Effect of Cell Division on Cell Differentiation (ChIP-Seq HUvsNoc). Effect of Cell Division on Cell Differentiation (ChIP-Seq HUvsNoc)

We addressed the role of cell division during differentiation by blocking the progression of the cell cycle during mesendoderm differentiation. Human pluripotent stem cells (H9 cells) were synchronized in their cell cycle and then induced to differentiate into mesendoderm for 24h. To block the cells in their S phase, we added Hydroxyurea (200 uM) at the beginning of the differentiation process. To block the cells in their G2/M phases, we added nocodazole (100 ng/ml) 8h after the induction of differentiation. At the end of the experiment, the cells were harvested and processed for RNA extraction. Overall design: Four different conditions were analyzed: Undifferentiated (UD), Mesendoderm (E24), Mesendoderm + Hydroxyurea (E24HU), and Mesendoderm + nocodazole (E24Noc). Two biological replicates were generated for each condition.

本研究通过阻断中内胚层分化过程中的细胞周期进程，探究细胞分裂在分化进程中的功能。将人多能干细胞（H9细胞）进行细胞周期同步化处理后，诱导其向中内胚层分化24小时。为将细胞阻滞于S期，我们于分化起始阶段添加200 μM的羟基脲（Hydroxyurea）；为将细胞阻滞于G2/M期，则在分化诱导8小时后加入100 ng/ml的诺考达唑（nocodazole）。实验结束后，收集细胞并开展RNA提取操作。
总体实验设计：共分析四种不同条件，分别为未分化组（UD）、中内胚层组（E24）、中内胚层+羟基脲组（E24HU）以及中内胚层+诺考达唑组（E24Noc），每组均设置两个生物学重复。