Sanger sequencing is no longer the gold standard, ... because there is potential for allele dropout due to polymorphic positions under primers or unknown heterozygous deletions. When this happens, the sequencing may either miss variants or may erroneously assign homozygosity to a heterozygous/hemizygous variant. What’s more, Sanger sequencing can only detect a minimum allele frequency of 15 percent to 20 percent.
says Josh Deignan, PhD, associate director of the UCLA molecular diagnostics laboratories.
Do you use Sanger confirmation in your lab? In which cases is it acceptable to skip Sanger sequencing?