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  • Bioperl samtools: problem with reference

    Hi all

    I am having problems processing a BAM file using Bio:B::Sam.

    I want to examine variants in a BAM file using code developed from the example offered by Lincoln Stein on the CPAN Bio:B::Sam page. When I run my code (below) I get errors saying:

    Use of uninitialized value in transliteration (tr///) at myScript.pl line 22

    and also

    Use of uninitialized value $refbase in concatenation (.) or string at myScript.pl line 24

    It seems that my code is not reading or accessing the reference genome correctly, can any one point me towards a solution?

    Thanks

    Ross

    #!usr/bin/perl -w

    use Bio:B::Sam;

    use strict;

    my $bamFile="myBamFile.bam";

    my $bam = Bio:B::Sam->new(-bam => $bamFile,
    -fasta=>"MyReference.fasta",
    -autoindex => 1);

    my @targets = $bam->seq_ids;

    my @SNPs; # this will be list of SNPs
    my $snp_caller = sub {
    my ($seqid,$pos,$p) = @_;

    my $segment=$bam->segment($seqid);
    my $refbase = $bam->segment($seqid,$pos,$pos)->dna;

    $refbase=~tr/a-z/A-Z/;

    print $refbase." \n";

    };

    foreach my $chr (@targets){
    print $chr."\n";
    $bam->pileup($chr,$snp_caller);
    }

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