Efficient CRISPR/Cas9-Mediated Genome Editing in Mice by Zygote Electroporation of Nuclease. Genetics 2015 Jun;200(2):423-30
Date
03/31/2015Pubmed ID
25819794Pubmed Central ID
PMC4492369DOI
10.1534/genetics.115.176594Scopus ID
2-s2.0-84931385060 (requires institutional sign-in at Scopus site) 234 CitationsAbstract
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.
Author List
Qin W, Dion SL, Kutny PM, Zhang Y, Cheng AW, Jillette NL, Malhotra A, Geurts AM, Chen YG, Wang HAuthors
Yi-Guang Chen PhD Professor in the Pediatrics department at Medical College of WisconsinAron Geurts PhD Professor in the Physiology department at Medical College of Wisconsin
MESH terms used to index this publication - Major topics in bold
AnimalsBase Sequence
CRISPR-Cas Systems
Electroporation
Endonucleases
Female
Gene Targeting
Genetic Loci
Genome
Genomics
INDEL Mutation
Mice
Molecular Sequence Data
RNA Editing
RNA, Messenger
Sequence Alignment
Zygote
