One-step generation of mice carrying mutations in multiple genes by CRISPR/cas-mediated genome engineering

Haoyi Wang, Hui Yang, Chikdu S. Shivalila, Meelad M. Dawlaty, Albert W. Cheng, Feng Zhang, Rudolf Jaenisch

Research output: Contribution to journalArticle

1855 Citations (Scopus)

Abstract

Mice carrying mutations in multiple genes are traditionally generated by sequential recombination in embryonic stem cells and/or time-consuming intercrossing of mice with a single mutation. The CRISPR/Cas system has been adapted as an efficient gene-targeting technology with the potential for multiplexed genome editing. We demonstrate that CRISPR/Cas-mediated gene editing allows the simultaneous disruption of five genes (Tet1, 2, 3, Sry, Uty - 8 alleles) in mouse embryonic stem (ES) cells with high efficiency. Coinjection of Cas9 mRNA and single-guide RNAs (sgRNAs) targeting Tet1 and Tet2 into zygotes generated mice with biallelic mutations in both genes with an efficiency of 80%. Finally, we show that coinjection of Cas9 mRNA/sgRNAs with mutant oligos generated precise point mutations simultaneously in two target genes. Thus, the CRISPR/Cas system allows the one-step generation of animals carrying mutations in multiple genes, an approach that will greatly accelerate the in vivo study of functionally redundant genes and of epistatic gene interactions.

Original languageEnglish (US)
Pages (from-to)910-918
Number of pages9
JournalCell
Volume153
Issue number4
DOIs
StatePublished - May 9 2013
Externally publishedYes

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Clustered Regularly Interspaced Short Palindromic Repeats
Genes
CRISPR-Cas Systems
Genome
Guide RNA
Mutation
Genetic Epistasis
Messenger RNA
Gene Targeting
Zygote
Embryonic Stem Cells
Stem cells
Point Mutation
Genetic Recombination
Alleles
Technology
Animals

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

One-step generation of mice carrying mutations in multiple genes by CRISPR/cas-mediated genome engineering. / Wang, Haoyi; Yang, Hui; Shivalila, Chikdu S.; Dawlaty, Meelad M.; Cheng, Albert W.; Zhang, Feng; Jaenisch, Rudolf.

In: Cell, Vol. 153, No. 4, 09.05.2013, p. 910-918.

Research output: Contribution to journalArticle

Wang, Haoyi ; Yang, Hui ; Shivalila, Chikdu S. ; Dawlaty, Meelad M. ; Cheng, Albert W. ; Zhang, Feng ; Jaenisch, Rudolf. / One-step generation of mice carrying mutations in multiple genes by CRISPR/cas-mediated genome engineering. In: Cell. 2013 ; Vol. 153, No. 4. pp. 910-918.
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