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ped.pl
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#!/usr/bin/perl
#
# This file is a script for Polymorphic Edge Detection.
#
# Copyright (C) 2019 National Agriculture and Food Research Organization. All Rights Reserved.
#
# License: refer to https://github.com/akiomiyao/ped
#
# Author: Akio Miyao <[email protected]>
#
$usage = '
ped.pl - program for polymorphic edge detection.
e.g. perl ped.pl target=ERR194147,ref=hg38
Results will be saved in the target directory.
If you want to detect polymorphisms between target and control,
perl ped.pl target=ERR3063487,control=ERR3063486,ref=WBcel235
If you want to specify the working directory,
perl ped.pl target=ERR194147,ref=hg38,wd=/home/you/work
If you want to specify both the working directory and tmp directory,
perl ped.pl target=ERR194147,ref=hg38,wd=/home/you/work,tmpdir=/mnt/ssd
To make only reference data.
perl ped.pl ref=hg38
To make the special reference data absent in config file.
mkdir refname
cp refname.fasta refname
perl ped.pl ref=refname,file=refname.fasta
If you want to set maximum number of threads (processes),
perl ped.pl target=ERR194147,ref=hg38,thread=14
In the case of file open error, reduce muximum number of threads.
In the case of over the threads (processes) number limit,
perl ped.pl target=ERR194147,ref=hg38,thread=14,wait=1
Default wait time to make new process is 0.1 seconds.
For kmer method,
perl ped.pl target=ERR3063487,control=ERR3063486,ref=WBcel235,method=kmer
For cnv detection,
perl ped.pl target=ttm2,control=ttm5,ref=IRGSP1.0,method=cnv
If control is not specified, reference seqeunce will be used as a control.
If short reads have different length sequences, sequence will be clipped.
If you want to specify the clipping length,
perl ped.pl target=SRR11542243,ref=SARS-CoV-2,clipping=100
If clipping value is omitted, the value will be adjusted by the program.
To know the optimal clipping length,
perl check_length.pl SRR11542243
An example,
perl download.pl accession=SRR11542243
perl ped.pl target=SRR11542243,ref=SARS-CoV-2
It takes 5 minuits.
SRR11542243.vcf in SRR11542243 directioy is the result.
SRR11542243.full.vcf : Unfilterd but verified result.
SRR11542243.count.vcf : Unfilterd raw result.
SRR11542243.bi.snp : PED format SNP result.
SRR11542243.sv : PED format structural variation result.
Some polymorphisms from mixed genome sequence like as viruses from human
will be filtered out during verify process.
In this casee, count.vcf is more informative.
perl search.pl target=SRR11542243,chr=NC_045512.2,pos=11185
will show bidirectional alignments.
Analysis for your fastq files
mkdir target_name
mkdir target_name/read
cp somewhere/fastq_files target_name/read
perl ped.pl.target=target_name,ref=ref_name
Results will be saved in the target directory.
Complessed fastq files with gz, bzip2 and xz format can be analyzed.
';
my $sort_opt = "-S 1M";
@nuc = ('A', 'C', 'G', 'T');
$ENV{LANG} = "C";
$start_time = time;
$start_timestamp = `date`;
$log = "script : ped.pl
argument : $ARGV[0]\n";
if ($ARGV[0] =~ /target|file/){
my @arg = split(',', $ARGV[0]);
foreach (sort @arg){
next if $_ eq "";
my ($name, $val) = split('=', $_);
$$name = $val;
$log .= "$name : $val\n";
}
}elsif ($ARGV[0] =~ /ref/){
my ($name, $val) = split('=', $ARGV[0]);
$$name = $val;
$log .= "$name : $val\n";
}elsif ($ARGV[1] ne ""){
$target = $ARGV[0];
$control = $ARGV[1];
$ref = $ARGV[2];
$method = $ARGV[3];
}elsif ($ENV{target} ne ""){
$target = $ENV{target};
$control = $ENV{control};
$ref = $ENV{ref};
$method = $ENV{method};
$wd = $ENV{wd};
$tmpdir = $ENV{tmpdir};
$max_semaphore = $ENV{thread};
$clipping = $ENV{clipping};
}else{
print $usage;
}
$cwd = `pwd`;
chomp($cwd);
if ($wd eq ""){
$wd = $cwd;
}
$uname = `uname`;
chomp($uname);
if ($method eq ""){
if ($ref eq ""){
$method = "kmer";
}else{
$method = "bidirectional";
}
$log .= "method : $method\n";
}
$zcat = "zcat";
if ($uname eq "FreeBSD"){
die "curl not found. Please install curl." if ! -e "/usr/local/bin/curl";
open(IN, "sysctl kern.smp.cpus |");
while(<IN>){
chomp;
$processor = (split(': ', $_))[1];
}
close(IN);
}elsif($uname eq "Darwin"){
$zcat = "gzcat";
open(IN, "sysctl hw.logicalcpu |");
while(<IN>){
chomp;
$processor = (split(': ', $_))[1];
}
close(IN);
}elsif($uname eq "Linux"){
die "curl not found. Please install curl.
