Fig. 4. LvAlx1 is required in mesomere descendants for transfating. (A) Experimental protocol. (B-G) Embryos were immunostained with mAb 6a9 and the same embryos were photographed using brightfield (B-D) and epifluorescence (E-G) optics. (B,E) Animal cap isolated at the 16-cell stage and cultured for 24 hours in the absence of LiCl. The embryo lacks mesoderm and endoderm and no 6a9-positive cells are present. (C,F) Animal cap treated with 50 mM LiCl for 3 hours after the operation and cultured for 24 hours. Endoderm and mesoderm have formed, including many 6a9-positive cells. (D,G) Animal cap isolated from a LvAlx1 MO-injected embryo, treated with 50 mM LiCl, and cultured for 48 hours. Although a gut (arrow) has formed, no 6a9-positive cells are present. (H) RT-PCR experiments showing that animal caps treated with 50 mM LiCl express Lvalx1 and one of its downstream targets (Lvp16). Each experiment was repeated three times (R1-R3).