Characterization of genomic deletion efficiency mediated by clustered regularly interspaced palindromic repeats (CRISPR)/cas9 nuclease system in mammalian cells

Matthew C. Canver, Daniel E. Bauer, Abhishek Dass, Yvette Y. Yien, Jacky Chung, Takeshi Masuda, Takahiro Maeda, Barry H. Paw, Stuart H. Orkin

研究成果: ジャーナルへの寄稿学術誌査読

242 被引用数 (Scopus)

抄録

The clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9 nuclease system has provided a powerful tool for genome engineering. Double strand breaks may trigger nonhomologous end joining repair, leading to frameshift mutations, or homology-directed repair using an extrachromosomal template. Alternatively, genomic deletions may be produced by a pair of double strand breaks. The efficiency of CRISPR/Cas9-mediated genomic deletions has not been systematically explored. Here, we present a methodology for the production of deletions in mammalian cells, ranging from 1.3 kb to greater than 1 Mb. We observed a high frequency of intended genomic deletions. Nondeleted alleles are nonetheless often edited with inversions or small insertion/deletions produced at CRISPR recognition sites. Deleted alleles also typically include small insertion/deletions at predicted deletion junctions. We retrieved cells with biallelic deletion at a frequency exceeding that of probabilistic expectation. We demonstrate an inverse relationship between deletion frequency and deletion size. This work suggests that CRISPR/Cas9 is a robust system to produce a spectrum of genomic deletions to allow investigation of genes and genetic elements.

本文言語英語
ページ(範囲)21312-21324
ページ数13
ジャーナルJournal of Biological Chemistry
289
31
DOI
出版ステータス出版済み - 8月 1 2014
外部発表はい

!!!All Science Journal Classification (ASJC) codes

  • 生化学
  • 分子生物学
  • 細胞生物学

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