sudo apt install curl (Ubuntu)
sudo yum install curl (CentOS)
" if ! -e "/usr/bin/curl";
open(IN, "/proc/cpuinfo");
while(<IN>){
$processor ++ if /processor/;
}
close(IN);
}else{
die "Operating system is unknown.\n";
}
if ($uname eq "Darwin"){
$max_process = 1;
}else{
$max_process = $processor;
}
$max_process = 3 if $max_process eq "";
$max_process = $thread if $thread ne "";
$log .= "max process : $max_process\n";
$refdir = "$wd/$ref";
if ($tmpdir eq "" and $target ne ""){
system("mkdir $wd/$target/tmp") if ! -e "$wd/$target/tmp";
$tmpdir = "$wd/$target/tmp";
}
$control = $ref if $control eq "" or $control eq "default";
if (! -e "config"){
system("curl -O https://raw.githubusercontent.com/akiomiyao/ped/master/config");
}
open(IN, "config");
while(<IN>){
chomp;
my @row = split('\t', $_);
if ($row[1] eq "description"){
$desc{$row[0]} = $row[2];
}elsif($row[1] eq "wget"){
$curl{$row[0]} = $row[2];
}elsif($row[1] eq "file"){
$file{$row[0]} = $row[2];
}elsif($row[0] eq $ref && $row[1] eq "chromosome"){
@row = split;
if ($row[3] != 0){
for ($i = $row[2]; $i <= $row[3]; $i++){
push(@chr, $i);
}
}
if ($row[4] ne ""){
foreach ($i = 4; $i <= $#row; $i++){
push(@chr, $row[$i]);
}
}
}
}
close(IN);
if ($ARGV[0] eq ""){
print " Currently, reference genomes listed below are suported.
Name Description\n";
foreach (sort keys %desc){
my $name = "$_ ";
$name = substr($name, 0, 16);
print " $name $desc{$_}\n";
}
print "
Author: Akio Miyao <miyao\@affrc.go.jp>
Scripts: https://github.com/akiomiyao/ped
Web page: https://akiomiyao.github.io/ped
Docker: https://hub.docker.com/r/akiomiyao/ped
";
exit;
}
if ($sub ne ""){
my $child = "$wd/child/child.$$";
system("touch $child");
&$sub($arg);
system("rm $child");
exit;
}else{
if (-e "$wd/child"){
system("rm -r $wd/child");
}
system("mkdir $wd/child");
}
if($ref ne ""){
system("mkdir $wd/$ref") if ! -e "$wd/$ref";
system("mkdir $wd/$ref/tmp") if ! -e "$wd/$ref/tmp";
open(REPORT, "> $wd/$ref/$ref.log") if $target eq "";
}
if ($target ne ""){
open(REPORT, "> $wd/$target/$target.report");
open(LOG, "> $wd/$target/$target.log");
}
print LOG $log;
report("Job begin: $method method");
if ($ref ne "" and ! (-s "$wd/$ref/ref20_uniq.TTT.gz" > 100 and -s "$wd/$ref/sort_uniq/$ref.sort_uniq.TTT.gz" > 100)){
&mkRef;
}
if ($target eq ""){
&finish;
}
if ($chr[0] eq ""){
opendir(REF, $refdir);
foreach(readdir(REF)){
chomp;
if (/^chr/){
($chr = $_) =~ s/^chr//;
push(@chr, $chr);
}
}
closedir(REF);
}
if (! -e "$wd/$target/sort_uniq/$target.sort_uniq.TTT.gz"){
&mkSortUniq($target);
}
if ($control ne $ref && ! -e "$wd/$control/sort_uniq/$control.sort_uniq.TTT.gz"){
&mkSortUniq($control);
}
if ($method eq "kmer"){
&kmer;
}elsif($method eq "cnv"){
&cnv;
}else{
&bidirectional;
}
&finish;
sub finish{
if ($tmpdir eq "$wd/$target/tmp"){
system("rm -r $tmpdir");
}elsif($tmpdir ne ""){
system("rm $tmpdir/*");
}
system("rm -r $wd/child") if -e "$wd/child";
system("rm -r $wd/$control/tmp") if -e "$wd/$control/tmp";
$end_time = time;
