Nakanishi Koji, Cavallo Francesca, Brunet Erika, Jasin Maria
Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
Methods Mol Biol. 2011;745:283-91. doi: 10.1007/978-1-61779-129-1_16.
DNA interstrand cross-links (ICLs) covalently link both strands of the DNA duplex, impeding cellular processes like DNA replication. Homologous recombination (HR) is considered to be a major pathway for the repair of ICLs in mammalian cells as mutants for HR components are highly sensitive to DNA-damaging agents that cause ICLs. This chapter describes GFP assays to measure HR following site-specific ICL formation with psoralen through DNA triplex technology. This approach can be used to determine the genetic requirements for ICL-induced HR in relation to those involved in HR repair of other DNA lesions such as double-strand breaks.
DNA链间交联（ICL）共价连接DNA双链的两条链，阻碍DNA复制等细胞过程。同源重组（HR）被认为是哺乳动物细胞中ICL修复的主要途径，因为HR组分的突变体对导致ICL的DNA损伤剂高度敏感。本章描述了通过DNA三链体技术用补骨脂素进行位点特异性ICL形成后测量HR的绿色荧光蛋白（GFP）检测方法。这种方法可用于确定ICL诱导的HR相对于参与其他DNA损伤（如双链断裂）的HR修复的遗传需求。
