
'patch=s' => \$patch,
'chim_search' => \$chim_search,
);
unless ($genome && $reads) {
die $usage;
}
if ($help_flag) {
die $usage;
}
if (@ARGV) {
die "Error, cannot recognize opts: @ARGV";
}
my $star_prog = `which $star_path`;
chomp $star_prog;
unless ($star_prog =~ /\w/) {
die "Error, cannot locate STAR program. Be sure it's in your PATH setting. ";
}
main: {
## ensure all full paths
$genome = &Pipeliner::ensure_full_path($genome);
$gtf_file = &Pipeliner::ensure_full_path($gtf_file) if $gtf_file;
my @read_files = split(/\s+/, $reads);
foreach my $read_file (@read_files) {
if ($read_file) {
$read_file = &Pipeliner::ensure_full_path($read_file);
}
}
$reads = join(" ", @read_files);
if ($out_dir) {
unless (-d $out_dir) {
mkdir $out_dir or die "Error, cannot mkdir $out_dir";
}
chdir $out_dir or die "Error, cannot cd to $out_dir";
}
my $star_index = "$genome.star.idx";
if (! -e "$star_index/build.ok") {
## build star index
unless (-d $star_index) {
mkdir($star_index) or die "Error, cannot mkdir $star_index";
}
my $cmd = "$star_prog --runThreadN $CPU --runMode genomeGenerate --genomeDir $star_index "
. " --genomeFastaFiles $genome "
. " --limitGenomeGenerateRAM 40419136213 ";
if ($gtf_file) {
$cmd .= " --sjdbGTFfile $gtf_file "
. " --sjdbOverhang 100 ";
}
&process_cmd($cmd);
&process_cmd("touch $star_index/build.ok");
}
## run STAR
my @tmpfiles;
my $pipeliner = new Pipeliner(-verbose => 1);
my $cmd = "$star_prog "
. " --runThreadN $CPU "
. " --genomeDir $star_index "
. " --outSAMtype BAM SortedByCoordinate "
. " --runMode alignReads "
. " --readFilesIn $reads "
. " --twopassMode Basic "
. " --alignSJDBoverhangMin 10 "
. " --outSAMstrandField intronMotif "
. " --outSAMunmapped Within "
. " --limitBAMsortRAM 20000000000";
if ($chim_search) {
$cmd .= " --chimJunctionOverhangMin 12 "
. " --chimSegmentMin 12 "
. " --chimSegmentReadGapMax parameter 3 "
}
if ($patch) {
$cmd .= " --genomeFastaFiles $patch ";
}
$cmd .= " --alignSJstitchMismatchNmax 5 -1 5 5 "; #which allows for up to 5 mismatches for non-canonical GC/AG, and AT/AC junctions, and any number of mismatches for canonical junctions (the default values 0 -1 0 0 replicate the old behavior (from AlexD)
if ($reads =~ /\.gz$/) {
$cmd .= " --readFilesCommand 'gunzip -c' ";
}
$pipeliner->add_commands( new Command($cmd, "star_align.ok") );
my $bam_outfile = "Aligned.sortedByCoord.out.bam";
my $renamed_bam_outfile = "$out_prefix.sortedByCoord.out.bam";
$pipeliner->add_commands( new Command("mv $bam_outfile $renamed_bam_outfile", "$renamed_bam_outfile.ok") );
$pipeliner->add_commands( new Command("samtools index $renamed_bam_outfile", "$renamed_bam_outfile.bai.ok") );
$pipeliner->run();
exit(0);
}
####
sub process_cmd {
my ($cmd) = @_;
print STDERR "CMD: $cmd\n";
#return;
my $ret = system($cmd);
if ($ret) {
die "Error, cmd: $cmd died with ret ($ret)";
}
return;
}
|