$elapsed_time = $end_time - $start_time;
$hour = int($elapsed_time / 3600);
$min = $elapsed_time % 3600;
$sec = $min % 60;
$min = int($min / 60);
if ($hour >= 24){
$day = int($hour / 24);
$hour = $hour % 24;
}
if ($day > 1){
$etime .= "$day days ";
}elsif($day == 1){
$etime .= "$day day ";
}
if ($hour > 1){
$etime .= "$hour hours ";
}elsif($hour == 1){
$etime .= "$hour hour ";
}
if ($min > 1){
$etime .= "$min minutes ";
}elsif($min == 1){
$etime .= "$min minute ";
}
if ($sec > 1){
$etime .= "$sec seconds ";
}elsif($sec == 1){
$etime .= "$sec second ";
}
$end_timestamp = `date`;
chomp($start_timestamp);
chomp($end_timestamp);
report("Job completed: $method method.$additionalReport
$etime ($elapsed_time seconds) elapsed.");
print LOG "start : $start_timestamp
end : $end_timestamp
elapsed : $etime
seconds : $elapsed_time\n";
close(LOG);
close(REPORT);
exit;
}
sub cnv{
&countKmer($target) if ! -e "$wd/$target/count/$target.count.TTT.gz";
&countKmer($control) if ! -e "$wd/$control/count/$control.count.TTT.gz";
&cnvJoin;
}
sub kmer{
&countKmer($target) if ! -e "$wd/$target/count/$target.count.TTT.gz";
&countKmer($control) if ! -e "$wd/$control/count/$control.count.TTT.gz";
&lbcKmer($target) if ! -e "$wd/$target/lbc/$target.lbc.TTT.gz";
&lbcKmer($control) if ! -e "$wd/$control/lbc/$control.lbc.TTT.gz";
&joinKmer;
if($ref eq ""){
system("cat $tmpdir/$target.snp.* > $wd/$target/$target.kmer.snp");
system("rm $tmpdir/$target.snp.*");
return;
}
&mapKmer;
&snpMkT;
&sortSeq;
&joinTarget;
&snpMkC;
&sortSeq;
&joinControl;
&kmerReadCount;
&toVcf;
&primer;
}
sub bidirectional{
if (! -e "$wd/$target/$target.index"){
system("rm $wd/$target/tmp/* > /dev/null 2>&1");
&mkData4MapF;
&sortData4Map;
↦
&mkData4MapR;
&sortData4Map;
↦
&align;
}
&canFork;
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=index,wd=$wd &");
&canFork;
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=svSort,wd=$wd &");
&waitChild;
&svMkT;
&sortSeq;
&joinTarget;
&svMkC;
&sortSeq;
&joinControl;
&svReadCount;
&snpMkT;
&sortSeq;
&joinTarget;
&snpMkC;
&sortSeq;
&joinControl;
&snpReadCount;
&primer(sv);
&primer;
&canFork;
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=bi2vcf,wd=$wd &");
&canFork;
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=biCount2vcf,wd=$wd &");
&waitChild;
}
sub primer{
my $type = shift;
my $count = "0001";
my $line = 0;
my $num;
if ($type eq "sv"){
open(IN, "$wd/$target/$target.sv");
open(OUT, "> $wd/$target/tmp/$target.$count");
}else{
if ($method eq "kmer"){
open(IN, "$wd/$target/$target.kmer.snp");
open(OUT, "> $wd/$target/tmp/$target.$count");
$type = "kmer";
}else{
open(IN, "$wd/$target/$target.bi.snp");
open(OUT, "> $wd/$target/tmp/$target.$count");
$type = "bi";
}
}
while(<IN>){
next if ! /H|M/;
$line++;
print OUT;
if ($line == 10000){
$line = 0;
close(OUT);
$count++;
$num = "000$count";
$num = substr($num, length($num) - 4, 4);
open(OUT, "> $wd/$target/tmp/$target.$num");
}
}
close(OUT);
for ($i = 1; $i <= $count; $i++){
&canFork;
$num = "000$i";
