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|
#!/usr/bin/perl -w
# Authors: Maria Nattestad and Mike Schatz
# Email: mnattest@cshl.edu
use strict;
my @chromosome_filter_choices = ("all-chromosomes","primary-chromosomes");
my @longrange_filter_choices = ("include-longrange","exclude-longrange","longrange-only");
my @output_file_choices = ("bed","bedpe");
my $USAGE = "Usage:\nAssemblytics_between_alignments.pl coords.tab minimum_event_size maximum_event_size [@chromosome_filter_choices] [@longrange_filter_choices] [@output_file_choices] > fusions.svs.bedpe ";
my $coordsfile = shift @ARGV or die $USAGE;
my $minimum_event_size = int(shift @ARGV);
my $maximum_event_size = int(shift @ARGV);
my $chromosome_filter = shift @ARGV or die $USAGE;
my $longrange_filter = shift @ARGV or die $USAGE;
my $output_file = shift @ARGV or die $USAGE;
# How close do alignments have to be in order to call deletions and insertions? (as opposed to contractions and expansions)
my $narrow_threshold = 50;
# Number of basepairs of distance in either the reference or the query before we call an SV long-range
my $longrange = $maximum_event_size;
# What is the longest two alignments can map apart in the query before we throw the variant between them away?
my $max_query_dist = 100000;
my %chromosome_filter_choices_hash = map { $_, 1 } @chromosome_filter_choices;
my %longrange_filter_choices_hash = map { $_, 1 } @longrange_filter_choices;
my %output_file_choices_hash = map { $_, 1 } @output_file_choices;
if ( $chromosome_filter_choices_hash{ $chromosome_filter } && $longrange_filter_choices_hash{ $longrange_filter } && $output_file_choices_hash { $output_file }) {
# All is well with the world
} else {
die $USAGE;
}
if ($longrange_filter ne "exclude-longrange" && $output_file eq "bed"){
die "Cannot output bed while allowing long-range variants\n$USAGE";
}
# open COORDS, "./bin/show-coords -rclHT $deltafile |"
# or die "Can't process $deltafile ($!)\n";
open COORDS, "$coordsfile" or die "Can't process $coordsfile ($!)\n";
##open COORDS, "show-coords -rclHT $deltafile |"
## or die "Can't process $deltafile ($!)\n";
## Require the flanking alignments are at least this long to call an SV
## Note there is no minimum length for fusions, this is determined by how
## the delta file was filtered
my $MIN_SV_ALIGN = 100;
#my $minimum_event_size = 50;
my $approximately_zero = $narrow_threshold;
my %alignments;
my $numalignments = 0;
while (<COORDS>)
{
chomp;
my @vals = split /\s+/, $_;
my $rid = $vals[6];
my $qid = $vals[7];
my $a;
$a->{"rstart"} = $vals[0];
$a->{"rend"} = $vals[1];
$a->{"qstart"} = $vals[2];
$a->{"qend"} = $vals[3];
$a->{"rlen"} = $vals[4];
$a->{"qlen"} = $vals[5];
$a->{"rid"} = $vals[6];
$a->{"qid"} = $vals[7];
$a->{"str"} = $_;
$a->{"qidx"} = 0;
$a->{"qrc"} = ($a->{"qend"} > $a->{"qstart"}) ? 0 : 1;
push @{$alignments{$qid}->{$rid}}, $a; # a is a hash with all the info for one alignment
$numalignments++;
}
print STDERR "Loaded $numalignments alignments\n";
my $candidatefusions = 0;
my $candidatesvs = 0;
my $sv_id_counter = 0;
my %svstats;
