Fig. 5

mir92a Directly Targets nog3 mRNA In Vitro and In Vivo

(A) The two putative binding sites of mir92a in nog3 mRNA. The position of the last base of the stop codon was numbered 0.

(B and C) The expression of the reporter GFP-nog3-ed32UTR was inhibited by mir92a duplex. We injected 100 pg of the reporter mRNA at the one-cell stage alone or together with 1 nl of control miRNA duplex (Ctrl duplex), mir92a duplex, or mir92a duplex (20 µM) plus 4 ng 92a-MO, and injected embryos were observed for GFP fluorescence at 24 hpf (B). The GFP protein levels at 24 hpf were also examined by western blot with the anti-GFP antibody (C). Scale bar, 1 mm.

(D and E) The expression of EGFP-nog3-BS1 but not EGFP-nog3-mBS1 reporter was inhibited by injection of mir92a duplex. The embryos were similarly injected and observed (D) as in (B). Scale bar, 1 mm. The GFP expression levels at 24 hpf were also examined by western blot with the anti-GFP antibody (E).

(F and G) Alteration of nog3 mRNA levels in embryos injected with 4 ng 92a-MO or 10 µM 92a duplex (1 nl per embryo). Uninjected (UIC) or injected embryos at 42 (F) and 48 hpf (G) were probed using the nog3 antisense probe. Ventral views with anterior to the left. The ratio of embryos with altered expression to the total was indicated.

See also Fig. S5.