Figure 2.

ESC Culture Conditions Influence the Activity of Oct4 Enhancers

(A and B) Derivation of ESCs from O4-DE-GFP/O4-PE-RFP blastocysts. O4-PE-RFP was initially expressed during the establishment of ESCs. Scale bars, 50 μm.

(C) Phase and fluorescence images of O4-DE-GFP/O4-PE-RFP ESCs in serum + LIF medium. Scale bars, 50 μm.

(D) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells in serum + LIF medium.

(E) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells from sorted O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells cultured in serum + LIF medium for ten passages.

(F) Phase and fluorescence images of O4-DE-GFP/O4-PE-RFP ESCs in serum + LIF + 2i medium. Scale bars, 50 μm.

(G) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells in serum + LIF + 2i medium.

(H) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells from sorted O4-DE-GFP⁺only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells cultured in serum + LIF + 2i medium for ten passages.

(I) Phase and fluorescence images of O4-DE-GFP/O4-PE-RFP ESCs in N2B27 + LIF + 2i medium. Scale bars, 50 μm.

(J and K) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells cultured in N2B27 + LIF + 2i medium for 2 days (J) or ten passages (K).

(L) Flow cytometry analysis of the proportion of O4-DE-GFP⁺ only cells or O4-DE-GFP⁺ and O4-PE-RFP⁺ cells cultured in serum + LIF medium for 2 days from N2B27 + LIF + 2i medium.