Department of Medical Genetics, University of Alberta, Edmonton, AB, Canada.
Johns Hopkins School of Medicine, Baltimore, MD, USA.
Methods Mol Biol. 2022;2579:87-97. doi: 10.1007/978-1-0716-2736-5_7.
The typical cell cycle in eukaryotes is composed of four phases including the G1, S, G2, and M phases. G1, S, and G2 together are called interphase. Cell synchronization is a process that brings cultured cells at different stages of the cell cycle to the same phase, which allows the study of phase-specific cellular events. While interphase cells can be easily distinguished from mitotic cells by examining their chromosome morphology, it is much more difficult to separate and distinguish the interphases from each other. Here, we describe drug-derived protocols for synchronizing HeLa cells to various interphases of the cell cycle: G1 phase, S phase, and G2 phase. G1 phase synchronization is achieved through serum starvation, S phase synchronization is achieved through a double thymidine block, and G2 phase synchronization is achieved through the release of the double thymidine block followed by roscovitine treatment. Successful synchronization can be assessed using flow cytometry to examine the DNA content and Western blot to examine the expression of various cyclins.
真核生物的典型细胞周期由 G1、S、G2 和 M 期四个阶段组成。G1、S 和 G2 合称为间期。细胞同步化是将处于不同细胞周期阶段的培养细胞带到同一阶段的过程，这允许研究特定于相的细胞事件。虽然间期细胞可以通过检查其染色体形态很容易地与有丝分裂细胞区分开来，但更难以将它们彼此分开并区分。在这里，我们描述了用于将 HeLa 细胞同步到细胞周期的各种间期的药物衍生方案：G1 期、S 期和 G2 期。通过血清饥饿实现 G1 期同步，通过双重胸苷阻断实现 S 期同步，通过释放双重胸苷阻断并随后用罗司维亭处理实现 G2 期同步。成功的同步可以使用流式细胞术检查 DNA 含量和 Western blot 检查各种细胞周期蛋白的表达来评估。
