Fig. 3

Loss of function phenotypes for the tcf3 gene in medaka. Embryos at the 1-cell stage were co-injected with 300 μM morpholino oligonucleotide (Tcf3MO) and 1 μg/mL FITC-dextran. The pairs b and f, c and g, d and h each show the same embryo at different stages (stage 22, a-d; stage 30, e-h). Control embryos (a,e) were co-injected with 1× Yamamoto’s and FITC-dextran. Pictures of embryos with a fluorescent signal were taken at the indicated stages. Classification was performed after the onset of eye pigmentation, according to the size of the eye: (b,f) weak, slightly reduced size; (c,g) moderate, severe size reduction; (d,h) strong, eye-less. Embryos at stage 16 (i-k, n-o) are shown in dorsal view, anterior to the top; except for (l,p) which are at stage 17 and shown in lateral view. For genotypic analysis (i-p), whole mount in situ hybridization experiments were performed on MO injected embryos and un-injected wild type controls at stage 16 using digoxigenin-labelled RNA probes against pax6 (i,m), pax2 (j,n), gbx1 (k,o), and wnt1 (l,p) at stage 17. (i-k, m-o) Broken lines indicate the outlines of the prospective neural domain estimated from the merged expression patterns formed by pax6, pax2 and gbx1 in wild type embryos (i-k). An anterior shift of the expression domains was observed for all four genes. (l,p; arrowhead) wnt1 expression in the mid-hindbrain boundary. Scale bars 100 μm; a, a-h; i, i-k and m-o; l, l and p. Abbreviations: MO, morpholino oligonucleotide