Institut de Génétique et de Biologie Moléculaire et Celullaire, INSERM U964, CNRS, UMR7104, Illkirch, France.
Université de Strasbourg, Strasbourg, France.
Methods Mol Biol. 2021;2153:439-445. doi: 10.1007/978-1-0716-0644-5_30.
CRISPR/Cas9 technology can be used to investigate how double-strand breaks (DSBs) occurring in constitutive heterochromatin are getting repaired. This technology can be used to induce specific breaks on mouse pericentromeric heterochromatin, by using a guide RNA specific for the major satellite repeats and co-expressing it with Cas9. Those clean DSBs can be visualized later by confocal microscopy. More specifically, immunofluorescence can be used to visualize the main factors of each DSB repair pathway and quantify their percentage and pattern of recruitment at the heterochromatic region.
CRISPR/Cas9 技术可用于研究在组成型异染色质中发生的双链断裂（DSBs）如何被修复。该技术可用于通过使用针对主要卫星重复序列的指导 RNA 并与 Cas9 共表达，在小鼠着丝粒异染色质上诱导特定的断裂。这些干净的 DSB 随后可以通过共聚焦显微镜观察到。更具体地说，可以使用免疫荧光法来观察每个 DSB 修复途径的主要因素，并量化它们在异染色质区域的募集百分比和模式。
