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|
#!/usr/bin/env perl
use strict;
use warnings;
use lib ("/usr/lib/trinityrnaseq/PerlLib");
use Process_cmd;
use Getopt::Long qw(:config no_ignore_case bundling pass_through);
use Pipeliner;
use File::Basename;
my $CPU = 2;
my $usage = <<__EOUSAGE;
############################################################
#
# Required:
#
# --genome <string> target genome.fasta file
#
# --samples_file <string> Trinity samples file
#
# Optional:
#
# --gtf <string> annotation in gtf format
#
# --CPU <int> multithreading (default: $CPU)
#
# --nameSorted sorts bam by read name
#
###########################################################
__EOUSAGE
;
my $help_flag;
my $genome_fa;
my $annotation_gtf;
my $samples_file;
my $nameSorted;
&GetOptions ( 'h' => \$help_flag,
'genome=s' => \$genome_fa,
'gtf=s' => \$annotation_gtf,
'samples_file=s' => \$samples_file,
'CPU=i' => \$CPU,
'nameSorted' => \$nameSorted,
);
if ($help_flag) { die $usage; }
unless ($genome_fa && $samples_file) {
die $usage;
}
$genome_fa = &Pipeliner::ensure_full_path($genome_fa);
$samples_file = &Pipeliner::ensure_full_path($samples_file);
$annotation_gtf = &Pipeliner::ensure_full_path($annotation_gtf) if $annotation_gtf;
main: {
my @read_sets = &parse_samples_file($samples_file);
## align reads to the mini-genome using hisat2
###########################
# first, build genome index
my $pipeliner = new Pipeliner(-verbose => 1);
if ($annotation_gtf) {
$pipeliner->add_commands(new Command("hisat2_extract_splice_sites.py $annotation_gtf > $annotation_gtf.ss",
"$annotation_gtf.ss.ok"));
$pipeliner->add_commands(new Command("hisat2_extract_exons.py $annotation_gtf > $annotation_gtf.exons",
"$annotation_gtf.exons.ok"));
$pipeliner->add_commands(new Command("hisat2-build --exon $annotation_gtf.exons --ss $annotation_gtf.ss -p $CPU $genome_fa $genome_fa",
"$genome_fa.hisat2.build.ok"));
$pipeliner->run();
}
else {
$pipeliner->add_commands(new Command("hisat2-build -p $CPU $genome_fa $genome_fa",
"$genome_fa.hisat2.nogtf.build.ok"));
$pipeliner->run();
}
#####################
## now run alignments
my $aln_checkpoints_dir = "hisat2_aln_chkpts." . basename($genome_fa);
unless (-d $aln_checkpoints_dir) {
mkdir($aln_checkpoints_dir) or die "Error, cannot mkdir $aln_checkpoints_dir";
}
my $sorted_opt = "";
my $sorted_token = "c";
if ($nameSorted) {
$sorted_opt = "-n";
$sorted_token = "n";
}
foreach my $read_set_aref (@read_sets) {
my ($sample_id, $left_fq, $right_fq) = @$read_set_aref;
my $bamfile = "$sample_id.${sorted_token}Sorted.hisat2." . basename($genome_fa) . ".bam";
$pipeliner->add_commands(new Command("bash -c \"set -eof pipefail; hisat2 --dta -x $genome_fa -p $CPU -1 $left_fq -2 $right_fq | samtools view -Sb -F 4 | samtools sort $sorted_opt -o $bamfile \" ",
"$aln_checkpoints_dir/$bamfile.ok"));
}
$pipeliner->run();
exit(0);
}
####
sub parse_samples_file {
my ($samples_file) = @_;
my @samples;
open(my $fh, $samples_file) or die "Error, cannot open file $samples_file";
while (<$fh>) {
chomp;
my @x = split(/\t/);
my ($cond, $rep, $fq_a, $fq_b) = @x;
$fq_a = &Pipeliner::ensure_full_path($fq_a);
$fq_b = &Pipeliner::ensure_full_path($fq_b) if $fq_b;
push (@samples, [$rep, $fq_a, $fq_b]);
}
close $fh;
return (@samples);
}
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