文章基本信息

标题：Validation of microinjection methods for generating knockout mice by CRISPR/Cas-mediated genome engineering
本地全文：下载
作者：Takuro Horii ; Yuji Arai ; Miho Yamazaki 等
期刊名称：Scientific Reports
电子版ISSN：2045-2322
出版年度：2014
卷号：4
DOI：10.1038/srep04513
出版社：Springer Nature
摘要：The CRISPR/Cas system, in which the Cas9 endonuclease and a guide RNA complementary to the target are sufficient for RNA-guided cleavage of the target DNA, is a powerful new approach recently developed for targeted gene disruption in various animal models. However, there is little verification of microinjection methods for generating knockout mice using this approach. Here, we report the verification of microinjection methods of the CRISPR/Cas system. We compared three methods for injection: (1) injection of DNA into the pronucleus, (2) injection of RNA into the pronucleus, and (3) injection of RNA into the cytoplasm. We found that injection of RNA into the cytoplasm was the most efficient method in terms of the numbers of viable blastocyst stage embryos and full-term pups generated. This method also showed the best overall knockout efficiency.