Lin Yunfeng, Ha Anh, Yan Shan
Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, NC, USA.
Methods Mol Biol. 2019;1999:161-172. doi: 10.1007/978-1-4939-9500-4_9.
DNA single-strand breaks (SSBs) are the most common type of DNA lesions as they are generated approximately 10,000 times per mammalian cell each day. Unrepaired SSBs compromise DNA replication and transcription programs, leading to genome instability, and have been implicated in many diseases including cancer. In this chapter, we introduce methods to study the ATR-Chk1 DNA damage response (DDR) pathway and DNA repair pathway in response to a site-specific, defined SSB plasmid in Xenopus laevis egg extracts. This experimental system can be applied in future studies to reveal many aspects of the molecular mechanisms of SSB repair and signaling in eukaryotes.
DNA单链断裂(SSB)是最常见的DNA损伤类型,因为它们每天在每个哺乳动物细胞中大约产生10000次。未修复的SSB会损害DNA复制和转录程序,导致基因组不稳定,并与包括癌症在内的许多疾病有关。在本章中,我们介绍了在非洲爪蟾卵提取物中研究ATR-Chk1 DNA损伤反应(DDR)途径和DNA修复途径的方法,以应对位点特异性、明确的SSB质粒。该实验系统可应用于未来的研究,以揭示真核生物中SSB修复和信号传导分子机制的许多方面。