Error in the creation of a consensus sequence from pileup

Question

[JW]

Hello all,

I want to create a consensus sequence from a Bam file I obtained through alignment. I have created a Vcf file as follows:

../../Code/samtools-1.2/samtools mpileup -uf ../../Reference.fasta Sorted.bam | ../../Code/bcftools-1.2/bcftools view > Pileup.vcf

The vcf file looks perfectly healthy.

Now I am trying to create a consensus Fastq sequence from it:

../../Code/bcftools-1.2/vcfutils.pl vcf2fq Pileup.vcf

But I keep getting the error:

Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 345.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 345.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 350.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 350.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 393.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 393.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 447.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 447.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 462.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 462.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 467.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 467.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 468.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 468.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 488.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 488.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 493.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 493.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 502.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 502.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 503.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 503.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 536.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 536.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 549.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 549.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 561.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 561.
Use of uninitialized value $q in numeric lt (<) at ../../Code/bcftools-1.2/vcfutils.pl line 529, <> line 789.
Use of uninitialized value $q in addition (+) at ../../Code/bcftools-1.2/vcfutils.pl line 539, <> line 789.
substr outside of string at ../../Code/bcftools-1.2/vcfutils.pl line 557, <> line 840.
Use of uninitialized value in lc at ../../Code/bcftools-1.2/vcfutils.pl line 557, <> line 840.
substr outside of string at ../../Code/bcftools-1.2/vcfutils.pl line 557, <> line 840.

What am I doing wrong?

Thanks in advance!

[swbarnes2]

I confess, I'm still on older version of samtools, but pileup format and vcf format are not the same, so when you call your file pileup.vcf, I'm concerned that you don't know quite what format you have, or want. Posting a few lines of it would help everyone to know that you do indeed have the right format.