使用ROPS（随机寡核苷酸引发合成）分析法检测和定量DNA链断裂。

Detection and quantification of DNA strand breaks using the ROPS (random oligonucleotide primed synthesis) assay.

作者信息

Boyko Alex, Kovalchuk Igor

机构信息

Department of Biological Sciences, University of Lethbridge, Lethbridge, AB, Canada.

出版信息

Methods Mol Biol. 2010;631:237-42. doi: 10.1007/978-1-60761-646-7_17.

DOI:10.1007/978-1-60761-646-7_17

PMID:20204879

Abstract

DNA double strand breaks (DSBs) arise from spontaneous DNA damage due to metabolic activities or from direct and indirect damaging effects of stress. DSBs are also formed transiently during such processes as replication, transcription, and DNA repair. The level of DSBs positively correlates with the activities of homologous and nonhomologous DNA repair pathways, which in turn inversely correlate with methylation levels and chromatin structure. Thus, measurement of strand breaks can provide an informative picture of genome stability of a given cell. The use of random oligonucleotide-primed synthesis for the analysis of DSB levels is described. Applications of the assay for quantitative detection of 3'OH, 3'P, or DNA strand breaks at a cleavage site of the deoxyribose residue are discussed.

摘要

DNA双链断裂（DSBs）源于代谢活动导致的自发DNA损伤，或应激的直接和间接损伤效应。在复制、转录和DNA修复等过程中，DSBs也会短暂形成。DSBs的水平与同源和非同源DNA修复途径的活性呈正相关，而这些修复途径的活性又与甲基化水平和染色质结构呈负相关。因此，测量链断裂可以提供给定细胞基因组稳定性的信息图。本文描述了使用随机寡核苷酸引发合成来分析DSB水平。讨论了该测定法在定量检测脱氧核糖残基切割位点处的3'OH、3'P或DNA链断裂中的应用。