$num = substr($num, length($num) - 4, 4);
&report("Output primer sequences. $num");
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=primerFunc,arg=$num:$type,wd=$wd &");
}
&waitChild;
system("cat $wd/$target/tmp/primer.* > $wd/$target/$target.$type.primer && rm $wd/$target/tmp/*");
}
sub primerFunc{
my(@row, $seq, $i, $f, $r, @f, @r, $gc, $flag, $hpos, $tpos, $fpos, $rpos, $length, $tg, $head, $tail, $minimum_length, $out, $fin, $fout, $fchr, $fchrt);
my ($num, $type) = split(':', $arg);
$fin = "in.$num";
$fout = "out.$num";
$fchr = "chr.$num";
$fchrt = "chrt.$num";
open($fin, "$wd/$target/tmp/$target.$num");
open($fout, "> $wd/$target/tmp/primer.$num");
while(<$fin>){
@row = split('\t', $_);
$dat = $_;
chomp($dat);
if ($prev_chr ne $row[0]){
open($fchr, "$wd/$ref/chr$row[0]");
binmode($fchr);
}
$prev_chr = $row[0];
@f = ();
@r = ();
if ($type eq "sv"){
if($row[5] =~ /ins|del/){
if ($row[1] <= $row[3]){
$hpos = $row[1];
$tpos = $row[3]
}else{
$hpos = $row[3];
$tpos = $row[1]
}
}else{
$hpos = $row[1];
$tpos = $row[3]
}
seek($fchr, $hpos - 250, 0);
read($fchr, $head, 250);
if ($row[0] ne $row[2]){
open($fchrt, "$wd/$ref/chr$row[2]");
binmode($fchrt);
seek($fchrt, $row[3], 0);
read($fchrt, $tail, 250);
close($fchrt);
}elsif($row[5] =~ /inversion/){
seek($fchr, $tpos, 0);
read($fchr, $tail, 250);
$tail = &complement($tail);
}else{
seek($fchr, $tpos, 0);
read($fchr, $tail, 250);
}
for($i = 0; $i < 230; $i++){
$f = substr($head, $i, 20);
($gc = $f) =~ y/AT//d;
if (length($gc) == 11){
push(@f, $f);
}
}
for($i = 0; $i < 230; $i++){
$r = substr($tail, $i, 20);
($gc = $r) =~ y/AT//d;
if (length($gc) == 11){
push(@r, &complement($r));
}
}
}else{
seek($fchr, $row[1] - 250, 0);
read($fchr, $seq, 500);
for($i = 0; $i < 230; $i++){
$f = substr($seq, $i, 20);
($gc = $f) =~ y/AT//d;
if (length($gc) == 11){
push(@f, $f);
}
}
for($i = 250; $i < 480; $i++){
$r = substr($seq, $i, 20);
($gc = $r) =~ y/AT//d;
if (length($gc) == 11){
push(@r, &complement($r));
}
}
}
$minimum_length = 10000;
$out = "";
$minimum_count = 0;
foreach $f (reverse @f){
foreach $r (@r){
if (&checkDimer($f, $r)){
if ($type eq "sv"){
$fpos = index($head, $f, 0);
$rpos = index($tail, complement($r), 0);
$length = 250 - $fpos + $rpos + length($row[12]);
$tg = substr($head, 229, 20) . " " . substr($tail, 0, 20);
next if $length < 70;
if ($minimum_length > $length){
$minimum_length = $length;
$out = "$dat\t$f\t$r\t$length\t$tg\n";
$minimum_count ++;
last if $minimum_count >= 3;
}
}else{
$fpos = index($seq, $f, 0);
$rpos = index($seq, complement($r), 0);
$length = $rpos - $fpos + 20;
next if $length < 70;
@row = split('\t', $dat);
$tg = substr($seq, 99, 150) . "[$row[3]/$row[2]]" . substr($seq, 250, 149);
}
if ($minimum_length > $length){
$minimum_length = $length;
$out = "$dat\t$f\t$r\t$length\t$tg\n";
$minimum_count ++;
last if $minimum_count >= 3;
}
}
}
last if $minimum_count >= 3;
}
if ($out ne ""){
print $fout $out;
}else{
print $fout "$dat\tN\tN\tN\tN\n";
}
}
close($fin);
close($fchr);
close($fout);
}
