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Mutect2: --force-active vs force calling --alleles
I have a ctDNA (liquid biopsy) sample that vendor reported EGFR L858R at 0.9%. When we did variant calling using varscan 2.4.4, varscan reported EGFR L858R at 1.01% but varscan fpfilter labeled EGFR L858R as false positive that failed filters VarBaseQual,MMQSdiff.
When we did variant calling using gatk 22.214.171.124 Mutect2 in tumor only mode following best practice, Mutect2 did not detect EGFR L858R although we can see the variant reads with IGV. But when we ran Mutect2 using --force-active or force calling --alleles options, Mutect2 called EGFR L858R both at ~ 0.86% vaf. To our surprise, when we ran FilterMutectCalls on the two vcf files, EGFR L858R passed filtering in both cases. Since Mutect2 did not detect EGFR L858R without --force-active or force calling --alleles options, we thought EGFR L858R would fail in FilterMutectCalls step. How to make sense of these scenarios?
This also lead us to think how should we proceed to analyze clinical samples since we do not know what variants a clinical sample has before analysis.
Thanks a lot!