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Copy pathACF_Step2_MuSeg.pl
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ACF_Step2_MuSeg.pl
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#!/usr/bin/perl -w
use strict;
# rescue circ spanning very short exons
die "Usage: $0 \"CB_splice_folder\" \"multiple_segments\" \"split_exon_gtf\" \"genome_folder\" \"output\" \"\(optional\) extend N bases\" \"\(DEBUG\)\"" if (@ARGV < 5);
my $DIR=$ARGV[0]; # /home/arthur/CB_splice/
my $filein1=$ARGV[1];
my $filein2=$ARGV[2];
my $genome=$ARGV[3];
my $fileout=$ARGV[4];
my $Ext=4;
if (scalar(@ARGV) > 5) {$Ext=$ARGV[5];}
die "Usage: $0 \"multiple_segments\" \"split_exon_gtf\" \"output\" \"\(optional\) extend positive N bases\" \"\(DEBUG set to 1\)\"" if ($Ext < 0);
print "Extension = $Ext\n";
my $debug=-1;
if (scalar(@ARGV) > 6) {$debug=$ARGV[6];}
my $cutoff=0.9;
my $Extend=15;
my $closegap=50;
my $command="rm -f Step2_MuSeg_finished";
system($command);
my %uniq;
open IN1,$filein2;
while(<IN1>) {
chomp;
my @a=split("\t",$_);
my $id=$a[7]."___".$a[8];
if ($a[6] eq "+") {
# 5' of exon
my $base=int($a[3]/1000);
if (exists $uniq{$a[0]}{$base}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base}{$a[3]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base}{$a[3]}=join("___",10,$id); }
if (exists $uniq{$a[0]}{$base+1}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base+1}{$a[3]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base+1}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base+1}{$a[3]}=join("___",10,$id); }
if (exists $uniq{$a[0]}{$base-1}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base-1}{$a[3]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base-1}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base-1}{$a[3]}=join("___",10,$id); }
# 3' of exon
$base=int($a[4]/1000);
if (exists $uniq{$a[0]}{$base}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base}{$a[4]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base}{$a[4]}=join("___",1,$id); }
if (exists $uniq{$a[0]}{$base+1}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base+1}{$a[4]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base+1}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base+1}{$a[4]}=join("___",1,$id); }
if (exists $uniq{$a[0]}{$base-1}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base-1}{$a[4]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base-1}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base-1}{$a[4]}=join("___",1,$id); }
}
else {
# 5' of exon
my $base=int($a[4] / 1000);
if (exists $uniq{$a[0]}{$base}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base}{$a[4]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base}{$a[4]}=join("___",10,$id); }
if (exists $uniq{$a[0]}{$base+1}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base+1}{$a[4]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base+1}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base+1}{$a[4]}=join("___",10,$id); }
if (exists $uniq{$a[0]}{$base-1}{$a[4]}) {
my @b=split("___",$uniq{$a[0]}{$base-1}{$a[4]});
if ($b[0] < 10) { $b[0]+=10; $uniq{$a[0]}{$base-1}{$a[4]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base-1}{$a[4]}=join("___",10,$id); }
# 3' of exon
$base=int($a[3] / 1000);
if (exists $uniq{$a[0]}{$base}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base}{$a[3]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base}{$a[3]}=join("___",1,$id); }
if (exists $uniq{$a[0]}{$base+1}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base+1}{$a[3]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base+1}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base+1}{$a[3]}=join("___",1,$id); }
if (exists $uniq{$a[0]}{$base-1}{$a[3]}) {
my @b=split("___",$uniq{$a[0]}{$base-1}{$a[3]});
if (($b[0] < 1) or ($b[0] eq 10)) { $b[0]+=1; $uniq{$a[0]}{$base-1}{$a[3]}=join("___",@b)}
}
else { $uniq{$a[0]}{$base-1}{$a[3]}=join("___",1,$id); }
}
}
close IN1;
open IN2,$filein1;
open OUT1,">".$fileout.".S1.pp";
open OUT11,">".$fileout.".S1.ppparsed";
open OUT2,">".$fileout.".S1.pm";
open OUT22,">".$fileout.".S1.pmparsed";
my %READ;
my %Overlap;
while(<IN2>) {
chomp;
my @a=split("\t",$_);
$READ{$a[0]}=join("\t",$_);
my $NR=scalar(@a)/5;
my %strand;
my $gap=-1;
for(my $i=1; $i<$NR; $i++) {
$strand{$a[5*$i + 4]}++;
if ($debug eq 1) { print join("\t",$a[0],$a[4],$a[5*$i],$a[5*$i+1],$a[5*$i+2],$a[5*$i+3],$a[5*$i+4]),"\n"; }
my $base=int(($a[5*$i + 2])/1000);
if ($debug eq 1) { print $base,"\n"; }
my $info="NA";
if (exists $uniq{$a[4]}) {
if (exists $uniq{$a[4]}{$base}) {
foreach my $pos (keys %{$uniq{$a[4]}{$base}}) {
if ($debug eq 1) {print join("\t",$a[0],$a[4],$a[5*$i],$a[5*$i+1],$a[5*$i+2],$a[5*$i+3],$a[5*$i+4],$pos,$uniq{$a[4]}{$base}{$pos}),"\n";}
if (abs($pos - $a[5*$i + 2]) <= $Ext) { $info=$uniq{$a[4]}{$base}{$pos}; last; }
}
}
}
$a[5*$i+2]=$a[5*$i + 2]."\t".$info;
$base=int(($a[5*$i + 3])/1000);
if ($debug eq 1) { print $base,"\n"; }
$info="NA";
if (exists $uniq{$a[4]}) {
if (exists $uniq{$a[4]}{$base}) {
foreach my $pos (keys %{$uniq{$a[4]}{$base}}) {
if ($debug eq 1) {print join("\t",$a[0],$a[4],$a[5*$i],$a[5*$i+1],$a[5*$i+2],$a[5*$i+3],$a[5*$i+4],$pos,$uniq{$a[4]}{$base}{$pos}),"\n";}
if (abs($pos - $a[5*$i + 3]) <= $Ext) { $info=$uniq{$a[4]}{$base}{$pos}; last; }
}
}
}
$a[5*$i+3]=$a[5*$i + 3]."\t".$info;
#if ($i > 1) {
# my $tmp_gap=$a[5*$i-5] + $a[5*$i-5+1] - $a[5*$i];
# if ($tmp_gap > $gap) { $gap=$tmp_gap;}
#}
#$Overlap{$a[0]}=$gap;
}
my $ss=0;
foreach my $i (keys %strand) { $ss++; }
if ($ss eq 1) {
print OUT1 join("\t",@a),"\n";
my @Outer_left=qw(999999999 999999999 NA);
my @Outer_right=qw(-1 -1 NA); # where the Back Splice took place
my @Inner_left=qw(-1 -1 NA);
my @Inner_right=qw(-1 -1 NA); # should be the first and the last segments, but NO (left < right) relation assumed!!!
