An efficient gene knock-in strategy using 5′-modified double-stranded DNA donors with short homology arms

Yi Yu, Yijun Guo, Qiqi Tian, Yuanqing Lan, Hugh Yeh, Meng Zhang, Ipek Tasan, Surbhi Jain, Huimin Zhao

Research output: Contribution to journalArticle

Abstract

Here, we report a rapid CRISPR–Cas9-mediated gene knock-in strategy that uses Cas9 ribonucleoprotein and 5′-modified double-stranded DNA donors with 50-base-pair homology arms and achieved unprecedented 65/40% knock-in rates for 0.7/2.5 kilobase inserts, respectively, in human embryonic kidney 293T cells. The identified 5′-end modification led to up to a fivefold increase in gene knock-in rates at various genomic loci in human cancer and stem cells.

Original languageEnglish (US)
JournalNature chemical biology
DOIs
StateAccepted/In press - Dec 23 2019

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ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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