In that case, I'd suggest using Tie::File:

use Tie::File;

...

tie my @seq_data, 'Tie::File', $sequence_fname
    or die "Can't open $sequence_fname: $!";

if(!($permute)){
    for (@seq_data) {
        my @line = split /\t/;
        ...
    }
    ...
{
else{
    open(OUT,">>$out")||die "Cannot open $out\n";

    for (...) {
        ...
        for (@seq_data) {
            my @line = split /\t/;
            ...
        }

        foreach my $key(keys %ktc){
            ...
            print OUT ...
            ...
        }
    }

    close OUT;
}

untie @seq_data;
[download]

That'll be a little slower because you'll be repeating the split /\t/, but at least you won't have memory issues.

Also, note the change I made to die. This is not for optimising the efficiency of your code; it's to improve feedback if things go wrong. When you terminate the die message with a newline, you prevent file and line information from being output. Also, "$!" provides addition information about why open failed, see "perlvar: Error Variables". This is a good practice to get into the habit of doing; alternatively, consider using autodie.