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Problems with callers of TCGA data

We have restricted access to the TCGA data portal and we downloaded VCF files from LUAD project.

We are facing 2 problems with the files that we don't understand:

VARSCAN2 VCF files annotate normal sample always like 0/0, why is that? Do not call the normal variants?

MUTECT VCF files annotate tumor samples always like heterozygous 0/1. Why is that? Aren't there homozygous variants?

Thank you very much for your attention.

tcga callers
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updated by Kevin Blighe Kevin Blighe • written by jaumesastretomas • jaumesastretomas

1 answer

I cannot give a definitive answer, but 0/0 in VCF format just means that the variant doesn't exist in the sample. I typically see 0/0 when I merge multiple samples into a single VCF. The VCF has to list each variant 1-per-row, so, many variants will exist in one sample but not others.

Regarding the heterozygous calls, I can only imagine that a very heterogeneous tumour bulk sample was sequenced. A heterogeneous bulk sample will contain many tumour clones (thousands or millions - who knows); thus, the frequencies of the variants would be diluted based on the number of different clones. Depending on the cut-offs used by MUTECT, a homozygous variant may never be called in such a scenario. I don't fully know how MUTECT functions, but this issue appears to have been reported on the Broad Institute's forum: https://gatkforums.broadinstitute.org/gatk/discussion/6865/mutect-unable-to-detect-some-germline-mutations

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written by Kevin Blighe Kevin Blighe

I cannot give a definitive answer, but 0/0 in VCF format just means that the variant doesn't exist in the sample.

Are you sure about that? I use it to indicate that the variant does not represent the majority of either the paternal or maternal chromosome copies in a diploid, though it may still exist as a somatic variation.

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written by Brian Bushnell Brian Bushnell

...fairly sure, but you've now cast some doubt in my mind! :/

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written by Kevin Blighe Kevin Blighe

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