my %POS;
for(my $i=1; $i<$NR; $i++) {
my @tmpa=split("\t",$a[5*$i+2]);
my @tmpb=split("\t",$a[5*$i+3]);
$POS{$tmpa[0]}{$i}=join("____",$tmpa[1],$tmpb[0],$tmpb[1],$a[5*$i+4]);
if ($Inner_left[0] eq -1) { $Inner_left[0]=$tmpa[0]; $Inner_left[1]=$tmpb[0]; $Inner_left[2]=$tmpa[1];}
$Inner_right[0]=$tmpa[0]; $Inner_right[1]=$tmpb[0]; $Inner_right[2]=$tmpb[1];
if ($Outer_left[0] > $tmpa[0]) {$Outer_left[0] = $tmpa[0]; $Outer_left[1]=$tmpb[0]; $Outer_left[2]=$tmpa[1]; }
if ($Outer_right[0] < $tmpb[0]) {$Outer_right[0] = $tmpa[0]; $Outer_right[1]=$tmpb[0]; $Outer_right[2]=$tmpb[1]; }
}
# check if Inner border is truely inner
my $flag=0; # 0:circle; 1:polymerase backslide or templateswitch; 10-or-100: backslide 110:linear
if ( (($Inner_right[0] <= $Inner_left[1]) and ($Inner_left[1] <= $Inner_right[1])) or (($Inner_left[0] <= $Inner_right[1]) and ($Inner_right[1] <= $Inner_left[1])) ) {$flag++;}
for(my $i=2; ($i<($NR-1))and($flag eq 0); $i++) {
my @tmpa=split("\t",$a[5*$i+2]);
my @tmpb=split("\t",$a[5*$i+3]);
if ( $Inner_left[1] < $Inner_right[0]) {
if (($Inner_left[1] <= $tmpa[0]) and ($tmpa[0] <= $Inner_right[0])) {$flag+=10;}
if (($Inner_left[1] <= $tmpb[0]) and ($tmpb[0] <= $Inner_right[0])) {$flag+=100;}
}
else {
if (($Inner_right[1] <= $tmpa[0]) and ($tmpa[0] <= $Inner_left[0])) {$flag+=10;}
if (($Inner_right[1] <= $tmpb[0]) and ($tmpb[0] <= $Inner_left[0])) {$flag+=100;}
}
}
# check for circularity
if ($flag eq 0) {
my $last=-1;
foreach my $left (sort {$a <=> $b} keys %POS) {
foreach my $rank (sort{$a <=> $b} keys %{$POS{$left}}) {
if ($last eq -1) {}
elsif (($last - $rank) > 1) {
if ($a[-1] eq "+") {}
else {$flag++;}
}
elsif (($rank - $last) > 1) {
if ($a[-1] eq "-") {}
else {$flag++;}
}
$last=$rank;
}
}
}
# report the segmental order with sticky-end info
my $info=join("\t",$a[0],$a[1],$a[2],$a[3],$a[4],$a[-1],$flag,$Outer_left[0],$Outer_right[1]);
if ($debug eq 3) { $info=join("\t",$a[0],$a[1],$a[2],$a[3],$a[4],$a[-1],$flag,$Outer_left[0],$Outer_right[1],$Inner_left[0],$Inner_left[1],$Inner_right[0],$Inner_right[1]) }
foreach my $left (sort {$a <=> $b} keys %POS) {
foreach my $rank (sort{$a <=> $b} keys %{$POS{$left}}) {
$info=$info."\t".$rank."____".$left."____".$POS{$left}{$rank};
}
}
# report
print OUT11 $info,"\n";
}
else {
print OUT2 join("\t",@a),"\n";
my $Plus_left=999999999;
my $Plus_right=-1;
my $Minus_left=999999999;
my $Minus_right=-1;
my %POS;
for(my $i=1; $i<$NR; $i++) {
my @tmpa=split("\t",$a[5*$i+2]);
my @tmpb=split("\t",$a[5*$i+3]);
$POS{$tmpa[0]}{$i}=join("____",$tmpa[1],$tmpb[0],$tmpb[1],$a[5*$i+4]);
if ($a[5*$i+4] eq "+") {
if ($Plus_left >= $tmpa[0]) {$Plus_left = $tmpa[0];}
if ($Plus_right <= $tmpb[0]) {$Plus_right = $tmpb[0];}
}
else {
if ($Minus_left >= $tmpa[0]) {$Minus_left = $tmpa[0];}
if ($Minus_right <= $tmpb[0]) {$Minus_right = $tmpb[0];}
}
}
# Not necessary to check if Inner border is truely inner
my $flag=1; # 0:circle; 1:polymerase backslide or templateswitch; 10-or-100: backslide 110:linear
# report the segmental order with sticky-end info
my $info=join("\t",$a[0],$a[1],$a[2],$a[3],$a[4],$Plus_left,$Plus_right,$Minus_left,$Minus_right);
foreach my $left (sort {$a <=> $b} keys %POS) {
foreach my $rank (sort{$a <=> $b} keys %{$POS{$left}}) {
$info=$info."\t".$rank."____".$left."____".$POS{$left}{$rank};
}
}
# report
print OUT22 $info,"\n";
}
}
close IN2;
close OUT1; # ".S1.pp"
close OUT11; # ".S1.ppparsed"
close OUT2; # ".S1.pm"
close OUT22; # ".S1.pmparsed"
my $tmpfile1=$DIR."/me2x5";
#my %me2x5 = &makescorematrix("~/CB_splice/me2x5");
my %me2x5 = &makescorematrix($tmpfile1);
my $tmpfile2=$DIR."/splicemodels/splice5sequences";