sub checkDimer{
my ($f, $r) = @_;
my @f = split('', $f);
my @r = split('', $r);
for ($i = 3; $i < 20; $i++){
my $match = 0;
for ($j = 0; $j < $i; $j++){
my $fn = $f[20 - $i + $j];
my $rn = &complement($r[19 - $j]);
if ($fn eq $rn){
$match ++;
}
}
if ($match >= $i - 1){
return 0;
}
}
return 1;
}
sub mapKmer{
my ($tag, $nuca, $nucb, $nucc);
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Mapping SNPs between $target and $control. $tag");
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=mapKmerSub,arg=$tag,wd=$wd &");
}
}
}
&waitChild;
}
sub mapKmerSub{
my $tag = shift;
my $fin = "map.in.$tag";
my $fout = "map.out.$tag";
my $fref = "map.ref.$tag";
my (@row, %dat, $seq, $nuc, $pos, @dat);
open($fin, "$tmpdir/$target.snp.$tag");
while(<$fin>){
chomp;
@row = split;
$seq = shift(@row);
if (length($row[0]) == 1){
$dat{$seq} = "$row[0]\t$row[1]\t$row[2]\t$row[3]\t$row[4]\t$row[5]\t$row[6]\t$row[7]\t$row[8]\t$row[9]";
}
}
close($fin);
open($fout, "> $tmpdir/$target.map.$tag");
open($fref, "zcat $wd/$ref/ref20_uniq.$tag.gz|");
while(<$fref>){
chomp;
@row = split;
$seq = substr($row[0], 0, 19);
$nuc = substr($row[0], 19, 1);
if ($dat{$seq}){
@dat = split('\t', $dat{$seq});
if($row[3] eq "f"){
$pos = $row[2] + 19;
if ($nuc eq $dat[0]){
print $fout "$row[1]\t$pos\t$seq\t$nuc\t$dat[0]\t$dat[1]\t$row[3]\t$dat[2]\t$dat[3]\t$dat[4]\t$dat[5]\t$dat[6]\t$dat[7]\t$dat[8]\t$dat[9]\n";
}
}else{
$pos = $row[2] - 19;
$dat[0] = complement($dat[0]);
$dat[1] = complement($dat[1]);
$seq = complement($seq);
$nuc = complement($nuc);
if ($nuc eq $dat[0]){
print $fout "$row[1]\t$pos\t$seq\t$nuc\t$dat[0]\t$dat[1]\t$row[3]\t$dat[5]\t$dat[4]\t$dat[3]\t$dat[2]\t$dat[9]\t$dat[8]\t$dat[7]\t$dat[6]\n";
}
}
}
}
close($fout);
close($fref);
}
sub joinKmer{
my ($tag, $nuca, $nucb, $nucc);
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Detection SNPs between $target and $control. $tag");
system("perl $cwd/ped.pl target=$target,control=$control,ref=$ref,sub=joinKmerSub,arg=$tag,wd=$wd &");
}
}
}
&waitChild;
}
sub joinKmerSub{
my $tag = shift;
$cutoff = 10 if $cutoff eq "";
my $fin = "join.in.$tag";
my $fout = "join.out.$tag";
system("zcat $wd/$target/lbc/$target.lbc.$tag.gz > $tmpdir/$target.lbc.$tag");
system("zcat $wd/$control/lbc/$control.lbc.$tag.gz > $tmpdir/$control.lbc.$tag");
open($fout, "> $tmpdir/$target.snp.$tag");
open($fin, "join $tmpdir/$control.lbc.$tag $tmpdir/$target.lbc.$tag |");
while(<$fin>){
my $rep = 0;
my $a = "";
my $b = "";
my $nohita = 0;
my $nohitb = 0;
my $pola = 0;
my $polb = 0;
my @row = split;
my $i;
for ($i = 1; $i <= 4; $i++){
if($row[$i] > 100){
$rep++;
}elsif ($row[$i] >= $cutoff){
$a .= $nuc[$i -1];
if ($row[$i + 4] <= 1){
$pola++;
}
}elsif($row[$i] <= 1){
$nohita++;
}
if($row[$i + 4] > 100){
$rep++;
}elsif ($row[$i + 4] >= $cutoff){
$b .= $nuc[$i -1];
if ($row[$i] <= 1){
$polb++;
}
}elsif($row[$i + 4] <= 1){
$nohitb++;
}
}
if ($a ne $b and $a ne "" and $b ne "" and $rep == 0 and $nohita >= 2 and $nohitb >= 2 and ($pola == 1 or $polb == 1)){
