Two-stage PCR protocol allowing introduction of multiple mutations, deletions and insertions using QuikChange™ site-directed mutagenesis

We developed a two-stage procedure, based on the QuikChange(TM) Sire-Directed Mutagenesis Protocol, that significantly expands its application to a variety of gene modification experiments. A pre-PCR, single-primer extension stage before the standard protocol allows the efficient introduction of not only point mutation but also multiple mutations and deletions and insertions to a sequence of interest.