Fig. 7

PT enhancer with 3′UTR of hes6 recapitulated a posterior-to-anterior gradient of hes6 expression within the PSM. (A) EGFP reporter genes were used to generate the transgenic fish lines, Tg(hes6:EGFP-hes6 3′UTR-SV40pA). To generate the reporter gene used in Tg(hes6:EGFP-hes6 3′UTR-SV40pA), a DNA fragment corresponding to hes6 3′UTR was inserted between EGFP and SV40pA in the reporter gene (shown in Fig. 2A). (B–J) Two-color fluorescent in situ hybridization. Tg(hes6:EGFP-SV40pA), Tg(hes6:EGFP-hes6 3′UTR), and Tg(hes6:EGFP-hes6 3′UTR-SV40pA ) embryos were fixed at the 10–12-somite stage. Confocal images were taken of flat-mounted embryos after yolk removal. Endogenous expression of hes6 is shown by magenta, and expression of EGFP mRNA is shown by green. Antisense probe for hes6 is specific to the open reading frames of hes6. Scale bar in J, 50 µm. (K–R) Expression of EGFP-hes6 3′UTR mRNA was observed in transgenic embryos after treatment with SU5402 and DEAB; expression was similar to the endogenous expression of hes6 mRNA. (K–N) Transient treatment of Tg(hes6:EGFP-hes6 3′UTR) embryos at the 1–2-somite stage with SU5402. EGFP expression was analyzed. (O–R) Treatment of Tg(hes6:EGFP-hes6 3′UTR) embryos from the sphere stage with DEAB. Embryos were fixed at the 10–12-somite stage and EGFP expression was examined. Dorsal views (K–R). Probes used for staining are shown at top-right of each panel. All images were taken at the same magnification. Scale bar in R is 200 µm.