Fig. 7: Mutation validation by amplicon-based sequencing.
From: MicroSEC filters sequence errors for formalin-fixed and paraffin-embedded samples
The MicroSEC analysis results were validated with amplicon-based sequencing that enriches target genomic regions by PCR. Ninety-seven mutations were randomly selected from 31 FFPE normal breast tissue samples, 12 FFPE breast tumor samples, 2 fresh frozen normal breast tissue samples, and 6 fresh frozen breast tumor samples. Each shape shows the variant allele frequencies (VAFs) in amplicon-based sequencing and capture-based sequencing for a specific mutation detected in a sample. The mutations that passed through the MicroSEC filter were detected with a similar level of VAF by both capture-based sequencing and amplicon-based sequencing (blue), with the exception of a CENPA mutation. Of the five potential CG-to-TG artifacts (red), two mutations in ESR1 or BRCA2 were not amplified by amplicon-based sequencing. Filtered out mutations were not detected by amplicon-based sequencing (green), with the exception of a NFYA mutation.
