Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
STAR Protoc. 2023 Sep 15;4(3):102487. doi: 10.1016/j.xpro.2023.102487. Epub 2023 Aug 6.
Detecting DNA breaks in defined regions of the genome is critical to advancing our understanding of genome stability maintenance. Here, we present exo-FISH, a protocol to label exposed single-stranded DNA in defined repetitive regions of mammalian genomes by combining in vitro restriction enzyme digestion on fixed cells with fluorescence in situ hybridization (FISH). We describe steps for cell harvesting and fixation, slide treatments, and FISH probe hybridization. We then detail procedures for imaging and analysis. For complete details on the use and execution of this protocol, please refer to Saayman et al. (2023)..
检测基因组特定区域的 DNA 断裂对于深入了解基因组稳定性维持至关重要。这里，我们介绍了 exo-FISH 技术，该技术将固定细胞的体外限制性内切酶消化与荧光原位杂交（FISH）相结合，标记哺乳动物基因组中特定重复区域的暴露单链 DNA。我们描述了细胞收获和固定、载玻片处理以及 FISH 探针杂交的步骤。然后，我们详细介绍了成像和分析的过程。如需详细了解该方案的使用和执行，请参阅 Saayman 等人（2023 年）的研究。
