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      One-Step Generation of Mice Carrying Mutations in Multiple Genes by CRISPR/Cas-Mediated Genome Engineering

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      Cell

      Elsevier BV

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          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. Copyright © 2013 Elsevier Inc. All rights reserved.

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          Author and article information

          Journal
          Cell
          Cell
          Elsevier BV
          00928674
          May 2013
          May 2013
          : 153
          : 4
          : 910-918
          Article
          10.1016/j.cell.2013.04.025
          3969854
          23643243
          © 2013

          https://www.elsevier.com/tdm/userlicense/1.0/

          https://www.elsevier.com/open-access/userlicense/1.0/

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