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ratio_calc.pl
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#!/usr/bin/perl
#############################################
#This program is a part of the SLOFE and takes inputs from it.
#This program indentifies the SRPS operons and calculates the ratio using the predicted stable stem-loops.
#
#Author: Yogendra Bhaskar
#
#License: GPL 3.0
#
#############################################
use strict;
use warnings;
use File::Basename;
use Cwd;
use Cwd 'abs_path';
use List::Util qw( min max ); #which is released with the Perl distribution v5.7.3
my $dirname_abs = abs_path(dirname(__FILE__));
my $Operon= $ARGV[0];
my $GB = $ARGV[1];
my $dir_out= $ARGV[2];
my $tmp_dir="$dir_out/tmp_slofe";
system_commands($Operon, $GB, $dir_out);
open(fh2,"$tmp_dir/srps_operon_genes_new");
#print "file opened for reading \n";
my @read2=<fh2>;
my $count_read2=0;
foreach (@read2) {
chomp;
$count_read2++;
}
my @OP2; my @strand2; my @op_count2; my @gene2; my @g_start2; my @g_end2;
for(my $i=0; $i<$count_read2; $i++){
my @values2= split("\t", $read2[$i]);
$OP2[$i] = $values2[0];
$strand2[$i]= $values2[1];
$op_count2[$i]= $values2[2];
$gene2[$i]= $values2[3];
$g_start2[$i]= $values2[4];
$g_end2[$i]= $values2[5];
}
close(fh2);
open(fh3,"$tmp_dir/final_stem-loops_improved_T_intergenic");
#print "file opened for reading \n";
my @read3=<fh3>;
my $count_read3=0;
foreach (@read3) {
chomp;
$count_read3++;
}
my @g_start3; my @g_end3; my @g_strand3; my @gene_new3; my @strand3; my @start3; my @end3; my @region3; my @length3; my @rm_seq3; my @rf_seq3; my @energy3; my @folding3; my @OP3; my @OP_region3; my @U_c3; my @PolyU3;
for(my $i=0; $i<$count_read3; $i++){
my @values3= split("\t", $read3[$i]);
$g_start3[$i]= $values3[0];
$g_end3[$i]= $values3[1];
$g_strand3[$i]= $values3[2];
$gene_new3[$i]= $values3[3];
$strand3[$i]= $values3[4];
$start3[$i]= $values3[5];
$end3[$i]= $values3[6];
$region3[$i]= $values3[7];
$length3[$i]= $values3[8];
$rm_seq3[$i]= $values3[9];
$rf_seq3[$i]= $values3[10];
$energy3[$i]= $values3[11];
$folding3[$i]= $values3[12];
$OP3[$i]= $values3[16];
$OP_region3[$i]= $values3[17];
$U_c3[$i]= $values3[18];
$PolyU3[$i]= $values3[19];
}
close(fh3);
no warnings 'uninitialized';
my @eng_new;
my @reg_new;
for(my $i=0; $i<$count_read2; $i++){
for(my $j=0; $j<$count_read3; $j++){
if($strand2[$i] eq '-'){
if($OP2[$i] == $OP3[$j] && $gene2[$i] eq $gene_new3[$j] && $OP2[$i] == $OP2[$i+1]){
$eng_new[$i+1]=$energy3[$j];
$reg_new[$i+1]=$region3[$j];
# print "$OP2[$i]\t$strand2[$i]\t$op_count2[$i]\t$gene2[$i]\t$g_start2[$i]\t$g_end2[$i]\t$eng_new[$i]\tmid\n";
}
elsif($OP2[$i] != $OP2[$i-1] && $OP3[$j] == ($OP2[$i] -1) && $OP_region3[$j] =~ m/$OP2[$i]/){
$eng_new[$i]=$energy3[$j];
$reg_new[$i]=$region3[$j];
# print "$OP2[$i]\t$strand2[$i]\t$op_count2[$i]\t$gene2[$i]\t$g_start2[$i]\t$g_end2[$i]\t$eng_new[$i]\tend\n";
}
}
elsif($strand2[$i] eq '+'){
if($OP2[$i] == $OP3[$j] && $gene2[$i] eq $gene_new3[$j]){
$eng_new[$i]=$energy3[$j];
$reg_new[$i]=$region3[$j];
}
}
}
}
