Perl Script that should move through array with i=3 prints indicies that aren't x3

Question

I have these arrays of Sequences and I wrote this script to walk through each sequence three letters at a time (eg. {0,1,2}, {3,4,5},{6,7,8}) and print the index of where it first encounters a certian 3 letter combination (TAA,TAG,TGA). (EX. if sequence were CGTAGCCCCTAACCCC, then the script would skip over the TAG in the 2 position because its not in the correct frame of 3 and report the TAA in the 9 position). Therefore, I am only expecting indices in multiples of 3 in my results.

On most strings there is no problem, however every once in a while it will index at 4 or other non multiples of three. I was wondering if anyone more advanced than I can figure out why this may happen. I know this script is ugly and I am sorry for that, I am a biologist and I mod it for whatever I am mining out of sequences at the time. I just can't figure out the bug.

Here are some sequences from my file. The 3rd line is the sequence that gives the strange result. Just for an example of what I am dealing with.

AGGTACGCGAGTCACCTTTCGTCTTCAATCTCGTTTGATCGAAGCTATTTGTCAAAAAGAGAGGATTTTTTTGCATCTCAATTATGATCATTCCTTAGGGTTTTCAGGGTTTTGGATTGTTGTTTTTGTTAACATTTATCTGATTCGTTTGTATTTGTGTGGCAGTCTAAAGTGGCATCAACAATGGCGTCTTTTATTATACATAAGCCAAAGGAGAGATCGCCTTTCACGAAAGCTGCTTTCAAAACGGTACCTTTAGTGATTCAGCATTTTTATCTGAAATATGTTTGTTGCATTATTGAATGATTCTGATGTGGTGTTGCTACCAACTTGTCTATGTTGGTTGATTTAGCTTGATAGCATCAAGGAGTTGGAACTGTTTATGTTGAAGCATCGAAAGGATTATGTTGATCTGCACCGGACTACAGAACAGGAAAAGGATAGTATTGAACAAGAAGTAAGTACTCTGAGCTAGGCTTGCCCGTAGTATATATCTGAACTCATGAAGTTACTGCGATAAATCTATGCTTGAGTTGAGATTGAACATATGGAACTATGGAATCATAAGAAATGTAGCAACTCATATTGAGATAACTCAGGAAGATTAATGTCTATTACTTTAGATAGCGAGGGAGTTAGTATATTGTGACACTGAGGAACTTGGATCTTGTATTCTTATACCTCTTGCAGTGTTTGATCGAGAACTATGTCTACTTATGTGTTGTGTAATATCATCAAACTCTCTCTCTCTCCCTCTTGCAGGTTGCTGCTTTTATTAAAGCTTGCAAAGAACAGATCGATATTCTCATAAACAGTATTAGAAATGAAGAAGCAAACTCCAAAGGATGGCTTGGCCTCCCCGCAGATAACTTCAATGCTGATTCTATAGCACACAAACATGGAGTGGTATGATATGCACCAATGTAGTAAGCCAACTTTGGTTTTTTTTTACTATGTTTTCTTTCAAAGTATCTAGATGTGTAGAAGTAATGGTAATTTTTTTTGTATGCAGGTTTTGATTCTGAGTGAGAAACTTCATTCAGTCACTGCCCAGTTTGATCAGCTTAGAGCTACTCGTTTCCAAGATATTATAAACAGAGCTATGCCGAGAAGAAAACCTAAGAGGGTCATAAAGGAAGCTACCCCAATTAATACAACTCTGGGAAATTCGGAGTCCATAGAACCGGATGAAATCCAGGCCCAACCTCGTAGATTACAACAACAACAACTTCTAGACGATGAAACACAAGCCCTTCAGGTAACAAGGCAAATATACATGATCTTCGAAAACTTGCATAAGTTTTGTAGTTATGCTAAATTTTGAAATTGATAATTTTTGCAGGTAGAGCTAAGTAATCTTTTAGATGGTGCTAGGCAGACAGAAACTAAGATGGTGGAGATGTCTGCATTAAACCACTTGATGGCAACTCATGTTCTGCAGCAAGCCCAACAGATAGAGTTTCTTTATGACCAGGTTAGGACTTATTAACTTCTCTAACGCTCTCATGTCAACACACTGTTTTGTTAGGCTTTCACTGTTCTTTACACTCCTTTGCTATCTCAAAGTTAAATTCGGATGCTTATTGTATTCAGAACTTTTCCTTGTCACATTCACCTAAATTAGGTATAGAGACGGGAAAGAAACTTTGTATTGGTCCAATTTTAATTGCTCTCCAATTTAGTGGTAGGAAATGGAACGGTTAATGTTTTTAGCTATGTAAAGTCTCTAAAACTCCATTTGAATGTGTCAATGACTCAATGCCATTCCCAATACTTTAGTTTATGGGGCTTTGCAGTTTTCCTACTCTGTAAACGTACAGCTTATGACTGACTTGGTGGCTCTCTTTATGTGTGTGTGTGTGTGTCTTGAGGCCCTTTTTCTCACTCAGTTTGACACTAAATGCAGGCAGTTGAGGCAACAAAGAACGTGGAGCTTGGAAACAAAGAGCTTTCTCAAGCAATCCAACGAAACAGCAGCAGCAGAACCTTTCTCTTACTGTTTTTCTTCGTCCTTACTTTCTCCGTCTTGTTCTTGGATTGGTACAGTTAAaaaacc 
AGGTGATTGTTTTGTTATTATAAATCAAGATCAGTACATATATATTTTTGTTTTTCTTGGTTTCATATGTAATATTTTGGACTTTTGGTGTTTAGGTTTTTGACTTGGAAGAAAAGAACGTAATGGATGAGTCACTACACGAGGTGTATAAATTTTGCCTCACCGATGTTGATGAGAGAAGCAAGAAAGAGACATCAATGAAAGATGATTACATAGAACATAAGAAGTCTACTAGATTGTTGGCTGAAAATGCGAAGAAGTCCGGTCACAGTTTAGAAATATTAAGGCCGGAATCTAAACCTGAGACTGAAAAAGAGGTGATTTTATTTTCTTGTTATATAAAGATTCGTAGACATATATTTGGTTTTTCTTTGGTTTCATAATATTTTGGACTTATGTGTGTTTAGGTCAATGAAGAGGAAGAGAAGAGAGTAATGGATCCGGATGTGGATATTAGTTGTTATGAAGAGTCACCACACGAGGTGTATAAATTTAGCCTCACCGATTTCGAAGAAGAGATAATGGAAGATGATTACAGAGAAGATATGAAGTGTAGAATGTTGGATGATATAGTGAAGAATTCCGGTCACCGTGTAGAAATATCAAGGCCGGAATATTATAAACCTGAGATTGAAAAACAGGTTTTATTTTTTTGGTTATTTTGTGATTAAGATCAGTTTTTTTTTTTTTTTTTTTTGGTTTAATAATATTTGATCTTGTGTGTGTTTAGGTATATGAAAAGGAAGAGAAGAAAGTAATGGATCCGGATATCTATATTAGATCTTATGAAGAGTCACCAAACGAGGTGTATAAATTTAGCCTCACTGATTTGGAAGAAGAGATAATGGAAAATGACTCCATAGAAGGTGTGAAGTGTAGAATGTTGGATGAAATAATGAAGAAGTCCGGTCACCATTTAAAAATATCAAGGCCGGAATATAAACCTGAGATTGAAAAACAGGTTAGTTTTTAATAAAAAGATCACTAGATATTTTTTTTTATTTTTTTTTGTTTTTGGTTTCATAATATTTGACTTGTGGCATGTGTTTAGGTATATGAAGAGGAAGAGAAGAAAGTAATGGATCCAGATGTGGATATTAGATGTTATGAAGAGTCACCACACGAGGTGTCTAAATTTAGCCTCACCGATTTCGAAGAAGAGATAATGGAAGATGATTACATAGAAGCTTTGAAGTGTAGAATGTTGGATGATATATTGAAGAAGTCCGGTCACCGTTTAGAAATATCAAGGCGGCAATATAATAAACCTGAGATTGAAATACAGGTGATTTTTTTTTTTTATTATTGTTGTTATAGTAAGATCAGTAGATATATATCTTGGTTTCATAATATTTTGGACTTGTGTGTGTTTAGGTCAATGAAAAGGAAGAGAAGAAAGTAATCAATACGGATATGGATATTAGATATGATGATGAGTCACCAGAAGAGGTGGAGACATATTCTAGTCTCACGGATGATGAAGAAGAGAGAAGCAAGGAAGATACATCAATGGAAGATGTGAAGTGTAGAATGTTGGATTAAAAAACGACGAAGCTCGGCCACCTTTTAGGAATATCAAGGCCGGAATATAGACCTGAGATTGAAAAACAGGTGATTTTATTTTGTTGTTAATTGTATTAGTAAAGATCAGTAGATATATATTTGTTTTTGTTTTTCGGTTTCATAATATTTTGGACGCTTGTGTTTAGGTCAATGAAGAGAAAGAAAGAAAGTAATGGATATTAGATCTGCTGGTCAGTCACAAACACGAGGTGTACAAATTTAGCCTCACCGATATCAAAGAAGAGAGAAGCAATGAAGATACATCAATGGAAGATTGTTGCATAGAAGAGGCTCAAGTCGGAAAAGATCAAAGAGTCTTCAGATTCAGAGAAAGTAGTGAAGAGAAGAGAAAATCCTCATCATCACCATTATCACCACTAACAGAGTTTAGGGATATGGAGAGTTTGACGTATTACATGAGGCAAAAAGGGATGCATCGAAGAAGAAGAAGATCATCAACATCACCACATTGTTGCCATAATGTAGTATACAATGAGTTTAAAGTGACGAAGGAAGAAGAAGAGGAAGAAAGACAAAGATTAACAACCAAACGTGTTCATTCTAAGCTTCATGAATACGAACAATTTTTAACTCAGTTTAAAAAGAAGAAGGAAGAAGAAAACGAGAGACGAAGATTATCACCCAAAGACTTTGAGCCTACGCTTCCTGATTACGACCAAGTGATTACTCGCTTTAGAGTGCTGGAGAAGGAAGAAGAAGAAAGACGAAGATTAGCAACAAAACATGTTCATCCTAAGCTTCCTGATTACGACCAGATTGCTACTAAGTTTAAACTCCTGAAGGAGGTAGAAAAAGAAAGACGAAGATTATTAACCAAACACAGTTCATCCTAAgcttcc 
TGGTAATTTTTGCATCTTCAAAATGTTCTAAAATTTTGGCAAATGGTTTTGTTAAGTTCGAATTTTTGGTTATGATACAGTTTGAACGTTTTTCTTCATAGATTACAGTTTTAGCAAATGTGAATCATTAAAAGTGGAATAGTTGGTTTGAAAACAATTGTCAATTTCATTTTTTTTTTGGTTTTATGGTTAGGCGAGGAAAGCATTAAGAGCTTTGAAAGGTATAGTGAAGCTACAAGCATTAGTGAGAGGATACTTAGTAAGGAAACGCGCGGCCGCAATGTTGCAGAGCATACAAACTTTGATCAGAGTCCAAACCGCTATGCGATCAAAACGCATCAATCGCAGCCTCAACAAAGAGTACAACAACATGTTTCAACCTCGACAATCCTTTGTAAAGAACTATTCTCATTTCCATTGGCTCTCTTTTTTTCTTTAAGCCAAAACAAGACTTAAAGTGTGTCCTCTGTTTGTAGGATAAGTTTGATGAAGCAACGTTCGATGACAGAAGAACAAAGATTGTAGAGAAGGACGATAGATACATGAGAAGATCAAGTTCAAGATCAAGATCTAGACAAGTGCACAATGTTGTTTCAATGTCTGACTATGAAGGCGATTTTGTTTACAAAGGGAATGATTTGGAGTTGTGTTTCTCGGATGAGAAGTGGAAGTTTGCTACCGCGCAGAACACGCCGAGATTATTGCATCACCATTCTGCTAATAATCGCTATTATGTAATGCAGTCTCCAGCTAAGAGTGTTGGTGGAAAGGCTTTGTGTGACTATGAAAGCAGTGTGAGTACTCCTGGCTACATGGAGAAAACTAAGTCCTTTAAGGCAAAAGTGCGTTCACACAGCGCACCGCGCCAGCGATCTGAGAGGCAGAGGTTGTCGCTAGATGAAGTTATGGCCTCTAAGAGTAGCGTTAGCGGTGTGAGTATGTCGCATCAGCATCCACCACGCCATTCTTGTTCCTGTGATCCGCTTTAActtaac 
GAGTTAGTAAACAAAGTGTTCACATTTTAGTAAACATTGTTGTTCGTTAATCACGTAACGTTTTGTTTTTCCAGTTTACACTGAGCTCTGATGAGTATATAACGGAGGTGAATGGTTACTACAAAACTACGTTTTCGGGAGAAGTCATAACGTCGTTGACGTTCAAGACGAACAAAAGGACATATGGGACTTACGGAAATAAAACCAGTAGCTACTTTTCTGTTGCCGCACCCAAAGATAACCAGATTGTCGGTTTTCTTGGAAGTAGCAGCCATGCTCTCAACTCCATCGACGCTCATTTTGCCCCTGCTCCTCCTCCTGGTAGCACCGGAGCTAAGCCCGGTGCTAGTGGCATCGGAAGTGATTCTGGTAGCATTGGTAGTGCCGGAACTAACCCTGGTGCTGATGGCACCAGAGAAACCGAAAAAAACGCTGGTGGCTCAAAACCTAGTAGTGGTAGTGCCGGAACTAACCCTGGTGCTAGTGCTGTTGGCAACGGAGAAACCGAAAAAAATGCTGGTGGCTCAAAACCTAGCAGTGGTAGTGCTGGAACTAACCCTGGTGCTAGTGCTGGTGGCAACGGAGAAACCGAAAAAAACGTTGGTGGCTCAAAACCTAGCAGTGGTAAAGCCGGAACTAACCCTGGTGCTAATGCTGGTGGCAACGGAGGAACCGAAAAAAACGCTGGTGGCTCAAAATCTAGCAGTGGTAGTGCTCGAACTAACCCTGGTGCTAGTGCTGGTGGCAACGGAGAAACTGTTTCCAACATTGGAGATACGGAAAGTAACGCTGGTGGCTCGAAAAGTAATGATGGTGCTAACAATGGTGCTAGTGGCATTGAAAGTAATGCTGGTAGCACTGGAACTAACTTTGGTGCTGGTGGCACCGGGGGAATTGGAGATACGGAAAGTGATGCTGGTGGCTCCAAAACTAACTCTGGAAACGGCGGAACTAACGATGGTGCTAGTGGTATTGGAAGTAATGATGGTAGCACTGGAACTAACCCTGGTGCTGGTGGAGGAACAGATTCAAACATCGAAGGTACTGAAAATAACGTTGGTGGCAAGGAAACTAACCCTGGTGCTAGTGGCATTGGAAATAGTGATGGTAGCACTGGAACTAGCCCCGAAGGTACCGAAAGTAACGCTGACGGCACAAAAACTAACACGGGAGGCAAAGAATCTAACACCGGAAGTGAATCCAACACCAATTCTAGTCCACAAAAGTTGGAAGCACAAGGAGGCAATGGAGGAAATCAATGGGACGACGGAACCGATCATGATGGTGTGATGAAGATACATGTTGCAGTTGGTGGTCTAGGAATTGAGCAAATTAGATTTGATTATGTCAAGAACGGACAGTTGAAGGAAGGACCCTTCCACGGTGTCAAAGGAAGAGGTGGCACTTCAACGGTGCGTAAATTTTTATTATTATGGCTCAATTACGTTTTTCGAATAAGTGTTAATTCAAGATTATTGATCTTCATGATTCTGCAGATTGAGATTAGCCATCCGGACGAGTATCTTGTTTCCGTCGAGGGGTTGTACGACTCTTCCAATATCATTCAAGGAATCCAGTTTCAATCCAACAAACACACTTCTCAGTACTTTGGATATGAATATTATGGAGATGGTACACAATTTTCACTTCAAGTTAATGAAAAGAAGATCATTGGTTTCCATGGTTTTGCCGACTCACACCTTAATTCTCTTGGAGCTTATTTCGTTCCAATCTCATCCTCTTCTTCCTCCTTGACTCCTCCTCCCAACAAAGTTAAAGCTCAAGGAGGAAGTTATGGAGAAACATTTGACGATGGTGCTTTCGATCATGTAAGAAAGGTTTATGTTGGTCAAGGTGATTCTGGTGTAGCTTATGTCAAGTTCGATTATGAAAAAGACGGTAAAAAGGAGACACAAGAACATGGAAAAATGACATTGTCAGGAACAGAGGAGTTTGAGGTTGATTCAGACGATTACATAACATCAATGGAGGTTTATGTCGACAAAGTCTACGGTTATAAAAGCGAAATCGTCATTGCTCTTACCTTCAAGACCTTTAAGGGTGAAACTTCTCCACGTTTTGGAATAGAGACTGAGAATAAATATGAAGTTAAAGACGGTAAAGGAGGAAAACTTGCTGGTTTCCATGGAAAAGCTAGCGATGTTCTTTATGCTATTGGTGCTTATTTCATTCCAGCAGCAAATTAGagagtt 
ACGTATGTCTTAGTTACTACTATCATACTATATTACTATGTATTGGAAAACTTTTGGTTAGAACCTGTTGGGAGGAAAGGGTTTATGTTCTGGTTCATTTTACGTGTACTAAGTACTTATAATTAAGATTAAAAGAAACATTTACAGCTTCACCCTCTGGTCGATGTATGTGGGCTGTGGGCATGTGGCCAATCTCTGAAGCGTTAGGTAGAGCAAATATAGAGTTGAGAGTTGCTTAAGTTAGTGAACGTGAATGACTAAAAAGATATGTTGCATTTAAATCGTATTGGGCCTCATCCCATCTAAAATATAGTAGGTGTAGGCCTTTTAGGTTAATTTGAATAAAATCAACCTTTTTGTAAGCAACATCGACGATTGTCACATTTTTCTCATACACATAGGTGTAATCTAGCTTTGAATGTTTTCTCATACACATAGGTGTAATCACCGTAATTATCATTTGTGAAGATATATGTTTTACCAAGTGGTTTGTATTGTCCATATATACTTTACCACTTTCATATTAACATATAATGTTTTTGTAAGTATTATACCATAAAGGATTGGTTTCTTAATATTATTAACAAAACGCAAAAATTCTTTTAAACGCAGGCGATTCCAATCCACAGCGTTGCGGTTAGAGTAGGATCAACACAAAGAGTAGTGATGGAGATCATAATCACATTCGCATTGGTCTACACTGTTTACGCCACAGCCATTGACTCCAACAATGGCACTCTCGGAACCATCGCTCCACTTGCTATCAGACTCATCGTTGGTGCTAACATTCTTGCAGCCGGCCCATTCTCTGGTGGTCCAATGAACCCTGGACGTTCTTTTGGATCATCTCTTGCCGTTGGAAATTTTTCAGGACATTAGgtttat 

