The type II bacterial CRISPR/Cas [clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas)] system is a very valuable genome engineering tool, which has been widely used in genome editing of a variety of organisms. Previously, we generated floxed alleles in rats by CRISPR/Cas9. Here, authors successfully use a two-cut strategy with one circular vector, which contains the exogenous cDNAs with homology arm regions, in generating knockin rats at the Trdmt1, Nestin and Cck loci. The efficiency of CRISPR/Cas9-mediated knockin was up to 54%. Furthermore, by crossing the Nestin-Cre rat with the Dnmt3b floxed rat and Cck-Cre with the Dnmt1 floxed rat, they detected Cre/loxP-mediated recombination in the F1 generation of rats. They also show that the knockin alleles were germline transmitted. These results provided a simple and flexible engineering strategy for the establishment of knockin rats.
Reference:Ma, Yuanwu, et al. "Generation of eGFP and Cre knockin rats by CRISPR/Cas9." FEBS Journal 281.17 (2014): 3779-3790.
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