open (wrfile1, ">$tmp_dir/with_eng_redef");
for(my $i=0; $i<$count_read2; $i++){
if($reg_new[$i] eq "" && $eng_new[$i] eq ""){
print wrfile1 "$OP2[$i]\t$strand2[$i]\t$op_count2[$i]\t$gene2[$i]\t$g_start2[$i]\t$g_end2[$i]\t0\t0\n";
}
else{
print wrfile1 "$OP2[$i]\t$strand2[$i]\t$op_count2[$i]\t$gene2[$i]\t$g_start2[$i]\t$g_end2[$i]\t$reg_new[$i]\t$eng_new[$i]\n";
}
}
close(wrfile1);
open(fh1,"$tmp_dir/with_eng_redef");
#print "file opened for reading \n";
my @read1=<fh1>;
my $count_read1=0;
foreach (@read1) {
chomp;
$count_read1++;
}
my @values1;
my (@OP, @strand,@op_count,@gene,@g_start,@g_end,@region,@eng);
for(my $i=0; $i<$count_read1; $i++){
@values1= split("\t", $read1[$i]);
$OP[$i] = $values1[0];
$strand[$i]= $values1[1];
$op_count[$i]= $values1[2];
$gene[$i]= $values1[3];
$g_start[$i]= $values1[4];
$g_end[$i]= $values1[5];
$region[$i]= $values1[6];
$eng[$i]= $values1[7];
# print "$OP[$i]\t$strand[$i]\t$op_count[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\n";
}
#Ratio calculation
my $a=1 ;
until($a>int((0.8*max(@op_count))+0.99)){
for(my $i=0; $i<$count_read1; $i++){
if($strand[$i] eq '+' && $OP[$i]==$OP[$i+1]){
if($eng[$i]==0){
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i+1]\tnewP\n";
$eng[$i]=$eng[$i+1];
}
else{
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\torgP\n";
$eng[$i]=$eng[$i];
}
}
elsif($strand[$i] eq '-' && $OP[$i]==$OP[$i-1]){
if($eng[$i]==0){
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i-1]\tnewN\n";
$eng[$i]=$eng[$i-1];
}
else{
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\torgN\n";
$eng[$i]=$eng[$i];
}
}
else{
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\tNEOP\n";
$eng[$i]=$eng[$i];
}
}
$a=$a+1;
}
my @eng2=0;
for(my $i=0; $i<$count_read1; $i++){
if($strand[$i] eq '+' && $OP[$i]==$OP[$i+1]){
if($strand[$i] eq '+' && $OP[$i]!=$OP[$i-1]){
$eng2[$i]=$eng[$i];
}
# $eng2[$i]=$eng[$i];
$eng2[$i+1]=$eng2[$i];
# $eng2[$i]= $eng2[$i+1];
}
# elsif($strand[$i] eq '-' && $OP[$i]==$OP[$i-1]){
# $eng2[$i]=$eng[$i];
# $eng2[$i-1]=$eng[$i];
# }
if($OP[$i]!=$OP[$i-1] && $OP[$i]!=$OP[$i+1]){
$eng[$i]=1;
$eng2[$i]=1;
}
}
for(my $i=$count_read1; $i>0; $i--){
if($strand[$i] eq '-' && $OP[$i]==$OP[$i-1]){
if($strand[$i] eq '-' && $OP[$i]!=$OP[$i+1]){
$eng2[$i]=$eng[$i];
}
$eng2[$i-1]=$eng2[$i];
}
}
# Ratio normalization
my @ratio=0;
for(my $i=0; $i<$count_read1; $i++){
$ratio[$i]= sprintf("%.2f", abs($eng[$i]/$eng2[$i]));
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\t$eng2[$i]\t$ratio[$i]\n";
}
#Ratio representation in this form: 1:0:0:0
for(my $i=0; $i<$count_read1; $i++){
if($OP[$i]==$OP[$i-1]){
$OP[$i]= $OP[$i];
$strand[$i]= $strand[$i];
$gene[$i]= "$gene[$i-1],$gene[$i]";
$g_start[$i]=$g_start[$i-1];
$g_end[$i]=$g_end[$i];
$region[$i]=$region[$i];
$eng[$i]="$eng[$i-1]:$eng[$i]";
$eng2[$i]="$eng2[$i-1]:$eng2[$i]";
$ratio[$i]="$ratio[$i-1]:$ratio[$i]";
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\t$eng2[$i]\t$ratio[$i]\n";
}
}
open (RatioCalc, ">$dir_out/Stoichiometry_ratio.csv");
my $b=0; my $wc=0;
print RatioCalc "#Operon\tStrand\tGenes\tOperon_start\tOperon_end\tdeltG_ratio\tRatio\n";