and here is the script I am running:

#!/usr/bin/perl

use strict; 
use warnings;

# A program to find the first inframe stop codon of non-spliced intron containing genes
print "ENTER THE FILENAME FOR DNA SEQUENCES:= ";
# Asks for Sequence file and if file does not exist prints error message
my $filename = <STDIN>;
#my $sequence;
my @sequence;
chomp $filename;
unless (open(DNAFILE, $filename) ) {
    print "Cannot open file \"$filename\"\n\n";
}
@sequence = <DNAFILE>;
close DNAFILE;
open (FILE, ">AtPTCindex.txt");
my $j;
my $i;
my $codon;
my $stopseq;
my $counter;
#Change $j<(375) to n=number of sequences
for ($j = 0; $j < @sequence; $j ++) {
    $counter = 0;
    for ($i = 0; $i < (length($sequence[$j]) - 2) && $counter < 1; $i += 3) {
        $codon = substr($sequence[$j], $i, 3);
        if ($codon =~ m/TAG|TGA|TAA/g) {
            # m added before /TAG... above
            $stopseq = substr($sequence[$j], $i, 9);
            my $result = index($sequence[$j], $stopseq);
            $counter = 1;
            #my $results = index($sequence[$j], $stopseq);
            print FILE "$result \n"; 
            #print FILE "$results $j \n";
        }
    }
    if ($counter == 0) {
        print FILE "\n"
    }
}
close FILE;
exit;

Thanks so much.

