Institute of Pathology, Albert Szent-Györgyi Medical School, University of Szeged, 1 Állomás Street H-6725, Szeged, Hungary.
Mutat Res Rev Mutat Res. 2021 Jul-Dec;788:108397. doi: 10.1016/j.mrrev.2021.108397. Epub 2021 Oct 29.
DNA double-strand breaks (DSBs) are one of the most frequent causes of initiating cancerous malformations, therefore, to reduce the risk, cells have developed sophisticated DNA repair mechanisms. These pathways ensure proper cellular function and genome integrity. However, any alteration or malfunction during DNA repair can influence cellular homeostasis, as improper recognition of the DNA damage or dysregulation of the repair process can lead to genome instability. Several powerful methods have been established to extend our current knowledge in the field of DNA repair. For this reason, in this review, we focus on the methods used to study DSB repair, and we summarize the advantages and disadvantages of the most commonly used techniques currently available for the site-specific induction of DSBs and the subsequent tracking of the repair processes in human cells. We highlight methods that are suitable for site-specific DSB induction (by restriction endonucleases, CRISPR-mediated DSB induction and laser microirradiation) as well as approaches [e.g., fluorescence-, confocal- and super-resolution microscopy, chromatin immunoprecipitation (ChIP), DSB-labeling and sequencing techniques] to visualize and follow the kinetics of DSB repair.
DNA 双链断裂 (DSBs) 是引发癌变畸形的最常见原因之一，因此，为了降低风险，细胞已经发展出了复杂的 DNA 修复机制。这些途径确保了细胞的正常功能和基因组完整性。然而，在 DNA 修复过程中的任何改变或故障都会影响细胞的动态平衡，因为对 DNA 损伤的不正确识别或修复过程的失调可能导致基因组不稳定。已经建立了几种强大的方法来扩展我们在 DNA 修复领域的现有知识。出于这个原因，在这篇综述中，我们专注于用于研究 DSB 修复的方法，并总结了目前用于在人类细胞中特异性诱导 DSB 并随后跟踪修复过程的最常用技术的优缺点。我们强调了适合特异性 DSB 诱导的方法（通过限制性内切酶、CRISPR 介导的 DSB 诱导和激光微照射）以及用于可视化和跟踪 DSB 修复动力学的方法 [例如，荧光、共聚焦和超分辨率显微镜、染色质免疫沉淀 (ChIP)、DSB 标记和测序技术]。
