CRISPR/Cas12a-assisted PCR tagging of mammalian genes

Abstract

Here we describe a simple strategy for tagging of genes in mammalian cells. The method enables efficient creation of endogenously expressed protein fusions. Only PCR for the generation of a DNA fragment is required. This avoids the handling of RNAs, recombinant proteins or cloning of plasmids. The fragment, termed ‘PCR cassette’, is then transfected into cells along with a CRISPR/Cas12a helper plasmid and integrates into the target locus specified by sequences provided by the oligonucleotides used for PCR. The method is robust and works in all cell lines tested with tagging efficiency of up to 20% without selection, and up to 60% when selection markers are used.