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batch_tetrad_based_recombination_profiling_by_reference_genome.pl
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#!/usr/bin/perl
use warnings FATAL => 'all';
#use warnings;
use strict;
use Getopt::Long;
##############################################################
# script: batch_tetrad_based_recombination_profiling_by_reference_genome.pl
# author: Jia-Xing Yue (GitHub ID: yjx1217)
# last edited: 2021.06.24
# description: run batch recombination profiling to characterize different crossover and gene conversion events
# example: perl batch_recombination_profiler_by_reference_genome.pl -s Master_Sample_Table.txt -n 3 -l 100 -d 5000 -q 30 -b batch_id
##############################################################
my $sample_table = "Master_Sample_Table.txt"; # master sample table
my $batch_id = "Batch_TEST";
my $genotype_dir;
my $n = 1; # minimal number of markers to be considered for linkage blocks
my $l = 10; # minimal marker-bounded block size to be considered for linkage blocks
my $d = 5000; # distance range for merging COs
my $qual_diff = 20; # net quality difference cutoff used for genotyping
my $output_dir = "$batch_id"; # the output directory
GetOptions('sample_table|s:s' => \$sample_table, # master sample table
'genotype_dir|g:s' => \$genotype_dir, # input directory for genotyping txt file
'batch_id|b:s' => \$batch_id, # batch name used for genotyping
'min_marker_number|n:i' => \$n, # minimal enclosed markers to be considered for defining a linkage block, default = 1
'min_block_size|l:i' => \$l, # minimal linkage block size (bp) to be considered, default = 10 bp
'qual_diff|q:i' => \$qual_diff, # net quality difference used in tetrad genotyping, default = 20
'distance|d:s' => \$d, # event merging distance (bp), default = 5000
'output_dir|o:s' => \$output_dir); # the output directory
check_nonnegative("n", $n);
check_nonnegative("l", $l);
check_nonnegative("d", $d);
check_nonnegative("qual_diff", $qual_diff);
print "parameter setting table for this run>>\n";
print "sample_table = $sample_table\n";
print "genotype_dir = $genotype_dir\n";
print "batch_id = $batch_id\n";
print "min_marker_number = $n\n";
print "min_block_size = $l\n";
print "qual_diff = $qual_diff\n";
print "merging distance = $d\n";
my $sample_table_fh = read_file($sample_table);
my %tetrads = parse_sample_table_by_tetrad($sample_table_fh);
# filter_tetrad_with_inviable_spores(\%tetrads);
calculate_spore_availibility(\%tetrads);
foreach my $tetrad_id (sort keys %tetrads) {
system("mkdir -p $output_dir/$tetrad_id");
my $cross_pair = $tetrads{$tetrad_id}{'cross_pair'};
my $spore_availibility = $tetrads{$tetrad_id}{'spore_availibility'};
my ($parent1, $parent2) = split "-", $cross_pair;
print "tetrad: $tetrad_id, cross_pair: $cross_pair, parent1: $parent1, parent2: $parent2, spore_availibility: $spore_availibility\n";
my @ref = ("ref");
foreach my $ref (@ref) {
my @mode = ("raw", "inferred");
foreach my $mode (@mode) {
my $genotype_input = "$genotype_dir/$batch_id/$cross_pair.$tetrad_id.$ref.q${qual_diff}.genotype.lite.$mode.txt.gz";
my $genotype_input_fh = read_file($genotype_input);
my $prefix = "$cross_pair.$tetrad_id.$ref.q${qual_diff}.genotype.lite.$mode.recombination_profile";
my %tetrad_genotypes = ();
my %preliminary_linkage_blocks = ();
my %markers = ();
parse_tetrad_genotypes_file($genotype_input_fh, $parent1, $parent2, $ref, $spore_availibility, \%tetrad_genotypes, \%preliminary_linkage_blocks, \%markers);
my %linkage_blocks = adjust_linkage_blocks($n, $l, \%preliminary_linkage_blocks);
my %blocks_by_segregation_pattern = ();
my @merging_log = ();
foreach my $chr (sort keys %linkage_blocks) {
foreach my $block_id (sort {$a <=> $b} keys %{$linkage_blocks{$chr}}) {
my $marker_pos_start = $linkage_blocks{$chr}{$block_id}{'marker_pos_start'};
my $marker_pos_end = $linkage_blocks{$chr}{$block_id}{'marker_pos_end'};
my $marker_raw_index_start = $linkage_blocks{$chr}{$block_id}{'marker_raw_index_start'};
my $marker_raw_index_end = $linkage_blocks{$chr}{$block_id}{'marker_raw_index_end'};
my $marker_effective_index_start = $linkage_blocks{$chr}{$block_id}{'marker_effective_index_start'};
my $marker_effective_index_end = $linkage_blocks{$chr}{$block_id}{'marker_effective_index_end'};
my $segregation_pattern = $linkage_blocks{$chr}{$block_id}{'segregation_pattern'}; # parent1_genotype_count:parent2_genotype_count:NA_genotype_count
my $genotype_pattern = $linkage_blocks{$chr}{$block_id}{'genotype_pattern'}; # genotype_a:genotype_b:genotype_c:genotype_d
# print "$genotype_pattern\t$block_id\t$chr\tmarker position range: $marker_pos_start-$marker_pos_end, marker raw index range: $marker_raw_index_start-$marker_raw_index_end, marker effective index range: $marker_effective_index_start-$marker_effective_index_end, segregation_pattern: $segregation_pattern\n";
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_pos_start'} = $marker_pos_start;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_pos_end'} = $marker_pos_end;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_raw_index_start'} = $marker_raw_index_start;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_raw_index_end'} = $marker_raw_index_end;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_effective_index_start'} = $marker_effective_index_start;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'marker_effective_index_end'} = $marker_effective_index_end;
$blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$block_id}{'genotype_pattern'} = $genotype_pattern;
}
}
my $recombination_event_id = 0;
my %recombination_events = ();
my %CO_associated_GC = ();
my @segregation_patterns = qw(2:2:0 3:1:0 1:3:0 4:0:0 0:4:0); # parent1:parent2:NA
foreach my $chr (sort keys %linkage_blocks) {
my $block_total_count = scalar keys %{$linkage_blocks{$chr}};
if ($block_total_count < 2) {
# print "no recombination on chromosome: $chr\n";
} else {
foreach my $segregation_pattern (@segregation_patterns) {
# print "segregation_pattern=$segregation_pattern\n";
if (exists $blocks_by_segregation_pattern{$chr}{$segregation_pattern}) {
# print "examing blocks with the same segregation_pattern: $segregation_pattern\n";
my @block_subclass = (sort {$a <=> $b} keys %{$blocks_by_segregation_pattern{$chr}{$segregation_pattern}});
my $block_subclass_count = scalar @block_subclass;
if ($segregation_pattern eq "2:2:0") {
if ($block_subclass_count < 2) {
# print "no CO on chromosome: $chr\n";
} else {
# slide through blocks with the same segregation pattern, two neighbouring blocks a time
for (my $i = 0; $i < $block_subclass_count - 1; $i++) {
my $current_block_id = $block_subclass[$i];
my $next_block_id = $block_subclass[$i + 1];
my $current_block_genotype_pattern = $blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$current_block_id}{'genotype_pattern'};
my $next_block_genotype_pattern = $blocks_by_segregation_pattern{$chr}{$segregation_pattern}{$next_block_id}{'genotype_pattern'};
my ($genotype_pattern_diff, $affected_spores) = compare_genotype_patterns($current_block_genotype_pattern, $next_block_genotype_pattern);
my ($genotype_switches_count_pattern, $genotype_switches_count_pattern_sorted, $uniq_single_switch_count, $genotype_switch_boundaries) = check_genotype_switches($current_block_id, $next_block_id, \%{$linkage_blocks{$chr}});
# for debug
# print "chr=$chr, i=$i, current_block_id=$current_block_id, next_block_id=$next_block_id\n";
# print "current_block_genotype_pattern=$current_block_genotype_pattern, next_block_genotype_pattern=$next_block_genotype_pattern\n";
# print "genotype_pattern_diff=$genotype_pattern_diff, affected_spores=$affected_spores\n";
# print "genotype_switches_count_pattern=$genotype_switches_count_pattern, genotype_switches_count_pattern_sorted: $genotype_switches_count_pattern_sorted\n";
