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

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

Nature chemical biology (2019-12-25)
Yi Yu, Yijun Guo, Qiqi Tian, Yuanqing Lan, Hugh Yeh, Meng Zhang, Ipek Tasan, Surbhi Jain, Huimin Zhao
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.

MATERIALS
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Brand
Product Description

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2-(Acetylthio)acetic acid, ≥90%
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(+)-Biotin N-hydroxysuccinimide ester, ≥98% (HPLC), powder
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