FGF-signalling in the differentiation of mouse ES cells towards ...

Figure S4Figure S4: Expression of TGF-β, Wnt and FGF signaling components during directed differentiation of EScells. RT-PCR analysis of T Gfp/+ cells cultured for 3 or 5 days in medium containing growth factors and/or inhibitor,as indicated. The expression of components of the nodal/activin, BMP, Wnt and FGF signaling pathway wasanalyzed. The expression of Tbp was analyzed as reference gene.Opposite, Figure S5: Wnt signaling is inhibited by addition of Dkk1 to activin-stimulated SuTOP-CFP cellscultured for 3 and 5 days. (A) Chimeric E10.5 embryo recovered after blastocyst injection of SuTOP-CFP EScells. Note CFP expression at sites known to harbor active Wnt signaling at this stage including the otic vesicle(white arrow), optic vesicle (white arrowhead), tail bud (red arrow), and somites and dorsal neural tube (redarrowhead). (B) SuTOP-CFP cells were cultured in the presence of 100 ng/ml activin for 3 days and supplementedwith 100 ng/ml Wnt3a or 320 ng/ml Dkk1 for a variable number of days as indicated. Scale bar is 80 µm. (C)SuTOP-CFP cells were cultured in the presence of 100 ng/ml activin for 5 days and supplemented with 100 ng/mlWnt3a or 320 ng/ml Dkk1 for a variable number of days as indicated in the figure. “–” indicates treatment withactivin alone. Scale bar is 200 µm.46