基因组编辑
诱导多能干细胞
清脆的
Cas9
生物
计算生物学
基因组工程
基因
人类基因组
遗传学
胞嘧啶
胚胎干细胞
DNA
基因组
作者
Giulia Pavani,Joshua G. Klein,Deborah L. French,Paul Gadue
出处
期刊:Methods in molecular biology
日期:2022-01-01
卷期号:: 321-333
标识
DOI:10.1007/7651_2022_461
摘要
The ability to engineer specific mutations in human embryonic stem cells (ECSs) or induced pluripotent stem cells (iPSCs) is extremely important in the modeling of human diseases and the study of biological processes. While CRISPR/Cas9 can robustly generate gene knockouts (KOs) and gene loci modifications in coding sequences of iPSCs, it remains difficult to produce monoallelic mutations or modify specific nucleotides in noncoding sequences due to technical constraints.Here, we describe how to leverage cytosine (BE4max) and adenine (ABEmax) base editors to introduce precise mutations in iPSCs without inducing DNA double-stranded breaks. This chapter illustrates how to design and clone gRNAs, evaluate editing efficiency, and detect genomic edits at specific sites in iPSCs through the utilization of base editing technology.
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