foreach my $qid (sort keys %alignments) # query name is the key for the alignments hash
{
my @refs = sort keys %{$alignments{$qid}}; # grab all alignments of that query
my $numref = scalar @refs;
## scan for fusions
# if ($numref > 1) # if query aligns to multiple chromosomes
# {
# my $allrefs = join " ", @refs; # join the names together for output
# print "== $qid [$numref] $allrefs\n"; # output the names of the chromosomes
# $candidatefusions++;
# my $rcnt = 0;
# foreach my $rid (@refs)
# {
# print "--\n" if ($rcnt > 0);
# $rcnt++;
# foreach my $a (@{$alignments{$qid}->{$rid}})
# {
# my $str = $a->{"str"};
# print "$str\n";
# }
# }
# print "\n";
# }
## Resort the alignments by query sort position
my @qaligns;
foreach my $rid (@refs)
{
foreach my $a (@{$alignments{$qid}->{$rid}})
{
push @qaligns, $a;
}
}
## Now record the index of the sorted query indices
@qaligns = sort { $a->{"qstart"} <=> $b->{"qstart"}} @qaligns;
for (my $i=0; $i < scalar @qaligns; $i++)
{
$qaligns[$i]->{"qidx"} = $i;
}
## scan for SVs
my $numalign = scalar @qaligns;
if ($numalign > 1) # if the query has more than 1 alignment
{
## note skip first one
for (my $j = 1; $j < $numalign; $j++)
{
my $ai = $qaligns[$j-1];
my $aj = $qaligns[$j];
my $istr = $ai->{"str"};
my $jstr = $aj->{"str"};
# if ($ai->{"rid"} ne $aj->{"rid"})
# {
# ## skip the fusions for now #############################################################################################
# next;
# }
my $rid = $ai->{"rid"};
if (($ai->{"rlen"} >= $MIN_SV_ALIGN) &&
($aj->{"rlen"} >= $MIN_SV_ALIGN))
{
## r alignments are always forward, q alignments may be flipped
my $rpos;
my $qpos;
my $rdist = 0;
my $qdist = 0;
my $svtype = 0;
my $chromi = $ai->{"rid"};
my $chromj = $aj->{"rid"};
my $posi;
my $posj;
my $strandi;
my $strandj;
$sv_id_counter++;
if (($ai->{"qrc"} == 0) && ($aj->{"qrc"} == 0))
{
## ri: [1 - 1000] | j: [2000 - 3000] => 1000
## qi: [1 - 1000] | j: [2000 - 3000] => 1000
$svtype = "FF";
$qdist = $aj->{"qstart"} - $ai->{"qend"};
$rdist = $aj->{"rstart"} - $ai->{"rend"};
if ($rdist >= 0) { $rpos = sprintf("%s:%d-%d:+", $rid, $ai->{"rend"}, $aj->{"rstart"}); }
else { $rpos = sprintf("%s:%d-%d:-", $rid, $aj->{"rstart"}, $ai->{"rend"}); }
if ($qdist >= 0) { $qpos = sprintf("%s:%d-%d:+", $qid, $ai->{"qend"}, $aj->{"qstart"}); }
else { $qpos = sprintf("%s:%d-%d:-", $qid, $aj->{"qstart"}, $ai->{"qend"}); }
# When the alignments are forward-forward, the connection point is at the end of the first (i: rend) and at the beginning of the second (j: rstart)
# i + - j
# ------> -------->
$posi = $ai->{"rend"};
$posj = $aj->{"rstart"};
$strandi = "+";
$strandj = "-";
}
elsif (($ai->{"qrc"} == 1) && ($aj->{"qrc"} == 1))
{
## ri: [2000 - 3000] | j: [1 - 1000] => 1000
## qi: [1000 - 1] | j: [3000 - 2000] => 1000
$svtype = "RR";
$rdist = $ai->{"rstart"} - $aj->{"rend"};
$qdist = $aj->{"qend"} - $ai->{"qstart"};
if ($rdist >= 0) { $rpos = sprintf("%s:%d-%d:+", $rid, $aj->{"rend"}, $ai->{"rstart"}); }
else { $rpos = sprintf("%s:%d-%d:-", $rid, $ai->{"rstart"}, $aj->{"rend"}); }
if ($qdist >= 0) { $qpos = sprintf("%s:%d-%d:+", $qid, $ai->{"qstart"}, $aj->{"qend"}); }
