TY - JOUR T1 - Precise Cas9 targeting enables genomic mutation prevention JF - bioRxiv DO - 10.1101/058974 SP - 058974 AU - Alejandro Chavez AU - Benjamin W Pruitt AU - Marcelle Tuttle AU - Rebecca S Shapiro AU - Ryan J Cecchi AU - Jordan Winston AU - Brian M Turczyk AU - Michael Tung AU - James J Collins AU - George M Church Y1 - 2016/01/01 UR - http://biorxiv.org/content/early/2016/06/14/058974.abstract N2 - Here we present a generalized method of guide RNA “tuning” that enables Cas9 to discriminate between two target sites that differ by a single nucleotide polymorphism. We employ our methodology to generate a novel in vivo mutation prevention system in which Cas9 actively restricts the occurrence of undesired gain-of-function mutations within a population of engineered organisms. We further demonstrate that the system is scalable to a multitude of targets and that the general tuning and prevention concepts are portable across engineered Cas9 variants and Cas9 orthologs. Finally, we show that the designed mutation prevention system maintains robust activity even when placed within the complex environment of the mouse gastrointestinal tract. ER -