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Fig. 6 Early Overexpression of you in Wild-type Embryos and Rescue of you Defects by shh mRNA Injection. (A–D) Dorsal views of the posterior trunk and tail bud of whole-mount embryos at 12 somites (15 hpf). (E–F) Lateral views of somites 2–7 in 24-hpf embryos. (G–H) Lateral views of somites 8–13 in 24-hpf embryos. (I–L) Lateral views of 24-hpf embryos. Anterior is to the left in all images. When 50 pg of you mRNA was injected into wild-type embryos at the 1–4-cell stage, no obvious expansion of myod (B), Engrailed (F), or nkx2.2 (J) expression was observed when compared either with wild-type embryos injected with equivalent amounts of mutant mRNA (A, E, and I) or with uninjected embryos (see Figure 1). Muscle pioneers were counted in a subset of the embryos; there were 4.0 ± 0.8 Engrailed-expressing muscle pioneers per somite in embryos injected with the control mRNA (n = 3 embryos, 33 somites) and 4.6 ± 1.1 muscle pioneers per somite in embryos injected with synthetic you mRNA (n = 8 embryos, 88 somites). Injection of 50 pg of shh mRNA into embryos at the 1–4-cell stage resulted in expansion of myod, Engrailed, and nkx2.2 expression in both wild-type (C, G, and K) and you mutant (D, H, and L) embryos. shh injection rescued adaxial expression of myod (D), muscle pioneer expression of Engrailed (H), and ventral spinal cord expression of nkx2.2 (L) in genotypically mutant you embryos (compare with Figure 1). Genotypes of all embryos were determined by PCR after photography.