Expression of CBF:H2B-Venus in ES cells and pre-streak stages. Immunostaining of the CBF:H2B-Venus ES cells (A-A3) for pluripotency-associated factors Nanog (grey) (A2) and Oct4 (red) (A3). Nuclei were stained with Hoechst (blue) (A). Embryoid body formation from CBF:H2B-Venus ES cells. Fluorescence was assessed after 1, 2, 4 and 6 days of EB culture (B-E). Expression of CBF:H2B-Venus in the epiblast at E6.0 (F) and E5.5 (G-I). Expression of Gata6 (G1, G2), Oct4 (H1, H2), and Eomes (I1, I2) in CBF:H2B-Venus embryos. At E6.0 CBF:H2B-Venus expression is detected in the epiblast in a salt and pepper distribution as well as in the extra-embryonic ectoderm (F, F1, F2). At E5.5 CBF:H2B-Venus expression is restricted to the epiblast (G, H and I). H2B-Venus positive cells express also Oct4 (H2). Note, Gata6 expression in the epiblast (white arrowhead, G2). Embryo has been counterstained with Hoechst (F1, F2) and Phalloidin (F2) to label nuclei and F-Actin. A, anterior; D, distal; em, embryonic; ex, extra-embryonic; P, posterior; Pr, proximal. Scale bars: 50 μm.