# print "uniq_single_switch_count=$uniq_single_switch_count\n";
# print "\n";
# here, $genotype_pattern_diff can only be 2, 4, or 0, corresponding to single CO, double CO or no CO.
# for $affected_spores, 1:0:1:0 means spore_a and spore_c have genotype pattern switches.
if ($genotype_pattern_diff == 2) {
# single CO
if ($next_block_id - $current_block_id == 1) {
# the two linkage blocks are neighbours to each other on the chromosome with no GC-related blocks in between
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type1_CO"; # single CO without associated GC
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type1_CO_block_start = $current_block_id;
my $type1_CO_block_end = $next_block_id;
my $type1_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type1_CO_block_start, $type1_CO_block_end, $type1_CO_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
# there is GC-related blocks in between
# check genotype switches in the interstitial GC-related blocks
# print "current block id: $current_block_id, next block id: $next_block_id, sorted genotype switch count pattern: $genotype_switches_count_pattern, uniq_single_switch count: $uniq_single_switch_count\n";
if (($genotype_switches_count_pattern_sorted eq "0:0:1:1") and ($uniq_single_switch_count == 2)) {
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type2_CO"; # single CO with associated GC on chromatids involved in the CO and no other GC existed
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type2_CO_block_start = $current_block_id;
my $type2_CO_block_end = $next_block_id;
my $type2_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type2_CO_block_start, $type2_CO_block_end, $type2_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type2_GC"; # GC associated with CO
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my $type2_GC_block_start = $type2_CO_block_start + 1;
my $type2_GC_block_end = $type2_CO_block_end - 1;
my $type2_GC_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type2_GC_block_start, $type2_GC_block_end, $type2_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} elsif (($genotype_switches_count_pattern_sorted eq "0:1:1:2") and ($uniq_single_switch_count == 1)) {
# chromatids with the single switch have switches at the same location
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type3_CO"; # single CO associated with GC on the chromatid not involved in the CO
my $type3_CO_block_start = "";
my $type3_CO_block_end = "";
my @genotype_switches_count_pattern = split /:/, $genotype_switches_count_pattern;
my @genotype_switch_boundaries = split /:/, $genotype_switch_boundaries;
@{$CO_associated_GC{$recombination_event_id}} = ();
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] == 1) {
($type3_CO_block_start, $type3_CO_block_end) = ($genotype_switch_boundaries[$i] =~ /(\S+)\|(\S+)/);
last;
}
}
my $type3_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type3_CO_block_start, $type3_CO_block_end, $type3_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type7_GC"; # GC on the chromatid not involved in CO
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my $type7_GC_block_start;
my $type7_GC_block_end;
my @type7_GC_affected_spores = (0, 0, 0, 0);
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] == 2) {
$type7_GC_affected_spores[$i] = 1;
my ($b1, $b2) = split ",", $genotype_switch_boundaries[$i];
($type7_GC_block_start) = ($b1 =~ /\|(\S+)/);
($type7_GC_block_end) = ($b2 =~ /(\S+)\|/);
last;
}
}
my $type7_GC_affected_spores = join ":", @type7_GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type7_GC_block_start, $type7_GC_block_end, $type7_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} elsif (($genotype_switches_count_pattern_sorted eq "0:1:1:2") and ($uniq_single_switch_count == 2)) {
# chromatids with the single switch have switches at different locations
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type4_CO"; # single CO with 2 associated GCs, one on the chromatids involved in the CO and the other on one of the chromatids not involved in the CO
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type4_CO_block_start = $current_block_id;
my $type4_CO_block_end = $next_block_id;
my $type4_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type4_CO_block_start, $type4_CO_block_end, $type4_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type9_GC"; # GC on one of the chromatids not involved in CO
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my $type9_GC_block_start;
my $type9_GC_block_end;
my @genotype_switches_count_pattern = split /:/, $genotype_switches_count_pattern;
my @genotype_switch_boundaries = split /:/, $genotype_switch_boundaries;
my @type9_GC_affected_spores = (0, 0, 0, 0);
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] == 2) {
$type9_GC_affected_spores[$i] = 1;
my ($b1, $b2) = split ",", $genotype_switch_boundaries[$i];
($type9_GC_block_start) = ($b1 =~ /\|(\S+)/);
($type9_GC_block_end) = ($b2 =~ /(\S+)\|/);
}
}
my $type9_GC_affected_spores = join ":", @type9_GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type9_GC_block_start, $type9_GC_block_end, $type9_GC_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type8_GC"; # GC on the chromatids involved in CO
push @{$CO_associated_GC{$recombination_event_id - 2}}, $recombination_event_id;
my $type8_GC_block_start;
my $type8_GC_block_end;
my @type8_GC_block_boundaries = ();
my @type8_GC_affected_spores = (0, 0, 0, 0);
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] == 1) {
$type8_GC_affected_spores[$i] = 1;
my ($b1, $b2) = split /\|/, $genotype_switch_boundaries[$i];
# print "chr=$chr, genotype_switch_boundary= $genotype_switch_boundaries[$i], b1=$b1, b2=$b2\n";
push @type8_GC_block_boundaries, $b1;
push @type8_GC_block_boundaries, $b2;
}
}
my @type8_GC_block_boundaries_sorted = sort {$a <=> $b} @type8_GC_block_boundaries;
$type8_GC_block_start = $type8_GC_block_boundaries_sorted[1];
$type8_GC_block_end = $type8_GC_block_boundaries_sorted[2];
my $type8_GC_affected_spores = join ":", @type8_GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type8_GC_block_start, $type8_GC_block_end, $type8_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type6_CO"; # single CO with complex GC patterns
