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simVirusInserts.pm
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simVirusInserts.pm
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package main;
use strict;
use warnings;
use IO::Zlib;
sub revcom($) {
my $str = $_[0];
$str =~ tr/acgtrymkswhbvdnxACGTRYMKSWHBVDNX/tgcayrkmswdvbhnxTGCAYRKMSWDVBHNX/;
my $rev = reverse $str;
$rev =~ tr/[](){}<>/][)(}{></;
return $rev;
}
sub openfile($) {
my ($filename)=@_;
my $infile;
if ($filename=~/.[bs]am(.gz)?$/) {
open( $infile,"-|","samtools view -F 256 $filename") or die "Error opening $filename: $!\n";
} elsif ($filename=~/.gz$/) {
open( $infile,"-|","gzip -dc $filename") or die "Error opening $filename: $!\n";
} elsif ($filename=~/.bz2$/) {
open( $infile,"-|","bzip2 -dc $filename") or die "Error opening $filename: $!\n";
} else {open( $infile,"<",$filename) or die "Error opening $filename: $!\n";}
return $infile;
}
sub getRefChr1stID($) {
my $GENOME = $_[0];
while (<$GENOME>) {
s/^>//;
/^(\S+)/ or next;
my $seqname = $1;
return $seqname;
}
}
sub getRefChr1st($) {
my $GENOME = $_[0];
while (<$GENOME>) {
s/^>//;
/^(\S+)/ or next;
my $seqname = $1;
print STDERR ">$seqname ...";
$/=">";
my $genome=<$GENOME>;
chomp $genome;
$genome=~s/\s//g;
$/="\n";
my $thelength = length $genome;
print STDERR "\b\b\b", $thelength, ".\n";
return $genome;
}
}
sub getticks($$$$$) {
my ($RefBorder,$Refstr,$RefLen,$PEinsertLen,$RefNratioMax) = @_;
my @theticks;
while (@theticks < 100) {
my $pos0 = int(rand($RefLen-(2*$RefBorder)))+$RefBorder;
my $str0 = substr $Refstr,($pos0-$PEinsertLen),2*$PEinsertLen;
my $seq = $str0;
my $N = $seq=~tr/Nn//;
next if $N > 2*$PEinsertLen*$RefNratioMax; # strlen is 2*$PEinsertLen
my $lc = $seq=~tr/atcg//;
next if $lc > 2*$PEinsertLen*$main::RefMratioMax;
push @theticks,$pos0;
}
@theticks = sort {$a<=>$b} @theticks;
#warn "$RefBorder,$RefLen";
return \@theticks;
}
sub getype($$) {
my ($R1,$R2)=@_;
my $type='I'; # !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJ
if ($R1 =~ /^[A-Z]+$/) {
if ($R2 =~ /^[A-Z]+[a-z]+$/) {
$type = '1';
} elsif ($R2 =~ /^[A-Z]+[a-z]+[A-Z]+$/) {
$type = '2';
} elsif ($R2 =~ /^[a-z]+$/) {
$type = '3';
} elsif ($R2 =~ /^[a-z]+[A-Z]+$/) {
$type = '4';
} elsif ($R2 =~ /^[A-Z]+$/) {
$type = '5';
}
} elsif ($R1 =~ /^[A-Z]+[a-z]+$/) {
if ($R2 =~ /^[A-Z]+[a-z]+$/) {
$type = '6';
} elsif ($R2 =~ /^[A-Z]+[a-z]+[A-Z]+$/) {
$type = '7';
} elsif ($R2 =~ /^[a-z]+$/) {
$type = '8';
} elsif ($R2 =~ /^[a-z]+[A-Z]+$/) {
$type = '9';
} elsif ($R2 =~ /^[A-Z]+$/) {
