Precision genome editing using CRISPR-Cas9 and linear repair templates in C. elegans

被引：141

作者：

Paix, Alexandre ^{[1
]}

Folkmann, Andrew ^{[1
]}

Seydoux, Geraldine ^{[1
]}

机构：

[1] Johns Hopkins Univ, Sch Med, Dept Mol Biol & Genet, 725 N Wolfe St, Baltimore, MD 21205 USA

来源：

METHODS | 2017年 / 121卷

基金：

美国国家卫生研究院;

关键词：

Genome editing; CRISPR; C; elegans; Cas9; RNP; Linear repair templates; Short homology; Gene conversion; Homology-dependent repair (HDR); CRISPR/CAS9; EFFICIENCY;

D O I：

10.1016/j.ymeth.2017.03.023

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

The ability to introduce targeted edits in the genome of model organisms is revolutionizing the field of genetics. State-of-the-art methods for precision genome editing use RNA-guided endonucleases to create double-strand breaks (DSBs) and DNA templates containing the edits to repair the DSBs. Following this strategy, we have developed a protocol to create precise edits in the C. elegans genome. The protocol takes advantage of two innovations to improve editing efficiency: direct injection of CRISPR-Cas9 ribonucleoprotein complexes and use of linear DNAs with short homology arms as repair templates. The protocol requires no cloning or selection, and can be used to generate base and gene-size edits in just 4 days. Point mutations, insertions, deletions and gene replacements can all be created using the same experimental pipeline. (C) 2017 The Authors. Published by Elsevier Inc.

引用

页码：86 / 93

页数：8

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