(First stage)

DE induction; (3 days)

(Second stage)

PE induction (Pdx-1⁺ cells); (5 days)

All groups (A, B, C, D)

RPMI + BSA (0.1%v/v) + Activin A (100 ng/ml +wnt-3α (25 ng/ml)

Group A: DMEM-HG + FBS (2%v/v) + bFGF (64 ng/ml)

Grope B: DMEM-HG + FBS (2%v/v) + cyclopamine (0.25 µM)

Group C: DMEM-HG + FBS (2%v/v) + bFGF + cyclopamine

Group D (control): DMEM-HG + FBS (2%v/v)