$type = 'A';
}
} elsif ($R1 =~ /^[A-Z]+[a-z]+[A-Z]+$/) {
if ($R2 =~ /^[A-Z]+[a-z]+[A-Z]+$/) {
$type = 'B';
} elsif ($R2 =~ /^[a-z]+[A-Z]+$/) {
$type = 'C';
} elsif ($R2 =~ /^[A-Z]+$/) {
$type = 'D';
}
} elsif ($R1 =~ /^[a-z]+$/) {
if ($R2 =~ /^[a-z]+[A-Z]+$/) {
$type = 'E';
} elsif ($R2 =~ /^[A-Z]+$/) {
$type = 'F';
}
} elsif ($R1 =~ /^[a-z]+[A-Z]+$/) {
if ($R2 =~ /^[a-z]+[A-Z]+$/) {
$type = 'H';
} elsif ($R2 =~ /^[A-Z]+$/) {
$type = 'G';
}
}
return $type;
}
sub base2qual($) {
my $len = length $_[0];
my $qual;
for my $i (0 .. $len-1) {
my $base = substr $_[0],$i,1;
if ($base=~/[A-Z]/) {
$qual .= 'H';
} elsif ($base=~/[a-z]/) {
$qual .= 'B';
} else {
die;
}
}
return $qual;
}
sub dosim($$$) {
my ($Refstr,$Virstr,$Paras)=@_;
my $PEinsertLen = $Paras->{PEinsertLen};
my $SeqReadLen = $Paras->{SeqReadLen};
my $RefBorder = $PEinsertLen + 1000;
open O,'>',$Paras->{OutPrefix}.'.Ref.fa';
#open R1,'>',$Paras->{OutPrefix}.'.1.fq';
#open R2,'>',$Paras->{OutPrefix}.'.2.fq';
tie *R1, 'IO::Zlib', $Paras->{OutPrefix}.'.1.fq.gz', "wb9";
tie *R2, 'IO::Zlib', $Paras->{OutPrefix}.'.2.fq.gz', "wb9";
#my @Refticks = @{getticks($RefBorder,$Refstr,$Paras->{RefLen},$PEinsertLen,$Paras->{RefNratioMax})};
#my @Virticks = @{getticks($Paras->{VirFrag},$Virstr,$Paras->{VirLen},int(0.9+ 0.5*$Paras->{VirFrag}),$Paras->{RefNratioMax})};
my @Refticks = @{$Paras->{pRefticks}}; # made a copy
my @Virticks = @{$Paras->{pVirticks}};
#ddx $Paras;
print STDERR "$Paras->{OutPrefix}:\tPE_Ins:$PEinsertLen, Vir_Frag:$Paras->{VirFrag}.\n";
for my $pRef (@Refticks) {
my $seqR1 = substr $Refstr,($pRef-$PEinsertLen),$PEinsertLen;
my $seqR2 = substr $Refstr,$pRef,$PEinsertLen;
my $pVir = shift @Virticks;
my $startV = $pVir-int(0.5*$Paras->{VirFrag});
my $seqV = substr $Virstr,$startV,$Paras->{VirFrag};
my $isReverse = int(rand(2));
$isReverse = $pRef % 2; # fixed upon odd/even
my $strand = '+';
if ($isReverse) {
$seqV = revcom($seqV);
$strand = '-';
}
my $RawNewSeq = join('',uc $seqR1,lc $seqV,uc $seqR2);
my @newSeqs = ($RawNewSeq,$RawNewSeq,$RawNewSeq);
# 根据 http://www.bioinformatics.babraham.ac.uk/training/Methylation_Course/BS-Seq%20theory%20and%20QC%20lecture.{pdf,pptx},bisulfite sequencing 按照回收接头种类分,有三种建库方式。常见的 Directional libraries是OT与OB,PBAT ~ 是CTOT与CTOB,Non-directional ~ 是四种都有。这里模拟的算是Non-directional的。下次有空再分出 Directional 与 PBAT。
# 现在已修改为最常见的 Directional 的,即bwa-meth所针对的。
$newSeqs[1] =~ tr /Cc/Tt/; # 100% un-methylation F
$newSeqs[2] =~ tr /Gg/Aa/; # 100% un-methylation R
#for my $mt (1 .. $#newSeqs) { # skip 0, only of 100% un-methylation.
