What does the “Potential Allele Dropout” warning mean?

This warning only affects amplicon-based target enrichment.

The presence of a variant in the region where the primer anneals can affect binding affinity, and low binding affinity can affect the PCR amplification. The end result is that for this particular allele, the entire amplicon may not amplify, and hence “”allele dropout” (no longer visible in the data). Only sequences from reads from the allele without the variant would be present in the data. If there are any other variants on the same allele (as the variant causing the allele drop-out) and they are not covered by other amplicons, they will be missed.

Variants that can cause an allele drop-out will only be detected if there are other amplicons overlapping the primer region. The end primer of the last amplicon of a target region or the primers of standalone amplicons will not be covered, so the SOPHiA Platform can not provide a warning for potential drop-outs in such amplicons.

TIP: In such cases, the relative coverage for that amplicon would be lower. If CNV analysis is available for a given panel, it can confirm an allele drop-out in the same region as a deletion / undetermined status. If there is no CNV result, we recommend comparing the relative coverage of this amplicon across several samples. The majority of SNVs will likely not cause an issue, since many enrichment kit providers include common polymorphisms in their primers. In the case of INDELs, an allele drop-out is very probable, and we recommend confirming the region with another method (e.g. Sanger sequencing.)