利用CRISPR/Cas9技术对人多能干细胞进行高效基因组编辑

摘要

CRISPR/Cas9技术提供了一个全新的基因组编辑体系。本文利用CRISPR/Cas9平台，在人胚胎干细胞株中对选取的一段特定基因组区域进行了多种基因组编辑：通过在基因编码框中引入移码突变进行基因敲除；通过单链DNA提供外源模板经由同源重组定点敲入FLAG序列；通过同时靶向多个位点诱导基因组大片段删除。研究结果表明 CRISPR/Cas9可以对多能干细胞进行高效基因编辑，获得的突变干细胞株有助于对基因和基因组区域的功能进行分析和干细胞疾病模型的建立。%The RNA-guided CRISPR (clustered regularly interspaced short palindromic repeat)-associated Cas9 nuclease has offered a new platform for genome editing with high efficiency. Here, we report the use of CRISPR/Cas9 technology to target a specific genomic region in human pluripotent stem cells. We show that CRISPR/Cas9 can be used to disrupt a gene by introducing frameshift mutations to gene coding region; to knock in specific sequences (e.g. FLAG tag DNA sequence) to targeted genomic locusvia homology directed repair; to induce large genomic deletion through dual-guide multiplex. Our results demonstrate the versatile application of CRISPR/Cas9 in stem cell genome editing, which can be widely utilized for functional studies of genes or genome loci in human pluripotent stem cells.