Genome-wide profiling of prime editor off-target sites in vitro and in vivo using PE-tag
Genome-wide profiling of prime editor off-target sites in vitro and in vivo using PE-tag
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DOI：
10.1038/s41592-023-01859-2
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发表时间：
2023-05-08
期刊：
影响因子：
48
通讯作者：
Wolfe, Scot A.
中科院分区：
文献类型：
--
作者：
Liang, Shun-Qing;Liu, Pengpeng;Wolfe, Scot A.
Prime编辑器具有广泛的潜在研究和临床应用。然而，描述其全基因组编辑活动的方法通常依赖于间接的全基因组编辑评估或近同源序列的计算预测。在这里，我们描述了一个全基因组的方法来识别潜在的主要编辑脱靶位点，我们称之为PE标签。该方法依赖于在引物编辑活性位点处连接或插入扩增标签以允许它们的鉴定。PE标签能够使用提取的基因组DNA在体外、哺乳动物细胞系和成年小鼠肝脏中对脱靶位点进行全基因组分析。PE标签组分可以以多种形式递送用于脱靶位点检测。我们的研究与先前描述的引物编辑系统的高特异性一致，但我们发现脱靶编辑率受到引物编辑指导RNA设计的影响。PE-tag是一种可用于全基因组范围内鉴定prime editor活性和评价prime editor安全性的方法，可用于体外和体内prime editing脱靶效应的全基因组定位。
Prime editors have a broad range of potential research and clinical applications. However, methods to delineate their genome-wide editing activities have generally relied on indirect genome-wide editing assessments or the computational prediction of near-cognate sequences. Here we describe a genome-wide approach for the identification of potential prime editor off-target sites, which we call PE-tag. This method relies on the attachment or insertion of an amplification tag at sites of prime editor activity to allow their identification. PE-tag enables genome-wide profiling of off-target sites in vitro using extracted genomic DNA, in mammalian cell lines and in the adult mouse liver. PE-tag components can be delivered in a variety of formats for off-target site detection. Our studies are consistent with the high specificity previously described for prime editor systems, but we find that off-target editing rates are influenced by prime editing guide RNA design. PE-tag represents an accessible, rapid and sensitive approach for the genome-wide identification of prime editor activity and the evaluation of prime editor safety.PE-tag is a genome-wide approach to map the off-target effects of in vitro and in vivo prime editing.