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. 2009 Jan 12;184(1):67–82. doi: 10.1083/jcb.200801009

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© 2009 Zhang et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Figure 3. — Changes in hESC cell cycle distribution as a result of NANOG overexpression. (A) Flow cytometry images showing movement of cells through the cell cycle after synchronization by nocodazole for 18 h assessed by propidium iodide staining. This figure represents an example of three independent experiments. (B) Chart representation of the fraction of cells in S phase assessed by propidium iodide staining over time after release from synchronization with nocodazole for 18 h. (C) Chart representation of the fraction of cells in S phase assessed by BrdU incorporation over time after release from synchronization with nocodazole for 18 h. The bottom panel shows flow cytometry images of BrdU-incorporating cells in NANOG and control H9 hESC sublines after 5 h of release from nocodazole. The dashed line represents the end of G1 and the start of S phase. (B and C) The figure represents an example of three independent experiments performed in H9 sublines. (D) Western blotting for expression of main components involved in G1 to S transition in NANOG-overexpressing and control clones. This summary is a representation of three independent experiments. GAPDH was used as a loading control. p16 and p15 were absent in all sublines, whereas p19 and p18 were expressed at very low but equal levels in NANOG-overexpressing and control clones (not depicted). Molecular masses are indicated in kilodaltons. HN, H1 NANOG; HC, H1 control; NN, hES-NCL1 NANOG; NC, hES-NCL1 control.