#my %seq = &makesequencematrix("/home/arthur/CB_splice/splicemodels/splice5sequences");
my %seq = &makesequencematrix($tmpfile2);
my %bgd;
$bgd{'A'} = 0.27;
$bgd{'C'} = 0.23;
$bgd{'G'} = 0.23;
$bgd{'T'} = 0.27;
sub makesequencematrix{
my $file = shift;
my %matrix;my $n=0;
open(SCOREF, $file) || die "Can't open $file!\n";
while(<SCOREF>) {
chomp;
$_=~ s/\s//;
$matrix{$_} = $n;
$n++;
}
close(SCOREF);
return %matrix;
}
sub makescorematrix{
my $file = shift;
my %matrix;my $n=0;
open(SCOREF, $file) || die "Can't open $file!\n";
while(<SCOREF>) {
chomp;
$_=~ s/\s//;
$matrix{$n} = $_;
$n++;
}
close(SCOREF);
return %matrix;
}
sub getrest5{
my $seq = shift;
my @seqa = split(//,uc($seq));
return $seqa[0].$seqa[1].$seqa[2].$seqa[5].$seqa[6].$seqa[7].$seqa[8];
}
sub scoreconsensus5{
my $seq = shift;
my @seqa = split(//,uc($seq));
my %bgd;
$bgd{'A'} = 0.27; $bgd{'C'} = 0.23; $bgd{'G'} = 0.23; $bgd{'T'} = 0.27;
my %cons1;
$cons1{'A'} = 0.004; $cons1{'C'} = 0.0032; $cons1{'G'} = 0.9896; $cons1{'T'} = 0.0032;
my %cons2;
$cons2{'A'} = 0.0034; $cons2{'C'} = 0.0039; $cons2{'G'} = 0.0042; $cons2{'T'} = 0.9884;
my $addscore = $cons1{$seqa[3]}*$cons2{$seqa[4]}/($bgd{$seqa[3]}*$bgd{$seqa[4]});
return $addscore;
}
sub log2{
my ($val) = @_;
return log($val)/log(2);
}
sub hashseq{
my $seq = shift;
$seq = uc($seq);
$seq =~ tr/ACGT/0123/;
my @seqa = split(//,$seq);
my $sum = 0;
my $len = length($seq);
my @four = (1,4,16,64,256,1024,4096,16384);
my $i=0;
while ($i<$len) {
$sum+= $seqa[$i] * $four[$len - $i -1] ;
$i++;
}
return $sum;
}
my @metables = &makemaxentscores;
sub makemaxentscores{
my $dir = $DIR."/splicemodels/";
#my $dir = "~/CB_splice/splicemodels/";
my @list = ('me2x3acc1','me2x3acc2','me2x3acc3','me2x3acc4','me2x3acc5','me2x3acc6','me2x3acc7','me2x3acc8','me2x3acc9');
my @metables;
my $num = 0 ;
foreach my $file (@list) {
my $n = 0;
open (SCOREF,"<".$dir.$file) || die "Can't open $file!\n";
while(<SCOREF>) {
chomp;
$_=~ s/\s//;
$metables[$num]{$n} = $_;
$n++;
}
close(SCOREF);
$num++;
}
return @metables;
}
sub makewmmscores{
my $dir = $DIR."/splicemodels/";
#my $dir = "/home/arthur/CB_splice/splicemodels/";
my @list = ('me1s0acc1','me1s0acc2','me1s0acc3','me1s0acc4','me1s0acc5','me1s0acc6','me1s0acc7','me1s0acc8','me1s0acc9');
my @metables;
my $num = 0 ;
foreach my $file (@list) {
my $n = 0;
open (SCOREF,"<".$dir.$file) || die "Can't open $file!\n";
while(<SCOREF>) {
chomp;
$_=~ s/\s//;
$metables[$num]{$n} = $_;
$n++;
}
close(SCOREF);
$num++;
}
return @metables;
}
sub makemmscores{
my $dir = $DIR."/splicemodels/";
#my $dir = "/home/arthur/CB_splice/splicemodels/";
my @list = ('me2s0acc1','me2s0acc2','me2s0acc3','me2s0acc4','me2s0acc5','me2s0acc6','me2s0acc7','me2s0acc8','me2s0acc9');
my @metables;
my $num = 0 ;
foreach my $file (@list) {
my $n = 0;
open (SCOREF,"<".$dir.$file) || die "Can't open $file!\n";
while(<SCOREF>) {
chomp;
$_=~ s/\s//;
$metables[$num]{$n} = $_;
$n++;
}
close(SCOREF);
$num++;
}
return @metables;
}
sub maxentscore{
my $seq = shift;
my $table_ref = shift;
my @metables = @$table_ref;
my @sc;
$sc[0] = $metables[0]{&hashseq(substr($seq,0,7))};
$sc[1] = $metables[1]{&hashseq(substr($seq,7,7))};
$sc[2] = $metables[2]{&hashseq(substr($seq,14,7))};
$sc[3] = $metables[3]{&hashseq(substr($seq,4,7))};
$sc[4] = $metables[4]{&hashseq(substr($seq,11,7))};
$sc[5] = $metables[5]{&hashseq(substr($seq,4,3))};
$sc[6] = $metables[6]{&hashseq(substr($seq,7,4))};
$sc[7] = $metables[7]{&hashseq(substr($seq,11,3))};
$sc[8] = $metables[8]{&hashseq(substr($seq,14,4))};
my $finalscore = $sc[0] * $sc[1] * $sc[2] * $sc[3] * $sc[4] / ($sc[5] * $sc[6] * $sc[7] * $sc[8]);
return $finalscore;
}
sub getrest3{
my $seq = shift;
