Fig. 1: ZFP281 is required for proper S phase progression in mouse ES cells.

a Western blot showing successful depletion of ZFP281 in mouse ES cells by CRISPR-Cas9. α-TUBULIN was used as a loading control. Four independent experiments show similar results. b Growth curve showing numbers of WT (yellow), ZFP281 KO-1 (green), and ZFP281 KO-2 (red) mouse ES cells counted at different time intervals after plating. Mean ± SEM from three independent experiments. c Flow cytometry analysis showing the percentage of WT, ZFP281 KO-1, and ZFP281 KO-2 ES cell at different cell cycle phases. Three independent experiments show similar results. d Western blot analysis (upper panel) showing the levels of Cyclin A2, Cyclin E1 and Cyclin C in WT, ZFP281 KO-1, and ZFP281 KO-2 ES cells. Histone H3 was used as a loading control. Gray values (lower panel) of Cyclin A2, Cyclin E1 and Cyclin C protein bands were determined by Image J and normalized to the loading control. Mean ± SEM from three independent experiments. Two-tailed, unpaired Student’s t tests were performed. (Cyclin A2, WT vs. ZFP281 KO-1, p = 0.0005; WT vs. ZFP281 KO-2, p = 0.0003. Cyclin E1, WT vs. ZFP281 KO-1, p = 0.0001; WT vs. ZFP281 KO-2, p = 0.0001. Cyclin C, WT vs. ZFP281 KO-1, p = 0.6566; WT vs. ZFP281 KO-2, p = 0.0500.) ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, n.s. = not significant. e EdU staining pattern (upper left) of WT and ZFP281 KO ES cells, DNA was counterstained using DAPI; Representative images showing the EdU staining pattern (upper right) of mouse ES cells at early, middle and late S phase; Percentage of WT (total n = 422) and ZFP281 KO ES (total n = 338) cells at different stages of S phase. Three independent experiments show similar results. Source data are provided as a Source Data file.