Validation on unamplified DNA from the tissue of origin provides confirmation that a candidate call is a genuine somatic variant and rules out the possibility that it corresponds to a DNA amplification artifact or a mutation that occurred during clonal expansion. Biological validation requires a variant to be present in multiple cells in the tissue of origin at a frequency above experimental detection limits. As such, the failure to validate a variant in the tissue of origin does not necessarily represent a false call. For example, only ~50% of CNVs manifested in hiPSC clones could be directly confirmed in the primary fibroblast cells used to derive hiPSCs (80).