my $seq_noconsensus = substr($seq,0,18).substr($seq,20,3);
return $seq_noconsensus;
}
sub scoreconsensus3{
my $seq = shift;
my @seqa = split(//,uc($seq));
my %bgd;
$bgd{'A'} = 0.27; $bgd{'C'} = 0.23; $bgd{'G'} = 0.23; $bgd{'T'} = 0.27;
my %cons1;
$cons1{'A'} = 0.9903; $cons1{'C'} = 0.0032; $cons1{'G'} = 0.0034; $cons1{'T'} = 0.0030;
my %cons2;
$cons2{'A'} = 0.0027; $cons2{'C'} = 0.0037; $cons2{'G'} = 0.9905; $cons2{'T'} = 0.0030;
my $addscore = $cons1{$seqa[18]} * $cons2{$seqa[19]}/ ($bgd{$seqa[18]} * $bgd{$seqa[19]});
return $addscore;
}
sub sanity {
my $seq = shift;
my @seqa = split(//,uc($seq));
my $Nr=scalar(@seqa);
my $flag=0;
for(my $i=0; $i<$Nr; $i++) {
if ($seqa[$i] !~m/[ATCG]/) {$flag++;}
}
return $flag;
}
open INf, $fileout.".S1.ppparsed";
open OUTf, ">".$fileout.".S1.ppparsed.id";
open OUTfm, ">".$fileout.".S1.ppparsed.loop";
open OUTf1, ">".$fileout.".S1";
open OUTfa1, ">".$fileout.".S1.fa";
open OUTf2, ">".$fileout.".S2";
open OUTfa2, ">".$fileout.".S2.fa";
open OUTf12, ">".$fileout.".S1.bed12";
open OUTf22, ">".$fileout.".S2.bed12";
open OUTerr,">".$fileout."_withN";
my %uniqf;
my %uniqF;
while(<INf>) {
chomp;
next if (m/^#/);
my @a=split("\t",$_);
if (($a[6] eq 0) and ($a[1] eq $a[2]) and ($a[1] > $cutoff*$a[3])){
my $id=join("_",$a[4],$a[8],$a[7],($a[7]-$a[8]));
# report seperately looped circles
# newid-83501973/1__1 151 151 151 10 + 0 111901776 111901810 2____111901776____NA____111901863____NA____+ 3____111901776____NA____111901810____NA____+ 1____111901827____NA____111901863____NA____+
my $last=-1;
my $f=0;
my $Nr=scalar(@a);
for(my $i=9; $i<$Nr; $i++) {
my @c=split(/\_\_\_\_/,$a[$i]);
if ($last eq -1) { $last=$c[3]; }
else {
if ($last > $c[1]) { $f++; }
$last=$c[3];
}
}
if ($f > 0) {
print OUTfm join("\t",@a),"\n";
next;
}
if (exists $uniqf{$id}) {
$uniqf{$id}=$uniqf{$id}.",".$a[0];
}
else {
$uniqf{$id}=$a[0];
}
$uniqF{$a[0]}=join("\t",@a);
my @b=split("\t",$READ{$a[0]});
my $NRb=scalar(@b)/5;
if ($a[5] eq "-") {
my @cl=split(/\_\_\_\_/,$a[9]);
my @cr=split(/\_\_\_\_/,$a[-1]);
$Overlap{$a[0]} = $b[5*$cl[0]] + $b[5*$cl[0]+1] - $b[5*$cr[0]];
}
else {
my @cl=split(/\_\_\_\_/,$a[9]);
my @cr=split(/\_\_\_\_/,$a[-1]);
$Overlap{$a[0]} = $b[5*$cr[0]] + $b[5*$cr[0]+1] - $b[5*$cl[0]];
}
}
}
my %candy;
foreach my $id (sort keys %uniqf) {
my @a=split(",",$uniqf{$id});
print OUTf $id,"\t",scalar(@a),"\t",$uniqf{$id},"\n";
my %POS;
my %break_left;
my %break_right;
my $f=0;
my $strand=0;
my $min_gap=999;
my $gstart=999999999;
my $gend=-1;
for(@a) {
my $tmpid=$_;
my @b=split("\t",$uniqF{$tmpid});
if ($b[5] eq "+") { $strand++; }
else {$strand--;}
if ($min_gap > $Overlap{$tmpid}) { $min_gap=$Overlap{$tmpid}; }
my $Nr=scalar(@b);
my $last_rank=-1;
my $last_pos=-1;
if ($f eq 0) {
for(my $i=9; $i<$Nr; $i++) {
my @c=split(/\_\_\_\_/,$b[$i]);
if ($c[1] < $gstart) { $gstart=$c[1];}
if ($c[1] > $gend) { $gend=$c[1];}
$POS{$c[1]}=$c[3];
$f++;
if ($last_rank eq -1) { }
else {
if(abs($last_rank - $c[0]) > 1) {
$break_left{$last_pos}=1;
$break_right{$c[1]}=1;
}
else {
$break_left{$last_pos}=2;
$break_right{$c[1]}=2;
}
}
$last_rank=$c[0];
$last_pos=$c[1];
}
}
else {
for(my $i=9; $i<$Nr; $i++) {
my @c=split(/\_\_\_\_/,$b[$i]);
if ($c[1] < $gstart) { $gstart=$c[1];}
if ($c[1] > $gend) { $gend=$c[1];}
my $overlap=0;
my $this_left;
my $this_right;
foreach my $left (sort{$a <=> $b} keys %POS) {
my $right=$POS{$left};
if (($c[3] < $left) or ($right < $c[1])) {
# no-overlap
}
else {
$overlap=$left;
last;
}
}
my $known=0;
if ($debug eq 10) { print $b[$i],"\n",$overlap,"\t",$POS{$overlap},"\n"; }
if ($overlap eq 0) {
$POS{$c[1]}=$c[3];
$f++;
$this_left=$c[1];
$this_right=$c[3];
}
else {
