Effect of antisense oligo injections on expression of hh genes and on rod recruitment. (A) Cryosection of a PTU-treated embryo injected with a nonsense oligo at 54 hpf, fixed at 56 hpf, and hybridized with shh cRNA. shh is expressed in the RPE (arrow), CNS (large arrowhead), and foregut (small arrowhead). Scale bar (applies to A, B, E, F, and G), 50 μm. (B) Cryosection of an embryo injected with the antisense shh/twhh oligo cocktail, hybridized with shh. shh is expressed in the CNS (large arrowhead) and foregut (small arrowhead), but not in the RPE. (C) Cryosection of a nonsense-injected embryo, labeled with the polyclonal anti-Hh antibody. Hedgehog immunoreactivity is present in the CNS (arrowhead), foregut (small arrowhead), and RPE. Scale bar (applies to C and D), 50 μm. (D) Cryosection of an antisense-injected embryo, labeled with the polyclonal anti-Hh antibody. Hedgehog immunoreactivity is still present in the CNS (arrowhead), but is virtually undetectable in the RPE (arrow indicates position of RPE) and is reduced in the foregut. (E) Eye from a nonsense oligo-injected embryo, fixed at 73 hpf and hybridized with GFrod, lateral view. A dense patch of opsin-expressing rods is present in ventral retina (bottom of photo), and many additional rods have begun to express opsin elsewhere in the eye. (F and G) Eyes from antisense oligo-injected embryos, hybridized with GFrod. In F, there is a dense ventral patch of opsin-expressing rods, but fewer rods are present elsewhere. In G, the ventral patch is considerably smaller and contains fewer rods, and no rods are seen elsewhere in the eye; the eye is also reduced in size compared to C.
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