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. 2014 Jan 15;9(1):e84881. doi: 10.1371/journal.pone.0084881

Figure 6. The Small Molecule Method Can Produce IM Cells without the Mesendoderm Step.

Figure 6

(A) Induction of BRACHYURY+ mesendoderm cells generated from OSR1-GFP knock-in hiPSCs (3D45) after 24, 48, 72, and 96 hrs of treatment with CHIR99021 and TTNPB or activin A. (B) Results of the qRT-PCR analyses showing mRNA expression of the mesendoderm markers, BRACHYURY, GOOSECOID, and MIXL1, in differentiation cultures from the growth factor and TTNPB methods. (C) Effects of adding a RAR inhibitor, BMS493, to differentiation cultures of 3D45 cells treated with CHIR99021 and activin A, analyzed by anti-BRACHYURY immunostaining. (D) Induction of OSR1+ cells on culture day 11 of the growth factor method, with or without addition of BMS493 at various concentrations. (E) Results of qRT-PCR analyses showing expression of RARB mRNA. 3D45 cells on day 1, before treatment, were used to normalize the data shown in (B) and (E). The data in (A–E) are means±SD of three independent experiments (n = 3). Scale bars, 100 m.