Have you looked at http://www.bioperl.org? This is not a script writing service and we don't really know much about DNA either. So, if you have a specific Perl problem, show your code, part of your input data, the output you get and the output you want.

If you want advice on the general approach, I think you will have to change your problem description to require less biologic background.

There are couple of ways to do it. If you are interested in using the Clustalw (EBI) then you have to install Bioperl module for your perl to give the multiple sequences as a set and get the output through it. The module within Bioperl package for using it is here: Clustalw. Have a look at the synopsis as it very clearly defined

The other way is by writing a Needleman Wunsch Program as a function in your perl program, providing the sequences one by one and making it compare with the ref seq sequence. Your Needleman Wunsch function is as follows:

# Needleman-Wunsch  Algorithm 

sub NeedlemanWunsch { 
my ($seq1, $seq2) = @ARGV;

# scoring scheme
my $MATCH    =  1; # +1 for letters that match
my $MISMATCH = -1; # -1 for letters that mismatch
my $GAP      = -1; # -1 for any gap

# initialization
my @matrix;
$matrix[0][0]{score}   = 0;
$matrix[0][0]{pointer} = "none";
for(my $j = 1; $j <= length($seq1); $j++) {
    $matrix[0][$j]{score}   = $GAP * $j;
    $matrix[0][$j]{pointer} = "left";
}
for (my $i = 1; $i <= length($seq2); $i++) {
    $matrix[$i][0]{score}   = $GAP * $i;
    $matrix[$i][0]{pointer} = "up";
}

# fill
for(my $i = 1; $i <= length($seq2); $i++) {
    for(my $j = 1; $j <= length($seq1); $j++) {
        my ($diagonal_score, $left_score, $up_score);

        # calculate match score
        my $letter1 = substr($seq1, $j-1, 1);
        my $letter2 = substr($seq2, $i-1, 1);                         
+   
        if ($letter1 eq $letter2) {
            $diagonal_score = $matrix[$i-1][$j-1]{score} + $MATCH;
        }
        else {
            $diagonal_score = $matrix[$i-1][$j-1]{score} + $MISMATCH;
        }

        # calculate gap scores
        $up_score   = $matrix[$i-1][$j]{score} + $GAP;
        $left_score = $matrix[$i][$j-1]{score} + $GAP;

        # choose best score
        if ($diagonal_score >= $up_score) {
            if ($diagonal_score >= $left_score) {
                $matrix[$i][$j]{score}   = $diagonal_score;
                $matrix[$i][$j]{pointer} = "diagonal";
            }
        else {
                $matrix[$i][$j]{score}   = $left_score;
                $matrix[$i][$j]{pointer} = "left";
            }
        } else {
            if ($up_score >= $left_score) {
                $matrix[$i][$j]{score}   = $up_score;
                $matrix[$i][$j]{pointer} = "up";
            }
            else {
                $matrix[$i][$j]{score}   = $left_score;
                $matrix[$i][$j]{pointer} = "left";
            }
        }
    }
}

# trace-back

my $align1 = "";
my $align2 = "";

# start at last cell of matrix
my $j = length($seq1);
my $i = length($seq2);

while (1) {
    last if $matrix[$i][$j]{pointer} eq "none"; # ends at first cell o
+f matrix

    if ($matrix[$i][$j]{pointer} eq "diagonal") {
        $align1 .= substr($seq1, $j-1, 1);
        $align2 .= substr($seq2, $i-1, 1);
        $i--;
        $j--;
    }
    elsif ($matrix[$i][$j]{pointer} eq "left") {
        $align1 .= substr($seq1, $j-1, 1);
        $align2 .= "-";
        $j--;
    }
    elsif ($matrix[$i][$j]{pointer} eq "up") {
        $align1 .= "-";
        $align2 .= substr($seq2, $i-1, 1);
        $i--;
    }    
 }
$align1 = reverse $align1;
$align2 = reverse $align2;
print "$align1\n";
print "$align2\n";
}
[download]

I would prefer that you should use the Clustalw rather than using a needleman wunsch program as it would answer your questions related to the matrices that you want to use. I hope it works for you. All the best

Thank you snape. I am aware of Needleman Wunsch Algorithm and clustalw /Muscle or other alignment programs. All these look into sequence similarities etc in generating MSA, Which I am not interested. I need to specifically align only using insert/deletions (indels) in respective sequences.

It's not clear to me what you are actually asking for help with here?

Do you already know where the insertion points are and just need help with adding the dashes in the right places?

Or are you looking for an algorithm that allows you to find the insertion points?

Also, little data and/or a worked example goes a long. If you have a set of sequences--shorted to say 200/300 chars; don't post megabytes sequences) that demonstrate what you need to do, it is usually much clearer to non-bio(*) monks.

(*)Good'ol fashioned soap was good enough for my mum...{drone}... :)

Sorry for more biological description. I have updated with very short input and output information. There are many multiple sequence alignments programs, but they all consider sequence similarity (based on physico-chemical). I mainly interested in inserting "dashes".

Are you saying you have (are given) the pairwise alignments (as strings with dashes) and all you need to produce is the "Final Multiple sequence alignment"?

Also, it there a typo?

............V
Ref1: CGACAAT--GCACGACAGAGGAAGCAGAACAGATATTTAGATTGCCTCTCATTTTCTCTCCC
Seq1: CGACAAAAAGCACGACAGAGGAAGCAGAACAGATA-----ATTGCCTCTCATTTTC-CTCCC
............^
[download]

---

Examine what is said, not who speaks -- Silence betokens consent -- Love the truth but pardon error.

"Science is about questioning the status quo. Questioning authority".

In the absence of evidence, opinion is indistinguishable from prejudice.

RIP an inspiration; A true Folk's Guy