$known=$overlap;
$this_left=$c[1] <= $overlap ? $c[1] : $overlap;
$this_right=$c[3] <= $POS{$overlap} ? $POS{$overlap} : $c[3];
if (($this_left ne $overlap) or ($this_right ne $POS{$overlap})) {
if (exists $break_left{$overlap}) {
my $tmp=$break_left{$overlap};
delete $break_left{$overlap};
$break_left{$this_left}=$tmp;
}
if (exists $break_right{$overlap}) {
my $tmp=$break_right{$overlap};
delete $break_right{$overlap};
$break_right{$this_left}=$tmp
}
delete $POS{$overlap};
$POS{$this_left}=$this_right;
}
$overlap=0;
# check if this expansion would overlap with the next exon
foreach my $left (sort{$a <=> $b} keys %POS) {
if ($left ne $this_left) {
my $right=$POS{$left};
if (($c[3] < $left) or ($right < $c[1])) {
# no-overlap
}
else {
$overlap=$left;
last;
}
}
}
if ($overlap ne 0) {
if ($debug eq 10) { print "Another merge","\n",$this_left,"\t",$this_right,"\n",$overlap,"\t",$POS{$overlap},"\n"; }
# merge exon {$this_left, $this_right} and {$overlap, $POS{$overlap}}
my $new_left=$this_left <= $overlap ? $this_left : $overlap;
my $new_right=$this_right <= $POS{$overlap} ? $POS{$overlap} : $this_right;
$break_left{$this_left}=2;
$break_right{$overlap}=2;
delete $POS{$this_left};
delete $POS{$overlap};
$POS{$new_left}=$new_right;
if (exists $break_right{$this_left}) {
my $tmp=$break_right{$this_left};
delete $break_right{$this_left};
$break_right{$new_left}=$tmp;
}
if (exists $break_left{$overlap}) {
my $tmp=$break_left{$overlap};
delete $break_left{$overlap};
$break_left{$new_left}=$tmp;
}
$this_left=$new_left;
$this_right=$new_right;
}
}
if ($last_rank eq -1) {
$last_rank=$c[0];
$last_pos=$this_left;
}
else {
if(abs($last_rank - $c[0]) > 1) {
if (($known eq 0) and ((!exists $break_left{$last_pos}) or ($break_left{$last_pos} ne 2)) and ($last_pos ne $gend)) { $break_left{$last_pos}=1; }
if (($known eq 0) and ((!exists $break_right{$this_left}) or ($break_right{$this_left} ne 2)) and ($this_left ne $gstart)) { $break_right{$this_left}=1; }
}
else {
$break_left{$last_pos}=2;
$break_right{$this_left}=2;
if ($known eq 0) {
if (((!exists $break_left{$this_left}) or ($break_left{$this_left} ne 2)) and ($this_left ne $gend)){
$break_left{$this_left}=1
}
}
}
$last_rank=$c[0];
$last_pos=$this_left;
}
if ($debug eq 10) {
foreach my $ttt (sort keys %break_left) { print join("\t","break_left",$ttt,$break_left{$ttt}),"\n"; }
foreach my $ttt (sort keys %break_right) { print join("\t","break_right",$ttt,$break_right{$ttt}),"\n"; }
foreach my $ttt (sort keys %POS) { print join("\t","POS",$ttt,$POS{$ttt}),"\n"; }
}
}
}
if ($debug eq 10) {
print $uniqF{$tmpid},"\n";
my $debug=$id."\t".$tmpid."\t".$f;
foreach my $left (sort{$a <=> $b} keys %POS) { $debug=$debug."\t".$left."\t".$POS{$left}; }
print $debug,"\n";
foreach my $ttt (sort keys %break_left) { print join("\t","break_left",$ttt,$break_left{$ttt}),"\n"; }
foreach my $ttt (sort keys %break_right) { print join("\t","break_right",$ttt,$break_right{$ttt}),"\n"; }
print join("\t",$gstart,$gend),"\n";
print "\n";
}
}
# picked the most used borders instead of simply the most-outer ones
my %Freq;
my %POSs;
foreach my $left (sort{$a <=> $b} keys %POS) {
my $right=$POS{$left};
for(@a) {
my $tmpid=$_;
my @b=split("\t",$uniqF{$tmpid});
my $Nr=scalar(@b);
for(my $i=9; $i<$Nr; $i++) {
my @c=split(/\_\_\_\_/,$b[$i]);
if (($left <= $c[1]) and ($c[3] <= $right)) {
$Freq{$left}{$c[1]}++;
$Freq{$right}{$c[3]}++;
}
}
}
my $new_left=-1;
foreach my $i (sort{$Freq{$left}{$b} <=> $Freq{$left}{$a}} keys %{$Freq{$left}}) {
if ($new_left eq -1) { $new_left=$i; }
elsif (($Freq{$left}{$i} >= $Freq{$left}{$new_left}) and ($i < $new_left)) { $new_left=$i; }
}
my $new_right=-1;
