Precise Cas9 targeting enables genomic mutation prevention
Precise Cas9 targeting enables genomic mutation prevention
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DOI：
10.1073/pnas.1718148115
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发表时间：
2018-04-03
影响因子：
11.1
通讯作者：
Church, George M.
中科院分区：
文献类型：
--
作者：
Chavez, Alejandro;Pruitt, Benjamin W.;Church, George M.
在这里，我们提出了一种通用的指导RNA“调谐”方法，使Cas9能够区分两个不同的单核苷酸多态性的靶位点。我们采用我们的方法来产生体内突变预防系统，其中Cas9主动限制工程生物群体中不期望的功能获得性突变的发生。我们进一步证明了该系统可扩展到多种靶标，并且一般的调节和预防概念在工程化的Cas9变体和Cas9直系同源物中是可移植的。最后，我们表明，突变预防系统保持强大的活动，即使在复杂的环境中放置的小鼠胃肠道。
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 ourmethodology to generate an 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 mutation prevention system maintains robust activity even when placed within the complex environment of the mouse gastrointestinal tract.