#!/usr/bin/env perl use strict; use warnings; use Getopt::Std; my %opts = (t=>1, n=>64); getopts("SPADsp:R:x:t:", \%opts); die(' Usage: run-bwamem [options] [file2] Options: -p STR prefix for output files [inferred from file1] -R STR read group header line such as \'@RG\tID:foo\tSM:bar\' [null] -x STR read type: pacbio, ont2d or intractg [default] -t INT number of threads [1] -P input are paired-end reads if file2 absent -A skip HiSeq2000/2500 PE resequencing adapter trimming (via trimadap) -D skip duplicate marking (via samblaster) -S for SAM/BAM input, don\'t shuffle -s sort the output alignment (more RAM) Note: File type is determined by the file extension of the first input sequence file. Recognized file extensions: fasta, fa, fastq, fq, fasta.gz, fa.gz, fastq.gz, fq.gz, sam, sam.gz and bam. SAM/BAM input will be converted to FASTQ. ') if @ARGV < 2; warn("WARNING: many programs require read groups. Please specify with -R if you can.\n") unless defined($opts{R}); my $idx = $ARGV[0]; my $exepath = $0 =~/^\S+\/[^\/\s]+/? $0 : &which($0); my $root = $0 =~/^(\S+)\/[^\/\s]+/? $1 : undef; die "ERROR: failed to locate the 'bwa.kit' directory\n" if !defined($root); my $prefix; if (defined $opts{p}) { $prefix = $opts{p}; } elsif ($ARGV[1] =~ /^(\S+)\.(fastq|fq|fasta|fa|mag|sam|sam\.gz|mag\.gz|fasta\.gz|fa\.gz|fastq\.gz|fq\.gz|bam)$/) { $prefix = $1; } else { die("ERROR: failed to identify the prefix for output. Please specify -p.\n") } my $prefix_dir = $prefix =~ /^(\S+)\/[^\/\s]+$/? $1 : "."; die("ERROR: directory $prefix_dir is not writable. Please specify a new output prefix with -p.\n") unless (-w $prefix_dir); die("ERROR: failed to locate the BWA index. Please run '$root/bwa index -p $idx ref.fa'.\n") unless (-f "$idx.bwt" && -f "$idx.pac" && -f "$idx.sa" && -f "$idx.ann" && -f "$idx.amb"); if (@ARGV >= 3 && $ARGV[1] =~ /\.(bam|sam|sam\.gz)$/) { warn("WARNING: for SAM/BAM input, only the first sequence file is used.\n"); @ARGV = 2; } if (defined($opts{P}) && @ARGV >= 3) { warn("WARNING: option -P is ignored as there are two input sequence files.\n"); delete $opts{P}; } my $size = 0; my $comp_ratio = 3.; for my $f (@ARGV[1..$#ARGV]) { my @a = stat($f); my $s = $a[7]; $s *= $comp_ratio if $f =~ /\.(gz|bam)$/; $size += int($s) + 1; } my $is_pe = (defined($opts{P}) || @ARGV >= 3)? 1 : 0; my $is_sam = $ARGV[1] =~ /\.(sam|sam\.gz)$/? 1 : 0; my $is_bam = $ARGV[1] =~ /\.bam$/? 1 : 0; my $cmd = ''; if ($is_sam || $is_bam) { my $cmd_sam2bam = $is_sam? "$root/htsbox samview -uS $ARGV[1] \\\n" : "cat $ARGV[1] \\\n"; my $ntmps = int($size / 4e9) + 1; my $cmd_shuf = ($is_bam || $is_sam) && !defined($opts{S})? " | $root/htsbox bamshuf -uOn$ntmps - $prefix.shuf \\\n" : ""; my $cmd_bam2fq = ""; $cmd_bam2fq = " | $root/htsbox bam2fq -O " . (defined($opts{P})? "-s $prefix.out.se.fq.gz " : "") . "- \\\n"; $cmd = $cmd_sam2bam . $cmd_shuf . $cmd_bam2fq; } elsif (@ARGV >= 3) { $cmd = " | $root/seqtk mergepe $ARGV[1] $ARGV[2] \\\n"; } else { $cmd = "cat $ARGV[1] \\\n"; } $cmd .= " | $root/trimadap \\\n" unless defined($opts{A}); $cmd .= " | $root/bwa mem -t$opts{t} " . (defined($opts{x})? "-x $opts{x} " : "") . ($is_pe? "-P " : "") . (defined($opts{R})? "-R'$opts{R}' " : "") . "$ARGV[0] - 2> $prefix.log.bwamem \\\n"; $cmd .= " | $root/samblaster 2> $prefix.log.dedup \\\n" unless defined($opts{D}); my $has_hla = 0; if (-f "$ARGV[0].alt") { my $fh; open($fh, "$ARGV[0].alt") || die; while (<$fh>) { $has_hla = 1 if /^HLA-[^\s\*]+\*\d+/; } close($fh); my $hla_pre = $has_hla? "-p $prefix.out " : ""; $cmd .= " | $root/k8 $root/bwa-postalt.js $hla_pre$ARGV[0].alt \\\n"; } my $t_sort = $opts{t} < 4? $opts{t} : 4; $cmd .= defined($opts{s})? " | $root/samtools sort -@ $t_sort -m1G - $prefix.out;\n" : " | gzip -1 > $prefix.out.sam.gz;\n"; $cmd .= "$root/run-HLA $prefix.out;\n" if ($has_hla); print $cmd; sub which { my $file = shift; my $path = (@_)? shift : $ENV{PATH}; return if (!defined($path)); foreach my $x (split(":", $path)) { $x =~ s/\/$//; return "$x/$file" if (-x "$x/$file"); } return; }