# my $newSeq = $newSeqs[$mt];
my $tID = join('_','Ref',$pRef-$PEinsertLen+1,$pRef,$pRef+$PEinsertLen,'Vir',$strand,$startV+1,$startV+$Paras->{VirFrag},'R',$PEinsertLen,$SeqReadLen);
print O '>',$tID,"\n$RawNewSeq\n\n";
my $maxP = length($RawNewSeq) - $PEinsertLen;
#for my $p ($Paras->{LeftStart} .. $Paras->{LeftEnd}) {
for my $p (0 .. $maxP) {
#last if $p > $maxP;
my $fPE = substr $newSeqs[1],$p,$PEinsertLen;
my $rPE = revcom(substr $newSeqs[2],$p,$PEinsertLen);
my $fR1 = substr $fPE,0,$SeqReadLen;
my $fR2 = substr $fPE,$PEinsertLen-$SeqReadLen,$SeqReadLen;
my $revfR2 = revcom($fR2);
my $type = getype($fR1,$fR2);
$type = '0' if $p == 0 or $p == $maxP;
my ($Part1,$Part2);
$Part1 = join '-',getInsertParts($PEinsertLen,$SeqReadLen,$Paras->{VirFrag},'f',$p,1);
$Part2 = join '-',getInsertParts($PEinsertLen,$SeqReadLen,$Paras->{VirFrag},'f',$p,2);
print R1 "\@sf${p}_${type}_${tID}/1 ${Part1}\n$fR1\n+\n",base2qual($fR1),"\n";
print R2 "\@sf${p}_${type}_${tID}/2 ${Part2}\n$revfR2\n+\n",base2qual($revfR2),"\n";
my $rR1 = substr $rPE,0,$SeqReadLen;
my $rR2 = substr $rPE,$PEinsertLen-$SeqReadLen,$SeqReadLen;
my $revrR2 = revcom($rR2);
$type =~ tr/123456789ABCDEFGH/GDFA5HCE94B728316/; # 反向后的对应关系
$Part2 = join '-',getInsertParts($PEinsertLen,$SeqReadLen,$Paras->{VirFrag},'r',$p,2);
$Part1 = join '-',getInsertParts($PEinsertLen,$SeqReadLen,$Paras->{VirFrag},'r',$p,1);
print R1 "\@sr${p}_${type}_${tID}/1 ${Part1}\n$rR1\n+\n",base2qual($rR1),"\n";
print R2 "\@sr${p}_${type}_${tID}/2 ${Part2}\n$revrR2\n+\n",base2qual($revrR2),"\n";
}
#}
}
close O;
close R1; close R2;
#print STDERR "\b\b\bdone.\n";
}
sub cigar2rpos($$) {
my ($cigar,$lpos) = @_;
my $reflen = 0;
my @cigar = $cigar =~ /(\d+)(\w)/g;
while (@cigar) {
my ($len,$op) = splice(@cigar,0,2);
$reflen += $len if $op =~ /^[MD]$/;
}
return $lpos + $reflen -1;
}
sub cigar2SMS($) {
my ($cigar) = @_;
my @cigars = $cigar =~ /(\d+\w)/g;
my ($Left,$Middle,$Right)=(0,0,0);
if ($cigars[0] =~ /(\d+)S/) {
shift @cigars;
$Left = $1;
}
if ($cigars[-1] =~ /(\d+)S/) {
pop @cigars;
$Right = $1;
}
$Middle = join('',@cigars);
$Middle = cigar2rpos($Middle,1);
return [$Left,$Middle,$Right];
}
sub getInsertPos($$$$$) { # 返回Read在模拟拼合片段上的左右端点。
my ($r1fr,$innerPos,$InsertSize,$ReadLen,$r12)=@_;
my ($FiveT,$ThreeT)=(0,0);
if (($r12 == 1 and $r1fr eq 'f') or ($r12 == 2 and $r1fr eq 'r')) {
$FiveT = $innerPos;
$ThreeT = $innerPos + $ReadLen;
} elsif (($r12 == 2 and $r1fr eq 'f') or ($r12 == 1 and $r1fr eq 'r')) {
my $InnerDis = $InsertSize - $ReadLen*2;
$FiveT = $innerPos + $ReadLen + $InnerDis;
$ThreeT = $innerPos + $InsertSize;
} else {die 'Y';}