Answer 1

As threatened, the following is a cleaned up version of your script:

#!/usr/bin/perl

use strict; 
use warnings;
use autodie;

die "Usage: $0 Filename\n" if @ARGV != 1;

my $file = shift;

open my $infh, '<', $file;
open my $outfh, '>', 'AtPTCindex.txt';

while (my $line = <$infh>) {
    chomp($line);

    my $result = '';

    for (my $i = 0; $i < (length($line) - 2); $i += 3) {
        my $codon = substr($line, $i, 3);
        if ($codon =~ m/TAG|TGA|TAA/) {
            # m added before /TAG... above
            my $stopseq = substr($line, $i, 9);
            $result = index($line, $stopseq);
            $result .= " ($i, $codon, $stopseq)";
            last;
        }
    }

    print "$result\n";
#   print $outfh "$result\n";
#   print $outfh "$result $.\n";
}

close $infh;
close $outfh;

For the 5 lines of data that you provided, the following is the output:

84 (84, TGA, TGATCATTC)
3 (3, TGA, TGATTGTTT)
3 (3, TAA, TAATTTTTG)
4 (27, TAG, TAGTAAACA)
123 (123, TAA, TAAGATTAA)

I believe your issue is with these lines:

my $stopseq = substr($line, $i, 9);
$result = index($line, $stopseq);

You're pulling a sequence from the $line at position $i, and then immediately doing an index for it. In the case of 4 of 5 of those lines, it immediately finds the same value $i. However, in the case of line 4, it finds a matching sequence earlier in the line.

If this isn't desired, you'll have to explain what your desired behavior actually is. Perhaps, you just want $i? Or are you looking for a matching stop sequence any point AFTER $i? You'll have to specify what your actual logic wants to be.

Answer 2

I took a different approach, unpacking it into groups of three instead of counting by indexes of three. I believe this script does what you want, and it looks a lot cleaner. It can also optionally take the filename as argument.

#!/usr/bin/perl

use strict;
use warnings;

my $filename = 'a'; # dummy value
my $resultfile = 'AtPTCindex.txt';

# User may have passed filename as arguement
if (@ARGV) {    if (-e $ARGV[0]) { $filename = $ARGV[0] } }
unless (-e $filename)
{
        print "ENTER THE FILENAME FOR DNA SEQUENCES: ";
        chomp($filename = <STDIN>)
}

open DNA,"<$filename" or die "Couldn't open $filename for reading: $!\n";
my @sequence = <DNA> or die "Couldn't read $filename: $!\n";;
close DNA;

# Uncomment the below line if you're braver than me
if (-e $resultfile) { die "Cowardly refusing to write to existing file" }
if (-e $resultfile) { unlink $resultfile };

open RESULT,">>$resultfile" or die "Courdn't open$!\n";
foreach my $string (@sequence)
{
        # split into groups of 3
        my @groups = unpack "(A3)*", $string;

        # Search for the group you want
        for (my $groupnum = 0; $groupnum < @groups - 1; $groupnum++)
        {
                if ($groups[$groupnum] =~ m/(TAG|TGA|TAA)/g)
                {
                        print RESULT (($groupnum + 0) * 3) . "\n";
                        print "$1 (" . $1 . ( $groups[$groupnum + 1]) . ($groups[$groupnum + 2]) . ") at index " . (($groupnum + 0) * 3) . "\n";
                        last;
                }
        }
}
close RESULT;

Running the script on your sample data, it outputs:

TGA (TGATCATTC) at index 84
TGA (TGATTGTTT) at index 3
TAA (TAATTTTTG) at index 3
TAG (TAGTAAACA) at index 27
TAA (TAAGATTAA) at index 123

...as well as writes the raw index numbers to the file specified.