print $fout "$row[0]\t$a\t$b\t$row[1]\t$row[2]\t$row[3]\t$row[4]\t$row[5]\t$row[6]\t$row[7]\t$row[8]\n";
}
}
close($fin);
close($fout);
system("rm $tmpdir/$target.lbc.$tag $tmpdir/$control.lbc.$tag");
}
sub countKmer{
my $target = shift;
my ($tag, $nuca, $nucb, $nucc);
system("mkdir $tmpdir") if ! -e "$tmpdir";
$countdir = "$wd/$target/count";
system("mkdir $countdir") if ! -e $countdir;
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Making kmer for $target. $tag");
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=countKmerSub,arg=$target:$tag,wd=$wd &");
}
}
}
&waitChild;
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Joining kmer for $target. $tag");
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=countKmerMerge,arg=$target:$tag,wd=$wd &");
}
}
}
&waitChild;
}
sub lbcKmer{
my $target = shift;
system("mkdir $wd/$target/lbc") if ! -e "$wd/$target/lbc";
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Joining kmer for $target. $tag");
system("perl $cwd/ped.pl target=$target,ref=$ref,sub=lbcKmerSub,arg=$target:$tag,wd=$wd &");
}
}
}
}
sub lbcKmerSub{
my ($target, $parent) = split(':', $arg);
$fin = "$parent.lbc.in";
$fout = "$parent.lbc.out";
open($fout, "> $wd/$target/lbc/$target.lbc.$parent");
open($fin, "zcat $wd/$target/count/$target.count.$parent.gz |");
while(<$fin>){
chomp;
($seq, $count) = split;
$head = substr($seq, 0, 19);
$nuc = substr($seq, 19, 1);
if ($head ne $prev and $prev ne ""){
print $fout "$prev\t$count{A}\t$count{C}\t$count{G}\t$count{T}\n";
$count{A} = 0;
$count{C} = 0;
$count{G} = 0;
$count{T} = 0;
}
$count{$nuc} = $count;
$prev = $head;
}
print $fout "$prev\t$count{A}\t$count{C}\t$count{G}\t$count{T}\n";
close($fin);
close($fout);
system("gzip $wd/$target/lbc/$target.lbc.$parent");
}
sub countKmerMerge{
my ($target, $parent) = split(':', $arg);
my ($nuca, $nucb, $nucc, $tag, $fin, $fout, @row, $count, $head, $prev, $nuc, %count);
system("touch $tmpdir/$parent.count");
my $countdir = "$wd/$target/count";
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
# &report("Making kmer for $target. $parent Merging of $tag subfile");
$fin = "$parent.$tag.join.in";
$fout = "$parent.$tag.join.out";
open($fout, "> $tmpdir/$parent.count.tmp");
open($fin, "$zcat $tmpdir/$parent.$tag.count.gz |join -a 1 -a 2 $tmpdir/$parent.count - |");
while(<$fin>){
chomp;
@row = split;
$count = $row[1] + $row[2];
print $fout "$row[0]\t$count\n";
}
close($fin);
close($fout);
system("mv $tmpdir/$parent.count.tmp $tmpdir/$parent.count && rm $tmpdir/$parent.$tag.count.gz");
}
}
}
system("mv $tmpdir/$parent.count $countdir/$target.count.$parent && gzip $countdir/$target.count.$parent");
}
sub countKmerSub{
my ($target, $parent) = split(':', $arg);
my ($nuca, $nucb, $nucc, $fin, $fout, $length, $seq, $ktag, $i);
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
$fout = "$tag.$parent.out";
open($fout, "> $tmpdir/$tag.$parent");
}
}
}
$fin = "$parent.in";
if (-e "$wd/$target/sort_uniq/$target.sort_uniq.$parent.gz"){