return ($FiveT,$ThreeT);
}
sub InsertPos2PartLVR($$$) { # 对getInsertPos返回的,模拟拼合片段上的点,返回其所属区段:[LVR],及到该区段末尾的长度(与读长无关)。
my ($Pos,$InsertSize,$VirFrag)=@_;
my ($type,$lastingLen)=('',0);
if ($Pos < $InsertSize) {
$type = 'L';
$lastingLen = $InsertSize - $Pos;
} elsif ( $Pos < ($InsertSize + $VirFrag) ) {
$type = 'V';
$lastingLen = $VirFrag - ($Pos - $InsertSize);
} else {
$type = 'R';
$lastingLen = 2*$InsertSize + $VirFrag - $Pos; # towards 3'
#$lastingLen = $Pos - ($InsertSize + $VirFrag); # towards Mid-point
}
# LVR模型没有考虑LVR各自悬空的情况,需要修正为LLVVRR。
=pod
if ($r1fr eq 'f') {
# those above
} elsif ($r1fr eq 'r') {
if ( $Pos > ($InsertSize + $VirFrag) ) {
$type = 'L';
$lastingLen = $Pos - $InsertSize - $VirFrag;
} elsif ( $Pos > $InsertSize ) {
$type = 'V';
$lastingLen = $Pos - $InsertSize;
} else {
$type = 'R';
$lastingLen = $Pos;
}
}
=cut
return ($type,$lastingLen);
}
sub Parts2List($$$$$$$$) { # 根据左右两个InsertPos2PartLVR返回值,计算理论覆盖模式。 [mmL,nnV,ddR] (mm+nn+dd = ReadLength)
my ($Pos,$type1,$lastingLen1,$type2,$lastingLen2,$ReadLen,$InsertSize,$VirFrag) = @_;
my %Type2Int = (
L => 1,
V => 2,
R => 3,
);
#ddx [$Pos,$type1,$lastingLen1,$type2,$lastingLen2,$ReadLen,$InsertSize,$VirFrag];
my %Int2Type = reverse %Type2Int;
if ($type1 eq $type2) { # 'L'的Read1记录这样到LR分界点最方便。需要`$nextLL`则手动替换。
return ( "${lastingLen1}${type1}" ); # 一律按左端点返回
} elsif ( $Type2Int{$type1} <= $Type2Int{$type2} ) {
my @ret = ( "${lastingLen1}${type1}" );
my $nextInt = 1+ $Type2Int{$type1};
my $nextPos = $Pos;
my $nextLL = $lastingLen1;
my $nextType;
my $RemainLen = $ReadLen;
while ($nextInt <= $Type2Int{$type2}) {
$nextPos += $nextLL;
$RemainLen -= $nextLL;
($nextType,$nextLL) = InsertPos2PartLVR($nextPos,$InsertSize,$VirFrag);
$nextLL=$RemainLen if $nextInt == $Type2Int{$type2};
#ddx [$nextInt,$nextType,$RemainLen,$nextLL,1,$r1fr,$Pos,$type1,$lastingLen1,$type2,$lastingLen2,$ReadLen,$InsertSize,$VirFrag];
push @ret,"${nextLL}$nextType";
++$nextInt;
}
return @ret;
} else { die 'E'; }
}
sub InsertPos2InsertParts($$$$$) {
my ($InsertSize,$ReadLen,$VirFrag,$FiveT,$ThreeT)=@_;
my ($type5,$lastingLen5) = InsertPos2PartLVR($FiveT,$InsertSize,$VirFrag);
my ($type3,$lastingLen3) = InsertPos2PartLVR($ThreeT,$InsertSize,$VirFrag);
my @Parts = Parts2List($FiveT,$type5,$lastingLen5,$type3,$lastingLen3,$ReadLen,$InsertSize,$VirFrag);
return @Parts;
}
sub getInsertParts($$$$$$) {
my ($InsertSize,$ReadLen,$VirFrag,$r1fr,$innerPos,$r12) = @_;
my ($FiveT,$ThreeT) = getInsertPos($r1fr,$innerPos,$InsertSize,$ReadLen,$r12);
my @Parts = InsertPos2InsertParts($InsertSize,$ReadLen,$VirFrag, $FiveT,$ThreeT);
return @Parts;
}
1;