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Fig. 3. Rescue of Zic3-attenuated notochord progenitor marker expression by co-injection of Siamois RNA. (A) Xnot expression in early gastrula. Vegetal views of the embryos with dorsal at the top. The embryos were injected with the RNAs indicated below (1000 pg each). Xnot expression was detected in the dorsal area adjacent to the yolk plug (purple signal) in the uninjected (uninj) embryo and the β-galactosidase (β-gal)-injected embryo, but disappeared in the Zic3 RNA (Zic3)-injected embryo. β-gal, Zic3, Bra, Sia and Nr3 mean the injected RNAs of β-galactosidase, Zic3, Bra, Siamois and Nr3, respectively. (B) Percentages of embryos with Xnot expression. Co-injection of Siamois fully restored normal Xnot expression. *P < 0.01; **P < 0.001; n.s., P > 0.05 in comparison to Zic3 alone injection in χ2 test. More than 50 embryos were analyzed in each group. |
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Fig. 1. Zic3 has a role in notochord development. EGFP RNA alone (A–C, J, L; EGFP, 400 pg) or EGFP RNA (400 pg) with Myc-tagged Zic3 RNA (D–F, M; Zic3, 200 pg) or Zic morpholino oligos (G–I, K, N; Z2MO + Z3MO, Zic2MO and Zic3MO, 30 ng each) were injected into the dorsal marginal zone of the two dorsal blastomeres of embryos at the 4-cell stage. The embryos were fixed at the tailbud stage (A–K, stage 33) or early neurula stage (L–N, stage 14). In panels A–I, expression of the posterior neural marker HoxB9 was analyzed by whole-mount in situ hybridization (WMISH; lateral view is shown in A, B, D, E, G, and H, and dorsal view is shown in C, F, and I). Panels J and K are hematoxylin-and-eosin-stained transverse sections of the embryos shown in panels B and H respectively (dashed lines indicate the section planes). n, notochord; hb, hindbrain; ot, otic vesicle. In panels L–N, expression of the notochord marker Xnot was analyzed by WMISH (n = 35–39). Images show dorsal views of the embryos. The indicated percentages of embryos had phenotypes similar to those shown in the images. Arrowheads, blastopores; long and short arrows, the rostrocaudal and left–right extent of Xnot expression. |