open($fin, "$zcat $wd/$target/sort_uniq/$target.sort_uniq.$parent.gz 2> /dev/null |");
}else{
open($fin, "$zcat $wd/$target/sort_uniq/$target.$parent.gz 2> /dev/null |");
}
while(<$fin>){
chomp;
$length = length($_);
for ($i = 0; $i <= $length - 20; $i++){
$seq = substr($_, $i, 20);
if ($seq !~ /N/){
$ktag = substr($seq, 0, 3);
$fout = "$ktag.$parent.out";
print $fout "$seq\n";
}
}
}
close($fin);
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
$fout = "$tag.$parent.out";
close($fout);
system("sort $sort_opt -T $tmpdir $tmpdir/$tag.$parent |uniq -c | awk '{print \$2 \"\t\" \$1}' |gzip -c > $tmpdir/$tag.$parent.count.gz && rm $tmpdir/$tag.$parent");
}
}
}
}
sub cnvJoin{
foreach $nuca (@nuc){
foreach $nucb (@nuc){
foreach $nucc (@nuc){
$tag = $nuca . $nucb . $nucc;
&canFork;
&report("Join kmer between $target and $control. $tag");
system("perl $cwd/ped.pl target=$target,control=$control,ref=$ref,sub=cnvJoinSub,tag=$tag,wd=$wd &");
}
}
}
&waitChild;
system("cat $tmpdir/map.* |sort -k 1 -n -k 2 -n $sort_opt -T $tmpdir > $wd/$target/$target.$control.cnv");
}
sub cnvJoinSub{
open(OUT, "> $tmpdir/cnv.$tag.tmp");
open(IN, "bash -c \"join -a 1 -a 2 <($zcat $wd/$control/count/$control.count.$tag.gz) <($zcat $wd/$target/count/$target.count.$tag.gz) \"|");
while(<IN>){
chomp;
@row = split;
print OUT "$row[0]\n";
}
close(OUT);
open(OUT, "> $tmpdir/cnv.$tag.$target");
open(IN, "zcat $wd/$target/count/$target.count.$tag.gz |join -a 1 -a 2 $tmpdir/cnv.$tag.tmp -|");
while(<IN>){
chomp;
@row = split;
$row[1] = 0 if $row[1] eq "";
print OUT "$row[0]\t$row[1]\n";
}
close(OUT);
open(OUT, "> $tmpdir/cnv.$tag.$control");
open(IN, "zcat $wd/$control/count/$control.count.$tag.gz |join -a 1 -a 2 $tmpdir/cnv.$tag.tmp -|");
while(<IN>){
chomp;
@row = split;
$row[1] = 0 if $row[1] eq "";
print OUT "$row[0]\t$row[1]\n";
}
close(OUT);
system("join $tmpdir/cnv.$tag.$target $tmpdir/cnv.$tag.$control > $tmpdir/join.$tag && rm $tmpdir/cnv.$tag.*");
open(OUT, "> $tmpdir/map.$tag");
open(IN, "zcat $wd/$ref/ref20_uniq.$tag | join - $tmpdir/join.$tag|");
while(<IN>){
chomp;
@row = split;
print OUT "$row[1]\t$row[2]\t$row[3]\t$row[4]\t$row[5]\n";
}
close(OUT);
}
sub mkRef{
my (@row, $i, $remote_file);
if ($file eq ""){
if ($file{$ref} ne "" and -e "$wd/$ref/$file{$ref}"){
if ($chr[0] ne ""){
&mkChr;
}else{
$file = $file{$ref};
&mkChrFromFile($file);
}
}else{
@row = split('/', $curl{$ref});
$remote_file = $row[$#row];
if (! -e "$wd/$ref/$remote_file" and $curl{$ref} ne ""){
if ($curl{$ref} !~/^[h|f]/){
&report($curl{$ref});
&finish;
}
&report("Downloading $curl{$ref}");
system("cd $wd/$ref && curl -O $curl{$ref} && cd $wd");
if ($ref eq "sacCer3"){
system("tar xfz S288C_reference_genome_R64-1-1_20110203.tgz");
system("mv S288C_reference_genome_R64-1-1_20110203/S288C_reference_sequence_R64-1-1_20110203.fsa $wd/$ref");
}
}
&mkChr;
}
}else{
&mkChrFromFile($file);
}
&mkChrFasta;
&mk20;
&mkControlRead;
system("rm -r $wd/$ref/tmp");
}
sub mkChrFromFile{
&report("Making chromosome file for $ref.");