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type6_CO_block_start = $current_block_id;
my $type6_CO_block_end = $next_block_id;
my $type6_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type6_CO_block_start, $type6_CO_block_end, $type6_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type15_GC"; # complex GC associated with Type6_CO
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my @type15_GC_block_boundaries = ();
my $type15_GC_block_start = $type6_CO_block_start + 1;;
my $type15_GC_block_end = $type6_CO_block_end - 1;
my @type15_GC_affected_spores = split /:/, $type6_CO_affected_spores;
my @genotype_switches_count_pattern = split /:/, $genotype_switches_count_pattern;
my @genotype_switch_boundaries = split /:/, $genotype_switch_boundaries;
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] > 1) {
$type15_GC_affected_spores[$i] = "1*";
# my @b = split ",", $genotype_switch_boundaries[$i];
# foreach my $b (@b) {
# my ($b1, $b2) = split /\|/, $b;
# push @type15_GC_block_boundaries, $b1;
# push @type15_GC_block_boundaries, $b2;
# }
# my @type15_GC_block_boundaries_sorted = sort {$a <=> $b} @type15_GC_block_boundaries;
# $type15_GC_block_start = $type15_GC_block_boundaries[1];
# $type15_GC_block_end = $type15_GC_block_boundaries[-2];
}
}
my $type15_GC_affected_spores = join ":", @type15_GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type15_GC_block_start, $type15_GC_block_end, $type15_GC_affected_spores, \%recombination_events, \%linkage_blocks);
}
}
} elsif ($genotype_pattern_diff == 4) {
# double CO
if ($next_block_id - $current_block_id == 1) {
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type7_CO"; # double CO without associated GC
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type7_CO_block_start = $current_block_id;
my $type7_CO_block_end = $next_block_id;
my $type7_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type7_CO_block_start, $type7_CO_block_end, $type7_CO_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
my ($genotype_switches_count_pattern, $genotype_switches_count_pattern_sorted, $uniq_single_switch_count, $genotype_switch_boundaries) = check_genotype_switches($current_block_id, $next_block_id, \%{$linkage_blocks{$chr}});
# print "current block id: $current_block_id, next block id: $next_block_id, sorted genotype switch count pattern: $genotype_switches_count_pattern, uniq_single_switch count: $uniq_single_switch_count\n";
if ($genotype_switches_count_pattern_sorted eq "1:1:1:1") {
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type8_CO"; # double CO with 1 or 2 associated GC(s)
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type8_CO_block_start = $current_block_id;
my $type8_CO_block_end = $next_block_id;
my $type8_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type8_CO_block_start, $type8_CO_block_end, $type8_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type12_GC"; # GC(s) assoicated with double CO
# for now, we use the Type6_CO coordinates for Type9_GC since there could be multiple scenarios considering the exact GC locations
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my $type12_GC_block_start = $type8_CO_block_start + 1;
my $type12_GC_block_end = $type8_CO_block_end - 1;
my $type12_GC_affected_spores = $affected_spores; # for now, consider all spores were affected. manual inspection will be needed to nail down the exact affected spores.
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type12_GC_block_start, $type12_GC_block_end, $type12_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
$recombination_event_id++;
my $event_type = "CO";
my $event_subtype = "Type9_CO"; # double CO with complex GC pattern
@{$CO_associated_GC{$recombination_event_id}} = ();
my $type9_CO_block_start = $current_block_id;
my $type9_CO_block_end = $next_block_id;
my $type9_CO_affected_spores = $affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type9_CO_block_start, $type9_CO_block_end, $type9_CO_affected_spores, \%recombination_events, \%linkage_blocks);
$recombination_event_id++;
$event_type = "GC";
$event_subtype = "Type16_GC"; # complex GC associated with Type9_CO
push @{$CO_associated_GC{$recombination_event_id - 1}}, $recombination_event_id;
my $type16_GC_block_start = $type9_CO_block_start + 1;
my $type16_GC_block_end = $type9_CO_block_end - 1;
my @type16_GC_affected_spores = (1, 1, 1, 1);
my @genotype_switches_count_pattern = split /:/, $genotype_switches_count_pattern;
for (my $i = 0; $i < 4; $i++) {
if ($genotype_switches_count_pattern[$i] > 1) {
$type16_GC_affected_spores[$i] = "1*";
}
}
my $type16_GC_affected_spores = join ":", @type16_GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type16_GC_block_start, $type16_GC_block_end, $type16_GC_affected_spores, \%recombination_events, \%linkage_blocks);
}
}
} else {
# print "no CO between block $current_block_id and $next_block_id\n";
# print "genotype_pattern_diff = $genotype_pattern_diff, current_block_id=$current_block_id, next_block_id=$next_block_id\n";
}
}
merge_nearby_COs($d, \%recombination_events, \%linkage_blocks, \%CO_associated_GC, \@merging_log);
}
} elsif (($segregation_pattern eq "1:3:0") or ($segregation_pattern eq "3:1:0")) {
foreach my $block_id (@block_subclass) {
if (($block_id == 1) or ($block_id == $block_total_count)) {
# GC at the chromosome start or end
$recombination_event_id++;
my $event_type = "GC";
my $event_subtype = "Type4_GC"; # GC adjacent to chromosome ends
my $type4_GC_block_start = $block_id;
my $type4_GC_block_end = $block_id;
my $type4_GC_affected_spores;
if ($block_id == 1) {
$type4_GC_affected_spores = $linkage_blocks{$chr}{$block_id+1}{'genotype_switch_pattern'};
} else {
$type4_GC_affected_spores = $linkage_blocks{$chr}{$block_id}{'genotype_switch_pattern'};
}
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type4_GC_block_start, $type4_GC_block_end, $type4_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
my $query_marker_pos_start = $linkage_blocks{$chr}{$block_id}{'marker_pos_start'};
my $query_marker_pos_end = $linkage_blocks{$chr}{$block_id}{'marker_pos_end'};
my %closest_CO = find_closest_event("CO", $chr, $query_marker_pos_start, $query_marker_pos_end, \%recombination_events, \%linkage_blocks);
my %closest_GC = find_closest_event("GC", $chr, $query_marker_pos_start, $query_marker_pos_end, \%recombination_events, \%linkage_blocks);
