Electrophoretic mobility shift assays for protein-DNA complexes involved in DNA repair.
Electrophoretic mobility shift assays for protein-DNA complexes involved in DNA repair.
复制标题
DOI：
10.1007/978-1-61779-998-3_5
复制
发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Chu, Gilbert
中科院分区：
文献类型：
--
作者：
Tsai, Chun;Smider, Vaughn;Hwang, Byung Joon;Chu, Gilbert
电泳迁移率变动分析 (EMSA) 可用于研究与 DNA 损伤剂产生的 DNA 结构结合的蛋白质。紫外线损伤的 DNA 结合蛋白 (UV-DDB) 参与核苷酸切除修复，与紫外线辐射或抗癌药物顺铂损伤的 DNA 结合。 Ku、XRCC4/Ligase IV 和 DNA-PKcs 参与通过非同源末端连接修复 DNA 双链断裂，在 DNA 末端组装成复合物。本章将描述用于检测不同 DNA 修复蛋白-DNA 复合物的几种 EMSA 方案。为了获得更多信息，可以应用 EMSA 的变体，其中包括用于检测 35S 标记蛋白质与受损 DNA 结合的反向 EMSA，以及用于检测蛋白质-DNA 复合物中特定蛋白质的存在的抗体超位移测定。
The electrophoretic mobility shift assay (EMSA) can be used to study proteins that bind to DNA structures created by DNA-damaging agents. UV-damaged DNA-binding protein (UV-DDB), which is involved in nucleotide excision repair, binds to DNA damaged by ultraviolet radiation or the anticancer drug cisplatin. Ku, XRCC4/Ligase IV, and DNA–PKcs, which are involved in the repair of DNA double-strand breaks by nonhomologous end joining, assemble in complexes at DNA ends. This chapter will describe several EMSA protocols for detecting different DNA repair protein–DNA complexes. To obtain additional information, one can apply variations of the EMSA, which include the reverse EMSA to detect binding of 35S-labeled protein to damaged DNA, and the antibody supershift assay to detect the presence of a specific protein in the protein–DNA complex.