A G to A (c.145G>C) mutation was engineered to change glycine codon 49 (GGC) to a serine codon (AGC) (p.G49S) in a genomic sequence fragment in the targeting vector, which also contains A loxP site flanked neomycin resistance gene cassette inserted into an intron. The targeting vector was injected into ES cells together with two gRNAs and CRISPR/Cas9 nickase mRNA encoding plasmid. The neo cassette was removed through subsequent cre-mediated recombination. The mutation mimics a polymorphism found in the human population. (J:278028)