if ($closest_CO{'distance_to_query'} <= 0) {
;
} elsif (($closest_GC{'distance_to_query'} <= 0) and ($closest_GC{'subtype'} =~ /Type(6|17)\_GC/)) {
;
} elsif (($closest_CO{'distance_to_query'} > 0) and ($closest_CO{'distance_to_query'} <= $d)) {
# the GC is in adjacency to COs
$recombination_event_id++;
my $event_type = "GC";
my $closest_event_id = $closest_CO{'id'};
my $closest_event_type = $closest_CO{'type'};
my $closest_event_subtype = $closest_CO{'subtype'};
my $closest_event_chr = $closest_CO{'chr'};
my $closest_event_block_start = $closest_CO{'block_start'};
my $closest_event_block_end = $closest_CO{'block_end'};
my $closest_event_marker_pos_start = $closest_CO{'marker_pos_start'};
my $closest_event_marker_pos_end = $closest_CO{'marker_pos_end'};
my $closest_event_affected_spores = $closest_CO{'affected_spores'};
my @closest_event_affected_spores = split /:/, $closest_event_affected_spores;
my $GC_affected_spores = $linkage_blocks{$chr}{$block_id}{'genotype_switch_pattern'};
my @GC_affected_spores = split /:/, $GC_affected_spores;
my $flag = 0;
for (my $i = 0; $i < 4; $i++) {
if (($closest_event_affected_spores[$i] == 1) and ($GC_affected_spores[$i] == 1)) {
$flag = 1;
last;
}
}
if ($flag == 0) {
# the GC is not on the chromatids involved in CO
my $event_subtype = "Type14_GC"; # GC near CO and GC is on one of the chromatids not involved in CO
my $type14_GC_block_start = $block_id;
my $type14_GC_block_end = $block_id;
my $type14_GC_affected_spores = $GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type14_GC_block_start, $type14_GC_block_end, $type14_GC_affected_spores, \%recombination_events, \%linkage_blocks);
# update CO subtype for the closest CO
# if ($closest_event_subtype =~ /Type1_CO/) {
# record_recombination_event($closest_event_id, $closest_event_type, "Type3_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# } elsif ($closest_event_subtype =~ /Type2_CO/) {
# my $closest_event_block_id = $closest_event_block_start;
# my $closest_event_GC_tract_genotype_pattern = $linkage_blocks{$closest_event_chr}{$closest_event_block_id}{'genotype_pattern'};
# my @closest_event_GC_tract_genotype_pattern = split /:/, $closest_event_GC_tract_genotype_pattern;
# my $type14_GC_genotype_pattern = $linkage_blocks{$chr}{$block_id}{'genotype_pattern'};
# my @type14_GC_genotype_pattern = split /:/, $type14_GC_genotype_pattern;
# my $closest_event_GC_tract_genotype;
# my $type14_GC_genotype;
# for(my $i = 0; $i < 4; $i++) {
# if ($closest_event_affected_spores[$i] == 1) {
# $closest_event_GC_tract_genotype = $closest_event_GC_tract_genotype_pattern[$i];
# last;
# }
# }
# for(my $i = 0; $i < 4; $i++) {
# if ($GC_affected_spores[$i] == 1) {
# $type14_GC_genotype = $type14_GC_genotype_pattern[$i];
# last;
# }
# }
# if ($closest_event_GC_tract_genotype eq $type14_GC_genotype) {
# record_recombination_event($closest_event_id, $closest_event_type, "Type4_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# } else {
# record_recombination_event($closest_event_id, $closest_event_type, "Type5_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# }
# } elsif ($closest_event_subtype =~ /Type(3|4|5|6)_CO/) {
# record_recombination_event($closest_event_id, $closest_event_type, "Type6_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# } else {
# record_recombination_event($closest_event_id, $closest_event_type, "Type9_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# }
} else {
# the GC is on the chromatids involved in CO
my $event_subtype = "Type13_GC"; # GC near CO but not contiguouns with it.
my $type13_GC_block_start = $block_id;
my $type13_GC_block_end = $block_id;
my $type13_GC_affected_spores = $GC_affected_spores;
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type13_GC_block_start, $type13_GC_block_end, $type13_GC_affected_spores, \%recombination_events, \%linkage_blocks);
# # update CO subtype for the closest CO
# if ($closest_event_subtype =~ /Type(1|2|3|4|5|6)_CO/) {
# record_recombination_event($closest_event_id, $closest_event_type, "Type6_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# } else {
# record_recombination_event($closest_event_id, $closest_event_type, "Type9_CO", $closest_event_chr, $closest_event_block_start, $closest_event_block_end, $closest_event_affected_spores, \%recombination_events, \%linkage_blocks);
# }
}
# } elsif (($closest_GC{'distance_to_query'} == 0) and ($closest_GC{'subtype'} =~ /Type(6|17)_GC/)) {
# # already counted when defining Type6_GC and Type17_GC, skip
# ;
} elsif ($closest_CO{'distance_to_query'} > $d) {
# the GC is not in adjacency to COs or Type6_GC, Type17_GC
$recombination_event_id++;
my $event_type = "GC";
my $event_subtype = "Type1_GC"; # NCO
my $type1_GC_block_start = $block_id;
my $type1_GC_block_end = $block_id;
my $type1_GC_affected_spores = $linkage_blocks{$chr}{$block_id}{'genotype_switch_pattern'};
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type1_GC_block_start, $type1_GC_block_end, $type1_GC_affected_spores, \%recombination_events, \%linkage_blocks);
}
}
}
} elsif (($segregation_pattern eq "0:4:0") or ($segregation_pattern eq "4:0:0")) {
foreach my $block_id (@block_subclass) {
if (($block_id == 1) or ($block_id == $block_total_count)) {
# GC at the chromosome start or end
$recombination_event_id++;
my $event_type = "GC";
my $event_subtype = "Type5_GC"; # double GC adjacent to chromosome ends
my $type5_GC_block_start = $block_id;
my $type5_GC_block_end = $block_id;
my $type5_GC_affected_spores;
if ($block_id == 1) {
$type5_GC_affected_spores = $linkage_blocks{$chr}{$block_id + 1}{'genotype_switch_pattern'};
} else {
$type5_GC_affected_spores = $linkage_blocks{$chr}{$block_id}{'genotype_switch_pattern'};
}
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type5_GC_block_start, $type5_GC_block_end, $type5_GC_affected_spores, \%recombination_events, \%linkage_blocks);
} else {
my $query_marker_pos_start = $linkage_blocks{$chr}{$block_id}{'marker_pos_start'};
my $query_marker_pos_end = $linkage_blocks{$chr}{$block_id}{'marker_pos_end'};
my %closest_CO = find_closest_event("CO", $chr, $query_marker_pos_start, $query_marker_pos_end, \%recombination_events, \%linkage_blocks);
my %closest_GC = find_closest_event("GC", $chr, $query_marker_pos_start, $query_marker_pos_end, \%recombination_events, \%linkage_blocks);
