利用果蝇原位缺口平移技术研究发育和凋亡过程中DNA链断裂的实验方案。

Maurya Deepak, Mondal Bama Charan

Cytogenetics Laboratory, Department of Zoology, Institute of Science, Banaras Hindu University, Varanasi 221005, India.

STAR Protoc. 2025 Jun 26;6(3):103921. doi: 10.1016/j.xpro.2025.103921.

Cellular stress causes DNA strand breaks that are typically repaired to maintain homeostasis and regulate cell fate. However, unrepaired DNA breaks can be lethal, leading to cell death. Here, we present a protocol to study DNA strand breaks in Drosophila during development and apoptosis using in situ nick translation. We describe the steps for labeling DNA strand breaks using digoxigenin (DIG)-labeled nucleotide (DIG-11-dUTP) and visualizing them with anti-DIG immunostaining. We then detail procedures for mounting, imaging, and analysis. For complete details on the use and execution of this protocol, please refer to Maurya et al. and Rigby et al..

细胞应激会导致DNA链断裂，这些断裂通常会被修复以维持体内平衡并调节细胞命运。然而，未修复的DNA断裂可能是致命的，会导致细胞死亡。在这里，我们展示了一种使用原位缺口平移法研究果蝇发育和凋亡过程中DNA链断裂的方案。我们描述了使用地高辛（DIG）标记的核苷酸（DIG-11-dUTP）标记DNA链断裂并通过抗DIG免疫染色进行可视化的步骤。然后，我们详细说明了固定、成像和分析的程序。有关此方案的使用和执行的完整详细信息，请参考Maurya等人和Rigby等人的文献。