else { $qpos = sprintf("%s:%d-%d:-", $qid, $aj->{"qend"}, $ai->{"qstart"}); }
# When the alignments are reverse-reverse, the connection point is at the beginning of the first (i: rstart) and at the end of the second (j: rend)
# j + - i
# <------- <--------
$posi = $ai->{"rstart"}; # rstart means first reference coordinate, not with respect to the contig
$posj = $aj->{"rend"}; # rend means last reference coordinate, not with respect to the contig
$strandi = "-";
$strandj = "+";
}
elsif (($ai->{"qrc"} == 0) && ($aj->{"qrc"} == 1))
{
## ri: [1 - 1000] | j: [2000 - 3000] => 1000
## qi: [1 - 1000] | j: [3000 - 2000] => 1000
$svtype = "FR";
$qdist = $aj->{"qend"} - $ai->{"qend"};
$rdist = $aj->{"rstart"} - $ai->{"rend"};
if ($rdist >= 0) { $rpos = sprintf("%s:%d-%d:+", $rid, $ai->{"rend"}, $aj->{"rstart"}); }
else { $rpos = sprintf("%s:%d-%d:-", $rid, $aj->{"rstart"}, $ai->{"rend"}); }
if ($qdist >= 0) { $qpos = sprintf("%s:%d-%d:+", $qid, $ai->{"qend"}, $aj->{"qend"}); }
else { $qpos = sprintf("%s:%d-%d:-", $qid, $aj->{"qend"}, $ai->{"qend"}); }
# When the alignments are forward-reverse, the connection point is at the beginning of the first (i: rstart) and at the end of the second (j: rend)
# i + j +
# -------> <--------
$posi = $ai->{"rend"};
$posj = $aj->{"rend"};
$strandi = "+";
$strandj = "+";
}
elsif (($ai->{"qrc"} == 1) && ($aj->{"qrc"} == 0))
{
## ri: [1 - 1000] | j: [2000 - 3000] => 1000
## qi: [1000 - 1] | j: [2000 - 3000] => 1000
$svtype = "RF";
$qdist = $ai->{"qend"} - $aj->{"qend"};
$rdist = $aj->{"rstart"} - $ai->{"rend"};
if ($rdist >= 0) { $rpos = sprintf("%s:%d-%d:+", $rid, $ai->{"rend"}, $aj->{"rstart"}); }
else { $rpos = sprintf("%s:%d-%d:-", $rid, $aj->{"rstart"}, $ai->{"rend"}); }
if ($qdist >= 0) { $qpos = sprintf("%s:%d-%d:+", $qid, $aj->{"qend"}, $ai->{"qend"}); }
else { $qpos = sprintf("%s:%d-%d:-", $qid, $ai->{"qend"}, $aj->{"qend"}); }
# When the alignments are reverse-forward:
# - i - j
# <------- -------->
$posi = $ai->{"rstart"};
$posj = $aj->{"rstart"};
$strandi = "-";
$strandj = "-";
}
else
{
my $irc = $ai->{"qrc"};
my $jrc = $aj->{"qrc"};
print "ERROR: Unknown SV: $irc $jrc\n";
print "$istr\n";
print "$jstr\n";
die "ERROR: Unknown SV: $irc $jrc\n";
}
my $totaldist = $rdist + $qdist;
my $typeguess = "";
my $abs_event_size = abs($rdist-$qdist);
if ($chromi ne $chromj) { # interchromosomal
$typeguess = "Interchromosomal";
$rdist = 0;
} else { # same chromosome
if ($strandi eq $strandj) {
$typeguess = "Inversion";
$abs_event_size = $rdist;
}
elsif ($qdist > $rdist) {
# both are significantly negative: (means the size of an overlapping region got larger, so tandem element expansion)
if ($rdist > -1*$approximately_zero && $rdist < $approximately_zero && $qdist > -1*$approximately_zero) {
$typeguess = "Insertion";
# split into out of nowhere (rdist ~ 0) vs. rdist is > 0: insertion_in_unmapped_region
}
else {
if ($rdist < 0 || $qdist < 0) {
$typeguess = "Tandem_expansion";
} else {
$typeguess = "Repeat_expansion";
}
}
}
elsif ($qdist < $rdist) {
# both are significantly negative: (means the size of an overlapping region got smaller, so tandem element contraction)
if ($rdist > -1*$approximately_zero && $qdist > -1*$approximately_zero && $qdist < $approximately_zero) {
