Germline variants

Exploring germline variants that might play a role in tumor analysis

In somatic T/N analysis, DRAGEN uses a subtraction model for SNVs. VCF will only emit variants where there is no (or very limited) alternate reads in the normal sample; in other words, variants that are found in both the normal and somatic sample will NOT be emitted in the SNV/INDEL VCF. This means that most germline variants will be excluded from the somatic VCF. To extract germline variants, it is recommended to run an additional analysis, the DRAGEN Germline pipeline, on the normal sample. In BSSH, there is an option to Enable Small Variant Calling on Normal Samples, while running Somatic in Tumor-Normal mode. As the germline and somatic variants are output from separate analyses, the variants will need to be combined before further exploration or explored separately.

Will enabling germline tagging in T/N workflows provide germline variants in the SNV/INDEV VCF?

Only those somatic variants that match a known allele from a database. In T/N mode, where the VCF usually includes only somatic variants, then it is possible that some of these somatic variants may match a known allele from a database, and then be tagged as germline.

Germline variants can be labeled in the somatic VCF by adding a flag to the command line recipe or by checking to Enable Common Germline Variant Tagging in the BSSH/ICA applications. Note that this flag has implications for other settings. First, annotation must be enabled. Second, if TMB and germline tagging are both enabled, then the TMB database filter needs to be set to false. With these two flags, germline variants are tagged in the VCF but ignored during TMB calculation.

--vc-enable-germline-tagging true
--tmb-skip-db-filter false

If I want both germline and somatic variants, should I run the Tumor sample in the Tumor/Only workflow, because that produces a VCF that contains both germline and somatic variants?

The matched normal provides a more reliable indicator of somatic variants than a database. Database germline tagging for SNV incorrectly tags ~2% of germline variants as somatic and falsely classifies some somatic variants as germline (i.e. false positives and false negatives for somatic SNVs). Additionally, there is limited support for differentiating between germline and somatic CNVs. If a paired normal sample is available, the highest performance for the detection of both somatic and germline variants is achieved by implementing the Tumor/Normal Somatic Variant Calling workflow (as described in this user guide) and also the Germline Caller on the Normal sample.