if (($closest_GC{'distance_to_query'} <= 0) and ($closest_GC{'subtype'} =~ /Type(6|17)\_GC/)) {
;
} elsif ($closest_CO{'distance_to_query'} > $d) {
# the GC is not in ajacency to COs
$recombination_event_id++;
my $event_type = "GC";
my $event_subtype = "Type3_GC"; # 4:0 GC
my $type3_GC_block_start = $block_id;
my $type3_GC_block_end = $block_id;
my $type3_GC_affected_spores = $linkage_blocks{$chr}{$block_id}{'genotype_switch_pattern'};
record_recombination_event($recombination_event_id, $event_type, $event_subtype, $chr, $type3_GC_block_start, $type3_GC_block_end, $type3_GC_affected_spores, \%recombination_events, \%linkage_blocks);
}
}
}
}
}
}
}
}
# generate outputs
my $recombination_event_output = "$output_dir/$tetrad_id/$prefix.events.txt";
my $recombination_event_output_fh = write_file($recombination_event_output);
print $recombination_event_output_fh "tetrad_id\tevent_id\tevent_type\tevent_subtype\tchr\tmarker_pos_start\tmarker_pos_end\tadjusted_pos_start\tadjusted_pos_end\tadjusted_size\tmarker_raw_index_start\tmarker_raw_index_end\tmarker_effective_index_start\tmarker_effective_index_end\taffected_spores\n";
my %event_type_count = ();
my $total_CO_count = 0;
my $total_GC_count = 0;
# foreach my $event_id (sort {(($a =~ /(\d+\.?\d+?)/)[0] || 0) <=> (($b =~ /(\d+\.?\d+?)/)[0] || 0)} keys %recombination_events) {
my %event_sort_by_coord = ();
foreach my $event_id (sort custom_sort keys %recombination_events) {
my $type = $recombination_events{$event_id}{'type'};
my $subtype = $recombination_events{$event_id}{'subtype'};
my $chr = $recombination_events{$event_id}{'chr'};
my $marker_pos_start = $recombination_events{$event_id}{'marker_pos_start'};
my $marker_pos_end = $recombination_events{$event_id}{'marker_pos_end'};
my $marker_raw_index_start = $recombination_events{$event_id}{'marker_raw_index_start'};
my $marker_raw_index_end = $recombination_events{$event_id}{'marker_raw_index_end'};
my $marker_effective_index_start = $recombination_events{$event_id}{'marker_effective_index_start'};
my $marker_effective_index_end = $recombination_events{$event_id}{'marker_effective_index_end'};
my $adjusted_pos_start;
my $adjusted_pos_end;
if ($type eq "CO") {
$total_CO_count++;
$adjusted_pos_start = ($marker_pos_start + $marker_pos_end)/2;
$adjusted_pos_end = ($marker_pos_start + $marker_pos_end)/2;
} else {
# type is GC
$total_GC_count++;
if ((exists $markers{'effective'}{$marker_effective_index_start - 1}) and ($markers{'effective'}{$marker_effective_index_start - 1}{'chr'} eq $chr)) {
# the previous marker is an internal marker on the same chromosome
my $previous_marker_pos = $markers{'effective'}{$marker_effective_index_start-1}{'pos'};
$adjusted_pos_start = ($previous_marker_pos + $marker_pos_start)/2;
} else {
# the GC starting marker is the first marker on the chromosome
$adjusted_pos_start = $marker_pos_start;
}
if ((exists $markers{'effective'}{$marker_effective_index_end + 1}) and ($markers{'effective'}{$marker_effective_index_end + 1}{'chr'} eq $chr)) {
# the next marker is an internal marker on the same chromosome
my $next_marker_pos = $markers{'effective'}{$marker_effective_index_end + 1}{'pos'};
$adjusted_pos_end = ($marker_pos_end + $next_marker_pos)/2;
} else {
# the GC ending marker is the last marker on the chromosome
$adjusted_pos_end = $marker_pos_end;
}
}
my $adjusted_size = $adjusted_pos_end - $adjusted_pos_start + 1;
my $affected_spores = $recombination_events{$event_id}{'affected_spores'};
$event_sort_by_coord{$chr}{$marker_pos_start}{$marker_pos_end}{$type}{$event_id} = "$tetrad_id\t$event_id\t$type\t$subtype\t$chr\t$marker_pos_start\t$marker_pos_end\t$adjusted_pos_start\t$adjusted_pos_end\t$adjusted_size\t$marker_raw_index_start\t$marker_raw_index_end\t$marker_effective_index_start\t$marker_effective_index_end\t$affected_spores\n";
if (exists $event_type_count{$type}{$subtype}) {
$event_type_count{$type}{$subtype}++;
} else {
$event_type_count{$type}{$subtype} = 1;
}
}
foreach my $chr (sort keys %event_sort_by_coord) {
foreach my $s (sort {$a <=> $b} keys %{$event_sort_by_coord{$chr}}) {
foreach my $e (sort {$a <=> $b} keys %{$event_sort_by_coord{$chr}{$s}}) {
foreach my $t (sort keys %{$event_sort_by_coord{$chr}{$s}{$e}}) {
foreach my $id (sort keys %{$event_sort_by_coord{$chr}{$s}{$e}{$t}}) {
print $recombination_event_output_fh $event_sort_by_coord{$chr}{$s}{$e}{$t}{$id}
}
}
}
}
}
# event type count summary output
my $event_type_count_out = "$output_dir/$tetrad_id/$prefix.event_type_count.txt";
my $event_type_count_out_fh = write_file($event_type_count_out);
print $event_type_count_out_fh "total CO events: $total_CO_count\n";
print $event_type_count_out_fh "total GC events: $total_GC_count\n";
print $event_type_count_out_fh "################################\n";
foreach my $type (sort keys %event_type_count) {
my %subtype_index = ();
foreach my $subtype (sort keys %{$event_type_count{$type}}) {
my ($subtype_index) = ($subtype =~ /Type(\d+)_/);
$subtype_index{$subtype} = $subtype_index;
}
foreach my $subtype (sort {$subtype_index{$a} <=> $subtype_index{$b}} keys %subtype_index) {
print $event_type_count_out_fh "$type\t$subtype\t$event_type_count{$type}{$subtype}\n";
}
}
print $event_type_count_out_fh "################################\n";
# output merging record
my $merging_log_out = "$output_dir/$tetrad_id/$prefix.merging_log.txt";
my $merging_log_out_fh = write_file($merging_log_out);
foreach my $line (@merging_log) {
print $merging_log_out_fh "$line\n";
}
# output CO_associated GC output
my $CO_associated_GC_out = "$output_dir/$tetrad_id/$prefix.co_associated_gc.txt";
my $CO_associated_GC_out_fh = write_file($CO_associated_GC_out);
print $CO_associated_GC_out_fh "event_id_for_CO\tCO_subtype\tevent_id_for_associated_GC\tassociated_GC_subtype\n";
foreach my $CO_id (sort keys %CO_associated_GC) {
my $CO_subtype = $recombination_events{$CO_id}{'subtype'};
my @GC_id = sort @{$CO_associated_GC{$CO_id}};
if ((scalar @GC_id) > 0) {
foreach my $GC_id (@GC_id) {
# print "CO_id: $CO_id, GC_id: $GC_id\n";
my $GC_subtype = $recombination_events{$GC_id}{'subtype'};
print $CO_associated_GC_out_fh "$CO_id\t$CO_subtype\t$GC_id\t$GC_subtype\n";
}
}
}
# output marker record
my $markers_out = "$output_dir/$tetrad_id/$prefix.markers.txt";