foreach my $i (sort{$Freq{$right}{$b} <=> $Freq{$right}{$a}} keys %{$Freq{$right}}) {
if ($new_right eq -1) { $new_right=$i; }
elsif (($Freq{$right}{$i} >= $Freq{$right}{$new_right}) and ($i > $new_right)) { $new_right=$i; }
}
$POSs{$new_left}=$new_right;
if (exists $break_left{$left}) {
my $tmp=$break_left{$left};
delete $break_left{$left};
$break_left{$new_left}=$tmp;
}
if (exists $break_right{$left}) {
my $tmp=$break_right{$left};
delete $break_right{$left};
$break_right{$new_left}=$tmp;
}
}
if ($debug eq 10) {
foreach my $ttt (sort keys %break_left) { print join("\t","break_left",$ttt,$break_left{$ttt}),"\n"; }
foreach my $ttt (sort keys %break_right) { print join("\t","break_right",$ttt,$break_right{$ttt}),"\n"; }
}
# close gap when gap is shorter than 100, then NOT extend the break-points by 50 on both side
my $last=-1;
foreach my $left (sort{$a <=> $b} keys %POSs) {
my $right=$POSs{$left};
if ($last eq -1) { $last=$left; }
else {
if ((exists $break_left{$last}) and (exists $break_right{$left}) and ($break_left{$last} eq 1) and ($break_right{$left} eq 1)) {
if (($left - $POSs{$last}) < 2*$closegap) {
$break_left{$last}=2;
if (exists $break_left{$left}) { $break_left{$last}=$break_left{$left}; delete $break_left{$left};}
$break_right{$left}=2;
delete $POSs{$left};
delete $POSs{$last};
$POSs{$last}=$right;
}
else {
#$break_right{$left-$closegap}=$break_right{$left};
#$break_right{$left}=2;
#$POSs{$last}+=$closegap;
#$POSs{$left-$closegap}=$POSs{$left};
#delete $POSs{$left};
}
}
$last=$left;
}
}
my @ID=split(/\_/,$id);
my $break="|";
my $blockSizes="";
my $blockStarts="";
my $blockcounts=0;
$gstart=999999999;
$gend=-1;
foreach my $left (sort{$a <=> $b} keys %POSs) {
if ((exists $break_left{$left}) and ($break_left{$left} eq 1)) { $break=$break.$POSs{$left}."|"; }
if ((exists $break_right{$left}) and ($break_right{$left} eq 1)) { $break=$break.$left."|"; }
if ($blockStarts eq "") {
$blockStarts=0;
$blockSizes=$POSs{$left} - $left;
}
else {
$blockStarts=$blockStarts.",".($left - $ID[2]);
$blockSizes=$blockSizes.",".($POSs{$left} - $left);
}
$blockcounts++;
if ($left < $gstart) { $gstart=$left;}
if ($POSs{$left} > $gend) { $gend=$POSs{$left};}
}
my $report=join("\t","chr".$ID[0],$gstart-1,$gend-1,$id."_".$break,scalar(@a));
if ($strand > 0) { $report=$report."\t-\t".($gstart-1)."\t".($gstart-1)."\t0\t".$blockcounts; }
else { $report=$report."\t+\t".($gstart-1)."\t".($gstart-1)."\t0\t".$blockcounts; }
$report=$report."\t".$blockSizes."\t".$blockStarts;
if (($gend - $gstart) < 1000000) { print OUTf12 $report,"\n"; }
$report=join("\t",$id,$id."_".$break,$ID[0]);
if ($strand > 0) { $report=$report."\t-";}
else { $report=$report."\t+"; }
$report=join("\t",$report,$gstart,$gend,$gstart,$gstart,$blockcounts);
my $ExonL="";
my $ExonR="";
foreach my $left (sort{$a <=> $b} keys %POSs) {
if ($ExonL eq "") {
$ExonL=$left;
$ExonR=$POSs{$left};
}
else {
$ExonL=$ExonL.",".$left;
$ExonR=$ExonR.",".$POSs{$left};
}
}
$report=join("\t",$report,$ExonL,$ExonR,$min_gap);
#print OUTf1 $report,"\n";
$candy{$ID[0]}{$gstart}{$gend}=join("\t",$id,$strand,$blockcounts,$ExonL,$ExonR,$min_gap,$break,scalar(@a));
}
close INf;
close OUTf;
my %reported;
my $f=0;
foreach my $chr (sort keys %candy) {
if ($f eq 1) {print "\t=======\tyes\n";}
elsif ($f eq -1) {print "\t=======\tno\n";}
print "searching chromosome : ".$chr.".fa";
$f=-1;
if (exists $reported{$chr}) {next;}
open (IN1, $genome."/".$chr.".fa") or next;
<IN1>;
my $SEQ;
$f=1;
$reported{$chr}=1;
while(<IN1>) {chomp; $SEQ=$SEQ.$_;}
close IN1;
foreach my $start (sort keys %{$candy{$chr}}) {
foreach my $end (sort keys %{$candy{$chr}{$start}}) {
