CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, Shanghai Institutes for Biological Sciences, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai 200031, P. R. China.
Institute for Stem Cell and Regeneration, Chinese Academy of Sciences, Beijing 100101, P. R. China.
STAR Protoc. 2021 Dec 23;3(1):101062. doi: 10.1016/j.xpro.2021.101062. eCollection 2022 Mar 18.
We provide a protocol for gene editing in mouse hepatocytes using the CRISPR-Cas9 technology via AAV delivery. This protocol describes the construction of AAV plasmids, AAV packaging, injection, and the detection of knockout efficiency. Using this protocol, we can get up to 10 AAV and knock out genes in hepatocytes efficiently within 15 days. Moreover, we describe an optimized protocol to simultaneously target two genes via AAV delivery of CRISPR-Cas9 materials in the liver. For complete details on the use and execution of this profile, please refer to Wei et al. (2020).
我们提供了一种使用 CRISPR-Cas9 技术通过 AAV 递送在小鼠肝细胞中进行基因编辑的方案。本方案描述了 AAV 质粒的构建、AAV 包装、注射以及敲除效率的检测。使用此方案，我们可以在 15 天内高效获得高达 10 个 AAV 和敲除肝细胞中的基因。此外，我们还描述了一种通过 AAV 递送 CRISPR-Cas9 材料在肝脏中同时靶向两个基因的优化方案。有关此方案使用和执行的完整详细信息，请参考 Wei 等人（2020 年）。
