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Fig. 4. Necab is downstream of Pax6. (A) X-gal staining showing Pax6 expression in the spinal cord, forebrain and optic vesicles. Immunofluorescence showing ß-galactosidase-positive cells in the optic vesicle of a Pax6 mutant embryo (Inset). (B-F) In situ hybridization showing that, in contrast to Otx2 (B), Six3 (C), Lhx2 (D) and Rx (E), Necab expression is absent in the optic vesicle and in the pretectum of Pax6 mutant embryos (arrowhead, F). Wild type, +/+; heterozygous Pax6 mutant, +/-; homozygous Pax6 mutant, -/-.