Question: Ploidy for variant calling in tumor pooled samples for amplicon next-generation sequencing
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gravatar for Alexandre Eeckhoutte
4 months ago by
Alexandre Eeckhoutte0 wrote:

Hello everyone,

I am trying to call rare variants in pools of four tumor samples for a specific gene of interest. For that we did a amplicon next-generation sequencing on our pools. They all come tumor with a monosomy for the chromosome supporting our gene of interest.

I aligned the fastq files to the hg19 assembly using bwa mem and removed primers with BAMClipper (Au CH., 2017). I then take the union of three variant caller : Mutect2, HaplotypeCaller and Freebayes in single sample mode to call variants. A biological step of validation is made with Sanger sequencing for all potentially interesting variants.

The tools that I use take into parameter to call variants the ploidy of the pool (i.e for a pool of 8 germline DNA, the ploidy should be set to 16 - 8x2).

My question is the following : how should I adjust the ploidy parameter for pools of four monosomic tumor ?

My guess is that I should maybe increase the ploidy, and instead of setting it to 4 I should set it to 8 for instance because of the possible tumor contamination.

Does anyone have some thoughts or experience on cases like this ?

Best regards
Alexandre

sequencing next-gen gene • 192 views
ADD COMMENTlink modified 4 months ago • written 4 months ago by Alexandre Eeckhoutte0
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