The biphasic effects of Smad1 on EB-derived EryP potential extend to other hematopoietic progenitors. The miSmad1 EBs were left untreated or induced with doxycycline on day 2 or day 4, and day 5.75 EBs were harvested and recultured in semisolid methylcellulose media permissive for differentiation into (A) macrophage (MacP) colonies, (B) megakaryocyte (MegaP) colonies, (C) definitive mixed lineage colonies, or (D) EryD definitive erythroid colonies. After replating dissociated EBs in media containing the relevant permissive cytokines, MacP, MegaP, and EryD colonies were identified by their distinctive morphologies and counted on day 7; mixed lineage colonies were counted on day 9. For each sample, n = 3, and each graph is a representative example taken from a pool of at least 4 experiments. Error bars indicate SEM; *P < .01 compared with uninduced controls.