Fig. 2. Specific effect of antisense Vg1 RBP oligonucleotide on Xenopus development. Two-cell stage embryos were injected in both blastomeres with equal amounts of CMO (B) or AMO (D,E) and allowed to develop to the tailbud stage. Embryos injected with CMO were identical to uninjected sibling embryos (A). Note the abnormal head development, lack of dorsal fin, curved neural tube, and severe reduction in normal pigmentation in embryos injected with AMO. (C,F) Close-up views of the head regions of B and E, respectively, show defective lens formation in the AMO-injected embryos (F). Note the presence, in these embryos, of a pigmented retinal epithelium below the undifferentiated, overlying ectoderm. (G) Sense Vg1 RBP mRNA can rescue AMO-injected embryos. Both blastomeres of two-cell stage embryos were co-injected with AMO and sense Vg1 RBP-GFP mRNA, lacking the 5' UTR that contains the AMO target sequence. Embryos were allowed to develop until tailbud stage. Note the rescue of lens and dorsal fin formation and of melanophore migration. (H) Overexpression of Vg1 RBP-GFP mRNA alone does not affect the normal development of the embryo. (I) Vg1 RBP translation is reduced specifically by AMO injection. Proteins were extracted from tailbud-stage, uninjected (control) embryos, and from embryos injected with either CMO (CMO) or AMO (AMO). Two embryo-equivalents were loaded in each lane and Vg1 RBP expression was analyzed by electrophoresis and western blot analysis using an anti-Vg1 RBP antibody. As measured by densitometry, Vg1 RBP levels are reduced to 20% of original levels in the AMO-injected embryos relative to both uninjected and CMO-injected embryos; samples were normalized to the internal ERK-2 control (detected by anti-ERK-2 antibody).

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