wnt9a, frzb and fzd7a are required for bapx1 expression. Dorsal views of embryo at 10-somites (A′–P′) and lateral views of embryo at 24 hpf (A–L) and 55 hpf (M–T). Wholemount RNA in situ hybridization for sox10 (A′–D′), sox9a (E′–H′), foxd3 (I′–L′) and dlx2a (M′–P′) in wildtype, wnt9a, frzb and fzd7a morphants; edn1 (A–D), endrA2 (E–H), dlx2a (I–L) sox9a (M–P) and bapx1 (Q–T) in wildtype, wnt9a, frzb and fzd7a morphants. Quantitative PCR for edn1, ednrA2, dlx2a in wildtype, wnt9a, frzb and fzd7a morphants at 24 hpf (U–W) and bapx1 at 55 hpf (X). Knockdown of wnt9a, frzb and fzd7a did not alter neural crest marker expression early in development (A′–P′). Similarly, edn1, ednrA2 and dlx2a expression (A–L) was unaffected in all morphants. Quantitative PCR analysis of whole embryos further confirms normal transcript levels at 24 hpf (U–W). Expression patterns of sox9a at 55 hpf in wnt9a, frzb and fzd7a morphants confirm the presence of CNCC mesenchyme within the first and second pharyngeal arches (M–P). However, the expression of bapx1 is reduced both qualitatively and quantitatively in wnt9a, frzb and fzd7a morphants at 55 hpf (Q–T, X).
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