for(my $i=0; $i<$count_read1; $i++){
if($OP[$i+1]==$OP[$i]){
$b=1;
}
else{
# print "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$region[$i]\t$eng[$i]\t$eng2[$i]\t$ratio[$i]\n";
print RatioCalc "$OP[$i]\t$strand[$i]\t$gene[$i]\t$g_start[$i]\t$g_end[$i]\t$eng[$i]\t$ratio[$i]\n";
$wc =$wc+1;
}
}
close(RatioCalc);
use warnings 'uninitialized';
#my $stoi_ratio= `sed '1d' Stoichiometry_ratio.csv| wc -l`;
#print "Number of SRPS operons and their stoichiometry in the from of ratio: $stoi_ratio Stoichiometry_ratio.csv";
print "Number of SRPS operons and their stoichiometry in the from of ratio: $wc $dir_out/Stoichiometry_ratio.csv\n";
print "Done!\n";
#system('mv '.$tmp_dir.'/rnamotif.out '.$tmp_dir.'/sequence.out '.$tmp_dir.'/rnafold.out '.$tmp_dir.'/cutoff5_stem-loops '.$tmp_dir.'/non-redundant_stem-loops '.$tmp_dir.'/final_stem-loops_sorted '.$tmp_dir.'/T_ext '.$dir_out.'');
#system('rm -rf '.$tmp_dir.'/*');
#system('mv ../rnamotif.out ../rnafold.out ../rnaM_rnaF_merged ../final_stem-loops_improved_T_intergenic ../with_eng_redef '.$tmp_dir.'/');
#system('mv '.$dir_out.'/rnamotif.out '.$dir_out.'/sequence.out '.$dir_out.'/rnafold.out '.$dir_out.'/cutoff5_stem-loops '.$dir_out.'/non-redundant_stem-loops '.$dir_out.'/final_stem-loops_sorted '.$dir_out.'/T_ext '.$tmp_dir.'/');
my $SSL; my $STSL;
sub system_commands{
system('cut -f2 '.$Operon.' |sed \'1i\\\\\'|awk \'{FS=","}{print NF}\'|sed \'1d\' > '.$tmp_dir.'/operon_count');
system('paste '.$Operon.' '.$tmp_dir.'/operon_count > '.$tmp_dir.'/improved_operon_map1');
system('grep -v Poly5U '.$dir_out.'/intergenic_intraOperonic_stem-loops |grep -v Poly4U|cut -f17|sort -g -u > '.$tmp_dir.'/tmp_srps_uniqOP');
#Stabilizer_stem-loops-SSL
system('grep -v Poly5U '.$dir_out.'/intergenic_intraOperonic_stem-loops |grep -v Poly4U|awk \'{FS="\t"}{if($19<=5)print $0}\' > '.$dir_out.'/Stabilizer_stem-loops-SSL');
$SSL= `wc -l $dir_out/Stabilizer_stem-loops-SSL`;
print "Number of predicted SSL: $SSL";
#Stabilizer_and_terminator_stem-loops-STSL
system('grep -v Poly5U '.$dir_out.'/intergenic_intraOperonic_stem-loops |grep -v Poly4U|awk \'{FS="\t"}{if($19>5)print $0}\' > '.$dir_out.'/Stabilizer_and_terminator_stem-loops-STSL');
$STSL= `wc -l $dir_out/Stabilizer_and_terminator_stem-loops-STSL`;
print "Number of predicted STSL: $STSL";
system('awk \'{FS="\t"}NR==FNR{h[$1]=$2"\t"$4; next}{print $0"\t"h[$1]}\' '.$tmp_dir.'/improved_operon_map1 '.$tmp_dir.'/tmp_srps_uniqOP| cut -f2|tr \',\' \'\n\' > '.$tmp_dir.'/tmp_all_srpsGene');
system('awk \'{FS="\t"}NR==FNR{h[$0]=1; next}{for(i in h) if($0~i){print $0"\t"i}}\' '.$tmp_dir.'/tmp_all_srpsGene '.$tmp_dir.'/improved_operon_map1|cut -f1,3,6,7 > '.$tmp_dir.'/genes_with_operon_number_new');
system('awk \'{FS="\t"}NR==FNR{h[$3]=$1"\t"$2; next}{print $0"\t"h[$4]}\' '.$GB.' '.$tmp_dir.'/genes_with_operon_number_new| sort -k4|sed \'/\t1\t/d\' > '.$tmp_dir.'/srps_operon_genes_new');
system('grep -v inside '.$dir_out.'/stable_stem-loops > '.$tmp_dir.'/final_stem-loops_improved_T_intergenic');
}
__END__