Tarpley Mason, Chen Yingling, Bhakat Kishor K
Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE, USA.
Fred and Pamela Buffett Cancer Center, Omaha, NE, USA.
Methods Mol Biol. 2023;2701:243-252. doi: 10.1007/978-1-0716-3373-1_16.
The base excision repair (BER) is the primary damage repair pathway for repairing most of the endogenous DNA damage including oxidative base lesions, apurinic/apyrimidinic (AP) sites, and single-strand breaks (SSBs) in the genome. Repair of these damages in cells relies on sequential recruitment and coordinated actions of multiple DNA repair enzymes, which include DNA glycosylases (such as OGG1), AP-endonucleases (APE1), DNA polymerases, and DNA ligases. APE1 plays a key role in the BER pathway by repairing the AP sites and SSBs in the genome. Several methods have been developed to generate a map of endogenous AP sites or SSBs in the genome and the binding of DNA repair proteins. In this chapter, we describe detailed approaches to map genome-wide occupancy or enrichment of APE1 in human cells using chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq). Further, we discuss standard bioinformatics approaches for analyzing ChIP-seq data to identify APE1 enrichment or binding peaks in the genome.
碱基切除修复（BER）是修复基因组中大多数内源性DNA损伤的主要损伤修复途径，这些损伤包括氧化碱基损伤、脱嘌呤/脱嘧啶（AP）位点和单链断裂（SSB）。细胞中这些损伤的修复依赖于多种DNA修复酶的顺序募集和协同作用，这些酶包括DNA糖基化酶（如OGG1）、AP核酸内切酶（APE1）、DNA聚合酶和DNA连接酶。APE1通过修复基因组中的AP位点和SSB在BER途径中起关键作用。已经开发了几种方法来绘制基因组中内源性AP位点或SSB的图谱以及DNA修复蛋白的结合情况。在本章中，我们描述了使用染色质免疫沉淀结合下一代测序（ChIP-seq）来绘制人类细胞中APE1全基因组占有率或富集情况的详细方法。此外，我们讨论了用于分析ChIP-seq数据以识别基因组中APE1富集或结合峰的标准生物信息学方法。