$typeguess = "Deletion";
# split into out of nowhere (rdist ~ 0) vs. rdist is > 0: deletion_in_unmapped_region
}
else {
if ($rdist < 0 || $qdist < 0) {
$typeguess = "Tandem_contraction";
} else {
$typeguess = "Repeat_contraction";
}
}
}
else {
$typeguess = "None";
}
if ($abs_event_size > $longrange) { # || abs($rdist) > $longrange || abs($qdist) > $longrange
$typeguess = "Longrange";
if (abs($qdist) > $max_query_dist) {
$typeguess = "None";
}
}
# my $ratio;
# if ($qdist != 0){
# # $ratio = abs(($rdist/$qdist)-1);
# # if ($ratio < 0.1) {
# # $typeguess = "Equilibrium";
# # }
# if ($rdist==$qdist || abs($qdist) > $longrange) {
# $typeguess = "None";
# }
# }
}
# my @chromosome_filter_choices = ("all-chromosomes","primary-chromosomes");
# my @longrange_filter_choices = ("include-longrange","exclude-longrange");
my $chromi_length = length $chromi; # length of the chromosome names: a way to filter to primary chromosomes and cut out alts and patches from the assembly
my $chromj_length = length $chromj;
if ($typeguess ne "Inversion" && $typeguess ne "None" && $abs_event_size >= $minimum_event_size) { # always required
if ($chromosome_filter eq "all-chromosomes" || ($chromi_length < 6 && $chromj_length < 6)) { # test for primary chromosomes unless "all-chromosomes" is chosen
if ($longrange_filter ne "exclude-longrange" || ($typeguess ne "Interchromosomal" && $typeguess ne "Longrange")) {
if ($longrange_filter ne "longrange-only" || ($typeguess eq "Interchromosomal" || $typeguess eq "Longrange")) {
if ($output_file eq "bedpe") {
print "$chromi\t$posi\t@{[$posi + 1]}\t$chromj\t$posj\t@{[$posj + 1]}\tAssemblytics_b_$sv_id_counter\t$abs_event_size\t$strandi\t$strandj\t$typeguess\t$rdist\t$qdist\t$qpos\t$abs_event_size\t$svtype\tbetween_alignments\n";
}
else {
use List::Util qw(min max);
my $ref_start = min(($posi, $posj));
my $ref_stop = max(($posi, $posj));
if ($ref_stop eq $ref_start) {
$ref_stop = $ref_start + 1;
}
# "chrom","start","stop","name","event.size","strand","event.type","ref.dist","query.dist","contig.name"
print "$chromi\t$ref_start\t$ref_stop\tAssemblytics_b_$sv_id_counter\t$abs_event_size\t+\t$typeguess\t$rdist\t$qdist\t$qpos\tbetween_alignments\n";
}
}
}
}
#if ($filter_type ~~ ("primary-allsizes","primary-shortrange") {
# && $typeguess ne "Interchromosomal" && $typeguess ne "Inversion" && $chromi_length < 6 && $chromj_length < 6 && $abs_event_size >= $minimum_event_size) {
}
$candidatesvs++;
#push @{$svstats{$svtype}}, $totaldist;
}
}
}
}
# print "Processed $numalignments alignments found $candidatefusions fusions and $candidatesvs SVs\n";
# print STDERR "Processed $numalignments alignments found $candidatefusions fusions and $candidatesvs SVs\n";
# foreach my $svtype (keys %svstats)
# {
# my @events = @{$svstats{$svtype}};
# my $cnt = scalar @events;
# my $sum = 0.0;
# foreach my $e (@events)
# {
# $sum += $e;
# }
# my $mean = sprintf ("%0.02f", $sum/$cnt);
# print "svtype[$svtype]: $cnt $mean\n";
# print STDERR "svtype[$svtype]: $cnt $mean\n";
# }
|
