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  • Consensus from mpileup for haploid sequences (forcing base calls - no ambiguities)

    I'm generating FASTQ consensus file with the samtools mpileup | bcftools view | vcfutils.pl vcf2fq chain from BAM files generated with BWA. These BAM files are alignments of Illumina reads to a mitochondrial (haploid) reference sequence.

    What I would like is a FASTQ file where the consensus base calls are made based on the most common (high quality) base in the reads at that site. That is, I don't want any ambiguity codes in my sequence. Alternatively, I'd like to be able to set a frequency threshold that a base has to exist at to be called, otherwise it is set to N.

    I have played with all of the parameters to samtools mpileup, bcftools view, and vcfutils.pl vcf2fq focusing on bcftools view's '-p', the variant probability threshold, but I can't get it to do what I want as it seems to be designed for SNP calling in diploid data and thus potentially valid heterozygous bases are called as such rather than being able to force a "homozygous" call of the most common base.

    As an example, I have a site that has 8 C's and 17 T's, all at high quality. I would like the consensus sequence of this site to have T rather than Y.

    In a perfect world, if I had a site with 8 C's and 8 T's, I'd like the option of making it either an N or calling it whatever base the reference has.

    If the mpileup -> bcftools -> vcf2fq chain won't do this, is there some other command line software that will do this from a BAM file? I need it to be command line driven because I'm doing this for 100s of BAM files as part of a batch pipeline I've written in R.

    Thanks in advance for any help.

  • #2
    I'm interested to know how to force haploid calls as well! Does anyone have any ideas how to do this?

    Thank you!

    Comment


    • #3
      Hi,
      did you ever find a way to solve this?

      Comment


      • #4
        Hello,
        I'm also interested to know if you have found a solution for SNP calls on haploid genomes.
        Thank you

        Comment


        • #5
          You may try this recent program SNVer.

          It models the number of haploids in its model so it is applicable to haplid genomes too.
          We have tried it on mitochondria for a collaborator. It works well.

          Comment


          • #6
            I still want this answer can someone please tell me how to solve this???

            Thank you

            Comment


            • #7
              For my applications, forcing an ambiguous call ended up not being necessary. However, I think the Genome Analysis Toolkit does a very nice job of genotyping and if I recall correctly it has a feature where you can set it to call haploid calls. Maybe dig into the forums over at GATK to find out more?

              Good luck!

              Comment

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