my @a=split("\t",$candy{$chr}{$start}{$end});
# R+m-
if ($a[1] > 0) {
my %SScore5;
my %SScore3;
my $left_seq=substr($SEQ,($start-2*$Extend-1),(4*$Extend + 1));
if ((length($left_seq) ne (4*$Extend + 1)) and ($debug eq 20)) {print join("\t",$a[0],$chr,$start,"left_404"),"\n"; next;}
if ((length($left_seq) ne (4*$Extend + 1))) { next;}
my $right_seq=substr($SEQ,($end-2*$Extend-1),(4*$Extend + 1));
if ((length($right_seq) ne (4*$Extend + 1)) and ($debug eq 20)) {print join("\t",$a[0],$chr,$start,$end,"right_404"),"\n";next;}
if ((length($right_seq) ne (4*$Extend + 1))) { next;}
if ((sanity($left_seq) > 0) or (sanity($right_seq) > 0)) { print OUTerr join("\t",@a),"\tsanity_fail\t",$left_seq,"\t",$right_seq,"\n"; next;}
my $t=$left_seq;
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_left_seq=scalar reverse $t;
$t=$right_seq;
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_right_seq=scalar reverse $t;
if ($debug eq 20) { print "R+m-S5\n",$rc_left_seq,"\n",$rc_right_seq,"\n"; }
for(my $i=0; $i<=$a[5]+2; $i++) {
my $str=uc substr($rc_left_seq,(2*$Extend+1-3 - $i),9);
if ((length($str) ne 9) and ($debug eq 20)) {print join("\t",@a),"\n";last;}
if ((length($str) ne 9)) {last;}
my $tmp_score=sprintf("%.2f",&log2(&scoreconsensus5($str)*$me2x5{$seq{&getrest5($str)}}));
$SScore5{$i}=$tmp_score;
if ($debug eq 20) { print "R+m-S5\n",$i,"\t",$str,"\t",$tmp_score,"\n"; }
}
for (my $i=0; $i<=$a[5]+2; $i++) {
my $str=substr($rc_right_seq,(2*$Extend+1-20 - $i),23);
if ((length($str) ne 23) and ($debug eq 20)) {print join("\t",@a),"\n";last;}
if (length($str) ne 23) { last;}
my $tmp_score=sprintf("%.2f", &log2(&scoreconsensus3($str)*&maxentscore(&getrest3($str),\@metables)));
$SScore3{$i}=$tmp_score;
if ($debug eq 20) { print "R+m-S3\n",$i,"\t",$str,"\t",$tmp_score,"\n"; }
}
my $SSum=-99999;
my $rc_left_pos=0;
my $rc_right_pos=0;
foreach my $leftpos (sort keys %SScore5) {
foreach my $rightpos (sort keys %SScore3) {
if ((($leftpos + $rightpos) >=0) and (($leftpos + $rightpos) <= $a[5]) and (($SScore5{$leftpos} + $SScore3{$rightpos}) > $SSum)) {
$rc_left_pos=$leftpos;
$rc_right_pos=$rightpos;
$SSum=$SScore5{$leftpos} + $SScore3{$rightpos};
}
}
}
my $Start=$start;
my $End=$end;
my @ExonL=split(/\,/,$a[3]);
my @ExonR=split(/\,/,$a[4]);
my @ID=split(/\_/,$a[0]);
my $report=join("\t",$a[0]."_".$a[6],$a[0],"chr".$ID[0]);
if ($a[1] > 0) { $report=$report."\t-";}
else { $report=$report."\t+"; }
$report=join("\t",$report,$Start,$End,$Start,$Start,$a[2]);
$report=join("\t",$report,join(",",@ExonL),join(",",@ExonR),$a[5],0,$SScore5{0},0,$SScore3{0},($SScore5{0}+$SScore3{0}));
print OUTf1 $report,"\n";
my $fasta="";
for(my $k=0; $k<$a[2]; $k++) {
if ($k eq 0) {
my $t=substr($SEQ,$ExonL[$k]-1,($ExonR[$k] - $ExonL[$k]));
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_t=scalar reverse $t;
$fasta=$rc_t;
if ($debug eq 30) {print join("\t",$k,$ExonL[$k]-1,$ExonR[$k]-1,$rc_t),"\n";}
}
else {
my $t=substr($SEQ,$ExonL[$k]-1,($ExonR[$k] - $ExonL[$k]));
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_t=scalar reverse $t;
$fasta=$rc_t.$fasta;
if ($debug eq 30) {print join("\t",$k,$ExonL[$k]-1,$ExonR[$k]-1,$rc_t),"\n";}
}
}
print OUTfa1 ">".$a[0],"\n",$fasta,"\n";
$Start=$start+$rc_left_pos;
$End=$end-$rc_right_pos;
$report=join("\t",$a[0]."_".$a[6],$a[0],"chr".$ID[0]);
if ($a[1] > 0) { $report=$report."\t-";}
else { $report=$report."\t+"; }
$report=join("\t",$report,$Start,$End,$Start,$Start,$a[2]);
$ExonL[0]=$Start;
$ExonR[-1]=$End;
$report=join("\t",$report,join(",",@ExonL),join(",",@ExonR),$SSum,$SScore5{$rc_left_pos},$SScore3{$rc_right_pos},$a[5],$rc_left_pos,$rc_right_pos);
print OUTf2 $report,"\n";