my $markers_out_fh = write_file($markers_out);
print $markers_out_fh "tetrad_id\tchr\tpos\traw_index\teffective_index\tgenotype_pattern\tsegregation_pattern\n";
foreach my $raw_index (sort {$a<=>$b} keys %{$markers{'raw'}}) {
my $chr = $markers{'raw'}{$raw_index}{'chr'};
my $pos = $markers{'raw'}{$raw_index}{'pos'};
my $genotype_pattern = $markers{'raw'}{$raw_index}{'genotype_pattern'};
my $segregation_pattern = $markers{'raw'}{$raw_index}{'segregation_pattern'};
my $effective_index = $markers{'raw'}{$raw_index}{'effective_index'};
print $markers_out_fh "$tetrad_id\t$chr\t$pos\t$raw_index\t$effective_index\t$genotype_pattern\t$segregation_pattern\n";
}
# output preliminary linkage blocks
my $preliminary_linkage_blocks_out = "$output_dir/$tetrad_id/$prefix.preliminary_linkage_blocks.txt";
my $preliminary_linkage_blocks_out_fh = write_file($preliminary_linkage_blocks_out);
print $preliminary_linkage_blocks_out_fh "tetrad_id\tchr:preliminary_block_id\tmarker_pos_start-marker_pos_end\tmarker_raw_index_start-marker_raw_index_end\tmarker_effective_index_start-marker_effective_index_end\tgenotype_pattern\tsegregation_pattern\n";
foreach my $chr (sort keys %preliminary_linkage_blocks) {
foreach my $block_id (sort {$a<=>$b} keys %{$preliminary_linkage_blocks{$chr}}) {
my $marker_pos_start = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_pos_start'};
my $marker_pos_end = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_pos_end'};
my $marker_raw_index_start = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_raw_index_start'};
my $marker_raw_index_end = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_raw_index_end'};
my $marker_effective_index_start = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_effective_index_start'};
my $marker_effective_index_end = $preliminary_linkage_blocks{$chr}{$block_id}{'marker_effective_index_end'};
my $genotype_pattern = $preliminary_linkage_blocks{$chr}{$block_id}{'genotype_pattern'}; # genotype_a:genotype_b:genotype_c:genotype_d
my $segregation_pattern = $preliminary_linkage_blocks{$chr}{$block_id}{'segregation_pattern'};
print $preliminary_linkage_blocks_out_fh "$tetrad_id\t$chr:$block_id\t$marker_pos_start-$marker_pos_end\t$marker_raw_index_start-$marker_raw_index_end\t$marker_effective_index_start-$marker_effective_index_end\t$genotype_pattern\t$segregation_pattern\n";
}
}
# output linkage blocks
my $linkage_blocks_out = "$output_dir/$tetrad_id/$prefix.linkage_blocks.txt";
my $linkage_blocks_out_fh = write_file($linkage_blocks_out);
print $linkage_blocks_out_fh "tetrad_id\tchr:block_id\tmarker_pos_start-marker_pos_end\tmarker_raw_index_start-marker_raw_index_end\tmarker_effective_index_start-marker_effective_index_end\tgenotype_pattern\tsegregation_pattern\n";
foreach my $chr (sort keys %linkage_blocks) {
foreach my $block_id (sort {$a<=>$b} keys %{$linkage_blocks{$chr}}) {
my $marker_pos_start = $linkage_blocks{$chr}{$block_id}{'marker_pos_start'};
my $marker_pos_end = $linkage_blocks{$chr}{$block_id}{'marker_pos_end'};
my $marker_raw_index_start = $linkage_blocks{$chr}{$block_id}{'marker_raw_index_start'};
my $marker_raw_index_end = $linkage_blocks{$chr}{$block_id}{'marker_raw_index_end'};
my $marker_effective_index_start = $linkage_blocks{$chr}{$block_id}{'marker_effective_index_start'};
my $marker_effective_index_end = $linkage_blocks{$chr}{$block_id}{'marker_effective_index_end'};
my $genotype_pattern = $linkage_blocks{$chr}{$block_id}{'genotype_pattern'}; # genotype_a:genotype_b:genotype_c:genotype_d
my $segregation_pattern = $linkage_blocks{$chr}{$block_id}{'segregation_pattern'};
print $linkage_blocks_out_fh "$tetrad_id\t$chr:$block_id\t$marker_pos_start-$marker_pos_end\t$marker_raw_index_start-$marker_raw_index_end\t$marker_effective_index_start-$marker_effective_index_end\t$genotype_pattern\t$segregation_pattern\n";
}
}
}
}
}
sub read_file {
my $file = shift @_;
my $fh;
if ($file =~ /\.gz$/) {
open($fh, "gunzip -c $file |") or die "can't open pipe to $file";
} else {
open($fh, $file) or die "can't open $file";
}
return $fh;
}
sub write_file {
my $file = shift @_;
my $fh;
if ($file =~ /\.gz$/) {
open($fh, "| gzip -c >$file") or die "can't open $file\n";
} else {
open($fh, ">$file") or die "can't open $file\n";
}
return $fh;
}
sub parse_sample_table_by_tetrad {
my $fh = shift @_;
my %sample_table = ();
while (<$fh>) {
chomp;
/^#/ and next;
/^\s*$/ and next;
my ($sample_id, $tetrad_id, $spore_id, $PE_reads_files, $cross_pair, $note) = split /\s+/, $_;
if (exists $sample_table{$tetrad_id}{'spore_index'}) {
push @{$sample_table{$tetrad_id}{'spore_index'}}, $spore_id;
} else {
@{$sample_table{$tetrad_id}{'spore_index'}} = ($spore_id);
}
$sample_table{$tetrad_id}{'cross_pair'} = $cross_pair;
$sample_table{$tetrad_id}{'note'} = $note;
}
return %sample_table;
}
sub calculate_spore_availibility {
my $tetrads_hashref = shift @_;
my @spore_index_array = qw(a b c d);
foreach my $tetrad_id (sort keys %$tetrads_hashref) {
my %spore_availibility_hash = (
'a' => 0,
'b' => 0,
'c' => 0,
'd' => 0
);
my @spore_availibility_array = ();
foreach my $spore_index (@{$$tetrads_hashref{$tetrad_id}{'spore_index'}}) {
$spore_availibility_hash{$spore_index} = 1;
}
foreach my $spore_index (@spore_index_array) {
push @spore_availibility_array, $spore_availibility_hash{$spore_index};
}
$$tetrads_hashref{$tetrad_id}{'spore_availibility'} = join ":", @spore_availibility_array;
}
}
sub filter_tetrad_with_inviable_spores {
my $tetrads_hashref = shift @_;
foreach my $tetrad_id (sort keys %$tetrads_hashref) {
my $viable_spore_count = scalar @{$$tetrads_hashref{$tetrad_id}{'spore_index'}};
if ($viable_spore_count < 4) {
delete $$tetrads_hashref{$tetrad_id};
}
}
}
sub compare_genotype_patterns {
my ($genotype_pattern1, $genotype_pattern2) = @_;
my @genotype_pattern1 = split /:/, $genotype_pattern1;
my @genotype_pattern2 = split /:/, $genotype_pattern2;
my $genotype_diff_count = 0;
my @affected_spores = (0, 0, 0, 0);
for (my $i = 0; $i < 4; $i++) {
if ($genotype_pattern1[$i] ne $genotype_pattern2[$i]) {
$genotype_diff_count++;
$affected_spores[$i] = 1;
}
}
my $affected_spores = join ":", @affected_spores;
return ($genotype_diff_count, $affected_spores);
}
sub compare_genotype_patterns_with_NA {
my ($genotype_pattern1, $genotype_pattern2) = @_;
