(A–H) In situ hybridization (ISH) and (I) quantitative, real-time PCR (qPCR) analysis of Wnt target gene expression; (A, C, E–H) lateral views, anterior to the left; (B, D) transverse sections through arch 2. (A, B) mycn mRNA is ventrally restricted (black arrowheads) in wild type (WT) embryos. Arches 1 and 2 are outlined by dotted lines. (C, D) GFP mRNA is ventrally restricted (arrowheads) in Tg(7xTCF-Xla.Siam:GFP)ia4 transgenics. (B, D) Both mycn (B) and 7xTCF;GFP (D) are expressed in ventral nc cells and pharyngeal endoderm, and excluded from pharyngeal ectoderm. (E–H) mycn and axin2 expression in controls (E, G), and dntcf3+ embryos at 26 hpf (F, H; heat shocked at 22 hpf). (I) qPCR analysis of axin2, lef1 and mycn expression in dntcf3+ embryos, normalized to nontransgenic, heat-shocked controls, with ef1alpha as an internal control. * P<0.05, ** P<0.001. Abbreviations: e, eye; D, dorsal arch; end, pharyngeal endoderm; hb, hindbrain; I, intermediate arch; mhb, mid-hindbrain boundary; nc, neural crest; pe, pharyngeal ectoderm; V, ventral arch. Scale bars: 100 µm.