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#!/usr/bin/env perl
use strict;
use warnings;
use lib ("/usr/lib/trinityrnaseq/PerlLib");
use SAM_reader;
use SAM_entry;
my $usage = "\n\nusage: aligned_reads.sam [debug]\n\n"
. "Note: uses the bitflag settings for counting entries. Proper-pairing supercedes other settings.\n\n";
my $sam_file = $ARGV[0] or die $usage;
my $DEBUG = $ARGV[1] || 0;
my $DEBUG_OFH;
if ($DEBUG) {
open ($DEBUG_OFH, ">_debug.frag_classes") or die $!;
}
=notes
Entirely works off flag settings.
Proper pairs trump individual left/right alignments.
Improper pairs: left and right read alignments exist anywhere but not flagged as proper pairs.
=cut
main: {
my $prev_read_name = "";
my $prev_scaff_name = "";
my %counts;
my $count = 0;
my $sam_reader = new SAM_reader($sam_file);
while ($sam_reader->has_next()) {
$count++;
my $read = $sam_reader->get_next();
if ($read->is_query_unmapped()) { next; } # ignore unaligned read entries.
my $scaff_name = $read->get_scaffold_name();
my $core_read_name = $read->get_core_read_name();
if ($read->is_paired()) {
if ($read->is_proper_pair()) {
## erase any single entries.
if (exists $counts{ $core_read_name . "::L" }) {
delete $counts{ $core_read_name . "::L" };
}
if (exists $counts{ $core_read_name . "::R" } ) {
delete $counts{ $core_read_name . "::R" };
}
$counts{ $core_read_name . "::PP" } = 1;
}
else {
# not propper pair
unless (exists $counts{ $core_read_name . "::PP" }) {
if ($read->is_first_in_pair()) {
$counts{ $core_read_name . "::L" } = 1;
}
elsif ($read->is_second_in_pair()) {
$counts{ $core_read_name . "::R" } = 1;
}
}
}
}
else {
$counts{ $core_read_name . "::S" } = 1;
}
if ($count % 100000 == 0) {
my $count_print = $count;
$count_print=~ s/(\d)(?=(\d{3})+(\D|$))/$1\,/g;
print STDERR "\r[$count_print] ";
}
}
print STDERR "\n\n";
## identify improper pairs
my @reads = keys %counts;
foreach my $read (@reads) {
if ($read =~ /::L$/) {
my $core_name = $read;
$core_name =~ s/::L$//;
if (exists($counts{ $core_name . "::R" }) ) {
## count as improper pair instead
$counts{ $core_name . "::IP" } = 1;
delete $counts{$read};
delete $counts{ $core_name . "::R" };
}
}
}
## sum counts, generate summary
my $count_PP = 0;
my $count_L = 0;
my $count_R = 0;
my $count_S = 0;
my $count_IP = 0; # improper pairs
my $total = 0;
foreach my $read (keys %counts) {
my @x = split(/::/, $read);
my $class = pop @x;
my $core_read_name = join("::", @x);
if ($class eq "PP") {
$count_PP += 2;
$total += 2;
}
elsif ($class eq "IP") {
$count_IP += 2;
$total += 2;
}
elsif ($class eq "L") {
$count_L++;
$total++;
}
elsif ($class eq "R") {
$count_R++;
$total++;
}
elsif ($class eq "S") {
$count_S++;
$total++;
}
if ($DEBUG) {
print $DEBUG_OFH join("\t", $core_read_name, $class) . "\n";
}
}
print "Proper_pair:\t$count_PP\t" . sprintf("%.3f%%", $count_PP/$total*100) . "\n"
. "Improper_pair:\t$count_IP\t" . sprintf("%.3f%%", $count_IP/$total*100) . "\n"
. "Left-only:\t$count_L\t" . sprintf("%.3f%%", $count_L/$total*100) . "\n"
. "Right-only:\t$count_R\t" . sprintf("%.3f%%", $count_R/$total*100) . "\n"
. "Single_end:\t$count_S\t" . sprintf("%.3f%%", $count_S/$total*100) . "\n\n";
print "Total aligned frags: $total\n\n";
close $DEBUG_OFH if $DEBUG;
exit(0);
}
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