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Fig. 2 Zic2a restricts six3b expression throughout the forebrain beginning at mid-somitogenesis. (A,B) six3b is similarly expressed in conMO- and zic2aMO-injected embryos at 10s-12s (A, 41/41 embryos, n=2; B, 57/57 embryos, n=2). (C,D) At 12s-15s, six3b expression is normal in control morphants (black arrowheads in C, 38/38 embryos, n=3), and expanded throughout the forebrain in zic2a morphants (green arrowheads in D, 58/58 embryos, n=3). (E,F) six3b expansion in zic2aMO-injected embryos persists at prim-5 (black arrowheads in E, 42/42 control morphants, n=3; green arrowheads in F, 40/41 embryos, n=3). (G) Real-time PCR showed that six3b levels, normalized to gapdh, were increased (P=0.0001) in ZicMO-injected embryos relative to control morphants (based on three biological replicates, with two technical replicates per biological replicate). A-F are lateral views; eyes have been removed from embryos in C-F. A′-B′ are ventral views of the same embryos. C′-F′ are dorsal views of the same embryos, anterior to the left.