Figure 2.

siRNA-mediated inhibition of miRNA biogenesis and disruption of tadpole development. (A) Impairment of miR-427 biogenesis by siRNAs. Accumulation of endogenous X. laevis miRNAs in untreated embryos (Non-inj.) or embryos injected with siRNA_NS at the one-cell stage was monitored over time by Northern blot analyses of total RNAs (one embryo equivalent of total RNAs per lane). Individual blots were probed for miR-427, miRNA-19b, or miRNA-16, as indicated. (B) PhosphorImager quantification of the miR-427 blots shown in A. (C) Dosage-dependent inhibition of pre-miRNA processing. ³²P-labeled pre-miR-427 RNAs were injected into one- to two-cell embryos alone (left and right panels) or together with different amounts of siRNA_NS, and Dicer processing was monitored over time post-injection by denaturing PAGE of total RNAs (one embryo equivalent per lane). (D) Abnormal development of tadpoles derived from siRNA-injected embryos. One-cell embryos were injected with 100 fmol of siRNA_NS or H₂O, and their development was monitored by visual inspection at stage 42. Images of tadpoles developed from noninjected embryos are shown in Supplemental Fig. S1.