Cancer Signaling and Epigenetics Program, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
Cancer Epigenetics Institute, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
STAR Protoc. 2022 Mar 15;3(2):101243. doi: 10.1016/j.xpro.2022.101243. eCollection 2022 Jun 17.
Classic approaches to characterizing cell cycle leverage chemicals or altered nucleotide pools, which could impact chromatin states at specific phases of the cell cycle. Such approaches could induce metabolic alterations and/or DNA damage, which could reshape protein recruitment and histone modifications. In this protocol, we describe ways to fix and sort cells across the cell cycle based on their DNA content. We further detail immunoprecipitation and library preparation, allowing analysis of the epigenome by chromatin immunoprecipitation sequencing (ChIP-seq) for small numbers of cells. For complete details on the use and execution of this protocol, please refer to Van Rechem et al. (2021).
经典的细胞周期特征描述方法利用化学物质或改变核苷酸池，这可能会影响细胞周期特定阶段的染色质状态。这种方法可能会引起代谢改变和/或 DNA 损伤，从而重塑蛋白质募集和组蛋白修饰。在本方案中，我们描述了根据细胞的 DNA 含量固定和分选细胞的方法。我们进一步详细描述了免疫沉淀和文库制备，允许对小数量细胞的表观基因组进行染色质免疫沉淀测序 (ChIP-seq) 分析。有关该方案使用和执行的完整详细信息，请参阅 Van Rechem 等人。（2021 年）。
