CRISPR clear? Dimeric Cas9-Fok1 nucleases improve genome-editing specificity.

tp://dx.doi.org/10.1016/j.gendis.2 52-3042/Copyright a 2014, Chongq Bacteria and archaea have been known for decades having evolvedadaptive immunedefenses calledclustered regularly interspaced short palindromic repeats (CRISPR)/CRISPRassociated (Cas) systems to degrade foreign nucleic acids. Recently, these RNA-guided Cas9 nucleases derived from CRISPR/Cas systems have shown promise in transforming our ability to edit mammalian genomes. While zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) have shown similar promise, the ease of producing targeting RNAs over the generation of unique sequence-directed nucleases to guide site-specific modifications makes the CRISPR/Cas9 system an appealing method for genomeediting.A short guideRNA (sgRNA)candirectCas9 to a specific genomic sequence where it induces doublestrand breaks that, when imperfectly repaired, yield mutations. Cas9 can also catalyze gene replacement through homologous recombination. Undoubtedly, Cas9-mediated genome editing and regulation should have transformative potential for basic science, genome engineering and therapeutics. However, the specificity of CRISPR/Cas9 targeting technology remains problematic. It’s been noted that sgRNA-Cas9 complexes are in general tolerant of 1w3, occasionally more, mismatches in their targets, as Cas9 was