Bandaru Viswanath, Blaisdell Jeffrey O, Wallace Susan S
Department of Microbiology and Molecular Genetics, Markey Center for Molecular Genetics, University of Vermont, Burlington, USA.
Methods Enzymol. 2006;408:15-33. doi: 10.1016/S0076-6879(06)08002-5.
As new organisms are being sequenced on a daily basis, new DNA glycosylases that recognize DNA damage can be easily identified in an effort to understand both their phylogenetics and substrate specificities. As a practical matter, existing bacterial and human homologs need to be readily available as laboratory reagents in order to compare the activities of the novel enzymes to existing enzymes. This chapter attempts to provide a primer for cloning, expression, and assay procedures for bacterial and human DNA glycosylases that recognize oxidative DNA damages. These methodologies can be translated readily to novel DNA glycosylases or to DNA glycosylases that recognize other types of DNA damages.
由于每天都有新的生物体被测序，为了了解它们的系统发育和底物特异性，识别DNA损伤的新型DNA糖基化酶很容易被鉴定出来。实际上，现有的细菌和人类同源物需要作为实验室试剂随时可用，以便将新型酶的活性与现有酶进行比较。本章试图为识别氧化性DNA损伤的细菌和人类DNA糖基化酶的克隆、表达和检测程序提供一个入门指南。这些方法可以很容易地应用于新型DNA糖基化酶或识别其他类型DNA损伤的DNA糖基化酶。
