Journal
GENETICS
Volume 200, Issue 2, Pages 423-+Publisher
GENETICS SOCIETY AMERICA
DOI: 10.1534/genetics.115.176594
Keywords
CRISPR; Cas9; electroporation; mouse zygote
Categories
Funding
- National Cancer Institute [P30CA034196]
- Jackson Laboratory
- JAX postdoctoral scholar fellowship
- National Institute of Diabetes and Digestive and Kidney Diseases [DK097605]
- National Natural Science Foundation of China [31471215]
- Strategic Priority Research Program of the Chinese Academy of Sciences [XDA01010409]
- State 863 Project [2015AA020307]
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The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas) system is an adaptive immune system in bacteria and archaea that has recently been exploited for genome engineering. Mutant mice can be generated in one step through direct delivery of the CRISPR/Cas9 components into a mouse zygote. Although the technology is robust, delivery remains a bottleneck, as it involves manual injection of the components into the pronuclei or the cytoplasm of mouse zygotes, which is technically demanding and inherently low throughput. To overcome this limitation, we employed electroporation as a means to deliver the CRISPR/Cas9 components, including Cas9 messenger RNA, single-guide RNA, and donor oligonucleotide, into mouse zygotes and recovered live mice with targeted nonhomologous end joining and homology-directed repair mutations with high efficiency. Our results demonstrate that mice carrying CRISPR/Cas9-mediated targeted mutations can be obtained with high efficiency by zygote electroporation.
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