Protocol for editing fibroblasts with in vitro transcribed Cas9 mRNA and profile off-target editing by optimized GUIDE-seq

被引：1

作者：

Li, Zhuokun ^{[1
]}

Reint, Ganna ^{[1
]}

Haapaniemi, Emma Maria ^{[1
]}

机构：

[1] Univ Oslo, Ctr Mol Med Norway, Oslo, Norway

来源：

STAR PROTOCOLS | 2023年 / 4卷 / 04期

基金：

芬兰科学院;

关键词：

Cell Culture; CRISPR; Sequencing;

D O I：

10.1016/j.xpro.2023.102662

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

CRISPR-Cas9 gene editing is an efficient technique to modify specific sites/regions of DNA. Delivery of the Cas9 by mRNA is particularly promising in pre -clinical genome editing applications for its transient, nonintegrating feature. However, the off-target of Cas9-gRNA still remains a concern and needs a specific monitor. Here, we present a revised protocol to edit fibroblasts by in vitro transcribed Cas9 mRNA and profile its off-target effect by the optimized GUIDE-seq method. This protocol can also be applied to other cell lines. For complete details on the use and execution of this protocol, please refer to Ganna Reint et al. (2021).1

引用

页数：21

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