Figure S2.

Cyclin D Expression in hESCs and during Germ Layer Specification, Related to Figure 3

(A) Smad2/3 transcriptional activity during progression of the cell cycle. Fucci-hESCs transfected with a reporter for Smad2/3 transcriptional activity (SBE4-luciferase construct) were incubated with Activin A for 3h followed by FACS sorting and analysis of luciferase activity.

(B) Cyclin D expression during cell cycle progression in hESCs. Western blot analysis for cyclin D1-D3 expression in FACS sorted Fucci-hESCs.

(C) Cyclin D expression in hESCs. Flow cytometry analysis of cyclin D expression in Oct4+ pluripotent cells.

(D) Cyclin D expression in hESCs is stable. hESCs were collected at day 3 after three days of splittings and analyzed by western blot.

(E) Cyclin D expression in pluripotent cells. Immunofluorescence microscopy of pluripotency markers and cyclin D1-3 in hESCs.

(F–K) Cyclin D expression during differentiation of hESCs into neuroectoderm (F, G), endoderm (H, I) and mesoderm (J, K) by Q-PCR and immunostaining, respectively. UD - undifferentiated H9; D1-D9 day 1 to 9. Data shown as mean ± SD (n = 3). Scale bar, 100μm.