my @genotype_pattern1 = split /:/, $genotype_pattern1;
my @genotype_pattern2 = split /:/, $genotype_pattern2;
my $genotype_diff_count = 0;
my $genotype_diff_by_NA_count = 0;
my @affected_spores = (0, 0, 0, 0);
for (my $i = 0; $i < 4; $i++) {
if ($genotype_pattern1[$i] ne $genotype_pattern2[$i]) {
$genotype_diff_count++;
if (($genotype_pattern1[$i] eq "NA") or ($genotype_pattern2[$i] eq "NA")) {
$genotype_diff_by_NA_count++;
} else {
$affected_spores[$i] = 1;
}
}
}
my $affected_spores = join ":", @affected_spores;
return ($genotype_diff_count, $genotype_diff_by_NA_count, $affected_spores);
}
sub parse_tetrad_genotypes_file {
my ($fh, $parent1, $parent2, $ref, $spore_availibility, $tetrad_genotypes_hashref, $linkage_blocks_hashref, $markers_hashref) = @_;
my $marker_raw_index = 0;
my $marker_effective_index = 0;
my $block_id = 0;
my $genotype_pattern_just_saw;
my @spore_availibility_array = split ":", $spore_availibility;
while (<$fh>) {
chomp;
/^#/ and next;
/^\s*$/ and next;
my ($chr, $pos, $reftag, $genotype_a, $genotype_b, $genotype_c, $genotype_d) = split /\t/, $_;
my $parent1_genotype_count = 0;
my $parent2_genotype_count = 0;
my $NA_genotype_count = 0;
my $missing_genotype_count = 0;
my @genotypes = ($genotype_a, $genotype_b, $genotype_c, $genotype_d);
for (my $i = 0; $i < (scalar @genotypes); $i++) {
if ($genotypes[$i] eq $parent1) {
$parent1_genotype_count++;
} elsif ($genotypes[$i] eq $parent2) {
$parent2_genotype_count++;
} else {
$NA_genotype_count++;
if ($spore_availibility_array[$i] == 1) {
$missing_genotype_count++;
}
}
}
my $segregation_pattern = "$parent1_genotype_count:$parent2_genotype_count:$NA_genotype_count";
my $genotype_pattern = "$genotype_a:$genotype_b:$genotype_c:$genotype_d";
$marker_raw_index++;
$$markers_hashref{'raw'}{$marker_raw_index}{'chr'} = $chr;
$$markers_hashref{'raw'}{$marker_raw_index}{'pos'} = $pos;
$$markers_hashref{'raw'}{$marker_raw_index}{'reftag'} = $reftag;
$$markers_hashref{'raw'}{$marker_raw_index}{'genotype_pattern'} = $genotype_pattern;
$$markers_hashref{'raw'}{$marker_raw_index}{'segregation_pattern'} = $segregation_pattern;
$$markers_hashref{'raw'}{$marker_raw_index}{'effective_index'} = "";
##################
# ignore all NA-containing markers for now
if ($NA_genotype_count > 0) {
next;
} else {
# only consider and index non-ambiguous markers, this will help calculating inter-marker boundaries
$marker_effective_index++;
$$markers_hashref{'raw'}{$marker_raw_index}{'effective_index'} = $marker_effective_index;
$$markers_hashref{'effective'}{$marker_effective_index}{'chr'} = $chr;
$$markers_hashref{'effective'}{$marker_effective_index}{'pos'} = $pos;
$$markers_hashref{'effective'}{$marker_effective_index}{'reftag'} = $reftag;
$$markers_hashref{'effective'}{$marker_effective_index}{'genotype_pattern'} = $genotype_pattern;
$$markers_hashref{'effective'}{$marker_effective_index}{'segregation_pattern'} = $segregation_pattern;
$$markers_hashref{'effective'}{$marker_effective_index}{'raw_index'} = $marker_raw_index;
}
$$tetrad_genotypes_hashref{$chr}{$pos}{'marker_raw_index'} = $marker_raw_index;
$$tetrad_genotypes_hashref{$chr}{$pos}{'marker_effective_index'} = $marker_effective_index;
$$tetrad_genotypes_hashref{$chr}{$pos}{'genotype_a'} = $genotype_a;
$$tetrad_genotypes_hashref{$chr}{$pos}{'genotype_b'} = $genotype_b;
$$tetrad_genotypes_hashref{$chr}{$pos}{'genotype_c'} = $genotype_c;
$$tetrad_genotypes_hashref{$chr}{$pos}{'genotype_d'} = $genotype_d;
$$tetrad_genotypes_hashref{$chr}{$pos}{'segregation_pattern'} = $segregation_pattern;
$$tetrad_genotypes_hashref{$chr}{$pos}{'genotype_pattern'} = $genotype_pattern;
if (not exists $$linkage_blocks_hashref{$chr}) {
# this is the first marker on the chromosome
$block_id++;
@{$$linkage_blocks_hashref{$chr}{$block_id}{'markers'}} = ($pos);
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_start'} = $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_end'} = $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_start'} = $marker_raw_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_end'} = $marker_raw_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_start'} = $marker_effective_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_end'} = $marker_effective_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'segregation_pattern'} = $segregation_pattern;
$$linkage_blocks_hashref{$chr}{$block_id}{'genotype_pattern'} = $genotype_pattern;
$$linkage_blocks_hashref{$chr}{$block_id}{'genotype_switch_pattern'} = "0:0:0:0"; # genotype switches relative to the previous linkeage block
} elsif ($genotype_pattern eq $genotype_pattern_just_saw) {
# markers with the same genotype pattern: genotype_a:genotype_b:genotype_c:genotype_d
push @{$$linkage_blocks_hashref{$chr}{$block_id}{'markers'}}, $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_end'} = $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_end'} = $marker_raw_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_end'} = $marker_effective_index;
} else {
# markers with different genotype pattern: genotype_a:genotype_b:genotype_c:genotype_d
$block_id++;
@{$$linkage_blocks_hashref{$chr}{$block_id}{'markers'}} = ($pos);
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_start'} = $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_end'} = $pos;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_start'} = $marker_raw_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_end'} = $marker_raw_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_start'} = $marker_effective_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_end'} = $marker_effective_index;
$$linkage_blocks_hashref{$chr}{$block_id}{'segregation_pattern'} = $segregation_pattern;
$$linkage_blocks_hashref{$chr}{$block_id}{'genotype_pattern'} = $genotype_pattern;
my $genotype_switch_count;
# genotype switches relative to the previous linkage block
($genotype_switch_count, $$linkage_blocks_hashref{$chr}{$block_id}{'genotype_switch_pattern'}) = compare_genotype_patterns($genotype_pattern_just_saw, $genotype_pattern);
}
$genotype_pattern_just_saw = $genotype_pattern;
}
}
sub adjust_linkage_blocks {
my ($n, $l, $preliminary_linkage_blocks_hashref) = @_;
my %new_linkage_blocks = ();
foreach my $chr (sort keys %$preliminary_linkage_blocks_hashref) {