my $blockSizes="";
my $blockStarts="";
$fasta="";
for(my $k=0; $k<$a[2]; $k++) {
if ($blockStarts eq "") {
$blockStarts=0;
$blockSizes=$ExonR[$k] - $ExonL[$k];
my $t=substr($SEQ,$ExonL[$k]-1,($ExonR[$k] - $ExonL[$k]));
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_t=scalar reverse $t;
$fasta=$rc_t;
if ($debug eq 30) {print join("\t",$k,$ExonL[$k]-1,$ExonR[$k]-1,$rc_t),"\n";}
}
else {
$blockStarts=$blockStarts.",".($ExonL[$k] - $Start);
$blockSizes=$blockSizes.",".($ExonR[$k] - $ExonL[$k]);
my $t=substr($SEQ,$ExonL[$k]-1,($ExonR[$k] - $ExonL[$k]));
$t=~tr/[atcgATCG]/[TAGCTAGC]/;
my $rc_t=scalar reverse $t;
$fasta=$rc_t.$fasta;
if ($debug eq 30) {print join("\t",$k,$ExonL[$k]-1,$ExonR[$k]-1,$rc_t),"\n";}
}
}
$report=join("\t","chr".$ID[0],$Start-1,$End-1,$a[0]."_".$a[6]."_".$SSum,$a[7],"-",$Start-1,$Start-1,0,$a[2],$blockSizes,$blockStarts);
if (($End - $Start) < 1000000) { print OUTf22 $report,"\n"; }
print OUTfa2 ">".$a[0],"\n",$fasta,"\n";
}
# R-m+
else {
my %SScore5;
my %SScore3;
my $left_seq=substr($SEQ,($start-2*$Extend-1),(4*$Extend + 1));
if ((length($left_seq) ne (4*$Extend + 1)) and ($debug eq 20)) {print join("\t",$a[0],$chr,$start,"left_404"),"\n"; next;}
if (length($left_seq) ne (4*$Extend + 1)) { next;}
my $right_seq=substr($SEQ,($end-2*$Extend-1),(4*$Extend + 1));
if (length($right_seq) ne (4*$Extend + 1)) {print join("\t",$a[0],$chr,$start,$end,"right_404"),"\n";next;}
if ((length($right_seq) ne (4*$Extend + 1)) and ($debug eq 20)){ next;}
if ((sanity($left_seq) > 0) or (sanity($right_seq) > 0)) { print OUTerr join("\t",@a),"\tsanity_fail\t",$left_seq,"\t",$right_seq,"\n"; next;}
if ($debug eq 20) { print "R-m+S5\n",$left_seq,"\n",$right_seq,"\n"; }
for(my $i=0; $i<=$a[5]+2; $i++) {
my $str=uc substr($right_seq,(2*$Extend+1-3 - $i),9);
if ((length($str) ne 9) and ($debug eq 20)) {print join("\t",@a),"\n";last;}
if (length($str) ne 9) { last;}
my $tmp_score=sprintf("%.2f",&log2(&scoreconsensus5($str)*$me2x5{$seq{&getrest5($str)}}));
$SScore5{$i}=$tmp_score;
if ($debug eq 20) { print "R-m+S5\n",$i,"\t",$str,"\t",$tmp_score,"\n"; }
}
for (my $i=0; $i<=$a[5]+2; $i++) {
my $str=substr($left_seq,(2*$Extend+1-20 + $i),23);
if ((length($str) ne 23) and ($debug eq 20)) {print join("\t",@a),"\n";last;}
if (length($str) ne 23) { last;}
my $tmp_score=sprintf("%.2f", &log2(&scoreconsensus3($str)*&maxentscore(&getrest3($str),\@metables)));
$SScore3{$i}=$tmp_score;
if ($debug eq 20) { print "R-m+S3\n",$i,"\t",$str,"\t",$tmp_score,"\n"; }
}
my $SSum=-99999;
my $rc_left_pos=0;
my $rc_right_pos=0;
foreach my $leftpos (sort keys %SScore5) {
foreach my $rightpos (sort keys %SScore3) {
if ((($leftpos + $rightpos) >=0) and (($leftpos + $rightpos) <= $a[5]) and (($SScore5{$leftpos} + $SScore3{$rightpos}) > $SSum)) {
$rc_left_pos=$leftpos;
$rc_right_pos=$rightpos;
$SSum=$SScore5{$leftpos} + $SScore3{$rightpos};
}
}
}
# refFlat format
my @ExonL=split(/\,/,$a[3]);
my @ExonR=split(/\,/,$a[4]);
#for(my $k=0; $k<$a[2]; $k++) { $ExonR[$k]++; }
my $Start=$start;
my $End=$end;
my @ID=split(/\_/,$a[0]);
my $report=join("\t",$a[0]."_".$a[6],$a[0],"chr".$ID[0]);
if ($a[1] > 0) { $report=$report."\t-";}
else { $report=$report."\t+"; }
$report=join("\t",$report,$Start,$End,$Start,$Start,$a[2]);
$ExonL[0]=$Start;
$ExonR[-1]=$End;
$report=join("\t",$report,join(",",@ExonL),join(",",@ExonR),$a[5],0,$SScore5{0},0,$SScore3{0},($SScore3{0}+$SScore5{0}));
print OUTf1 $report,"\n";
my $fasta="";
for(my $k=0; $k<$a[2]; $k++) {
if ($k eq 0) {
my $tmpseq=substr($SEQ,$ExonL[$k]-1,($ExonR[$k] - $ExonL[$k] - 0));
$fasta=$tmpseq;
if ($debug eq 30) {print join("\t",$k,$ExonL[$k],$ExonR[$k],$tmpseq),"\n";}
}