my $new_block_id = 0;
foreach my $block_id (sort {$a <=> $b} keys %{$$preliminary_linkage_blocks_hashref{$chr}}) {
my $block_marker_count = scalar @{$$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'markers'}};
my $min_block_size = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_end'} - $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_start'} + 1;
my $marker_count_by_index = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_end'} - $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_start'} + 1;
# print "checkpoint 1: chr=$chr, block_id=$block_id, block_marker_count=$block_marker_count, marker_count_by_index=$marker_count_by_index, min_block_size=$min_block_size\n";
# for debug
if (($block_marker_count >= $n) and ($min_block_size >= $l)) {
# qualified block
if ($new_block_id == 0) {
# this is the first new block on the chromosome
$new_block_id++;
%{$new_linkage_blocks{$chr}{$new_block_id}} = %{$$preliminary_linkage_blocks_hashref{$chr}{$block_id}};
} else {
# check if merging with the previous qualified block is needed
my $current_block_genotype_pattern = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'genotype_pattern'};
my $previous_new_block_genotype_pattern = $new_linkage_blocks{$chr}{$new_block_id}{'genotype_pattern'};
my ($genotype_diff_count, $genotype_switch_pattern) = compare_genotype_patterns($current_block_genotype_pattern, $previous_new_block_genotype_pattern);
if ($genotype_diff_count == 0) {
# this block has the same genotype as the previous qualified block, merging the two blocks
push @{$new_linkage_blocks{$chr}{$new_block_id}{'markers'}}, @{$$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'markers'}};
$new_linkage_blocks{$chr}{$new_block_id}{'marker_pos_end'} = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_pos_end'};
$new_linkage_blocks{$chr}{$new_block_id}{'marker_raw_index_end'} = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_raw_index_end'};
$new_linkage_blocks{$chr}{$new_block_id}{'marker_effective_index_end'} = $$preliminary_linkage_blocks_hashref{$chr}{$block_id}{'marker_effective_index_end'};
} else {
# this block has different genotype as the previous qualified block, initialize a new block
$new_block_id++;
%{$new_linkage_blocks{$chr}{$new_block_id}} = %{$$preliminary_linkage_blocks_hashref{$chr}{$block_id}};
$new_linkage_blocks{$chr}{$new_block_id}{'genotype_switch_pattern'} = $genotype_switch_pattern;
}
}
}
}
}
return %new_linkage_blocks;
}
sub check_genotype_switches {
# this function will also consider the interstitial blocks between the two input blocks if there is any
my ($block_start, $block_end, $linkage_blocks_on_chr_hashref) = @_;
my @spores = qw(a b c d);
my %genotype_switches = ();
foreach my $spore (@spores) {
$genotype_switches{$spore}{'count'} = 0;
@{$genotype_switches{$spore}{'block_boundary'}} = ();
}
for (my $i = $block_start; $i < $block_end; $i++) {
my @current_block_genotypes = split /:/, $$linkage_blocks_on_chr_hashref{$i}{'genotype_pattern'};
my @next_block_genotypes = split /:/, $$linkage_blocks_on_chr_hashref{$i + 1}{'genotype_pattern'};
for (my $j = 0; $j < scalar @spores; $j++) {
if ($current_block_genotypes[$j] ne $next_block_genotypes[$j]) {
$genotype_switches{$spores[$j]}{'count'}++;
my $next_block_id = $i + 1;
push @{$genotype_switches{$spores[$j]}{'block_boundary'}}, "$i|$next_block_id";
}
}
}
my @genotype_switches_count = ();
my %uniq_single_switches = (); # a non-redundant set of block boundaries with a single spore-specific genotype switch
my @genotype_switch_boundaries = ();
foreach my $spore (@spores) {
push @genotype_switches_count, $genotype_switches{$spore}{'count'};
my $spore_genotype_switch_boundaries = join ",", @{$genotype_switches{$spore}{'block_boundary'}};
push @genotype_switch_boundaries, $spore_genotype_switch_boundaries;
if ($genotype_switches{$spore}{'count'} == 1) {
# single switch
my $switch_boundary = ${$genotype_switches{$spore}{'block_boundary'}}[0];
if (not exists $uniq_single_switches{$switch_boundary}) {
$uniq_single_switches{$switch_boundary} = 1;
} else {
$uniq_single_switches{$switch_boundary}++;
}
}
}
my $genotype_switches_count_pattern = join ":", @genotype_switches_count;
my @genotype_switches_count_sorted = sort {$a <=> $b} @genotype_switches_count;
my $genotype_switches_count_pattern_sorted = join ":", @genotype_switches_count_sorted;
my $uniq_single_switch_count = scalar keys %uniq_single_switches;
my $genotype_switch_boundaries = join ":", @genotype_switch_boundaries;
return ($genotype_switches_count_pattern, $genotype_switches_count_pattern_sorted, $uniq_single_switch_count, $genotype_switch_boundaries);
}
sub record_recombination_event {
my ($id, $type, $subtype, $chr, $block_start, $block_end, $affected_spores, $events_hashref, $linkage_blocks_hashref) = @_;
# print "record_event: id=$id, type=$type, subtype=$subtype, chr=$chr, block_start=$block_start, block_end=$block_end, affected_spores=$affected_spores\n";
$$events_hashref{$id}{'type'} = $type;
$$events_hashref{$id}{'subtype'} = $subtype;
$$events_hashref{$id}{'chr'} = $chr;
$$events_hashref{$id}{'block_start'} = $block_start;
$$events_hashref{$id}{'block_end'} = $block_end;
if ($type eq "CO") {
# if event is CO
$$events_hashref{$id}{'marker_pos_start'} = $$linkage_blocks_hashref{$chr}{$block_start}{'marker_pos_end'};
$$events_hashref{$id}{'marker_pos_end'} = $$linkage_blocks_hashref{$chr}{$block_end}{'marker_pos_start'};
$$events_hashref{$id}{'marker_raw_index_start'} = $$linkage_blocks_hashref{$chr}{$block_start}{'marker_raw_index_end'};
$$events_hashref{$id}{'marker_raw_index_end'} = $$linkage_blocks_hashref{$chr}{$block_end}{'marker_raw_index_start'};
$$events_hashref{$id}{'marker_effective_index_start'} = $$linkage_blocks_hashref{$chr}{$block_start}{'marker_effective_index_end'};
$$events_hashref{$id}{'marker_effective_index_end'} = $$linkage_blocks_hashref{$chr}{$block_end}{'marker_effective_index_start'};
} else {
# if event is GC
$$events_hashref{$id}{'marker_pos_start'} = $$linkage_blocks_hashref{$chr}{$block_start}{'marker_pos_start'};
$$events_hashref{$id}{'marker_pos_end'} = $$linkage_blocks_hashref{$chr}{$block_end}{'marker_pos_end'};
$$events_hashref{$id}{'marker_raw_index_start'} = $$linkage_blocks_hashref{$chr}{$block_start}{'marker_raw_index_start'};