Dicing Assays.

Processing of long dsRNA into siRNAs was analyzed with an ultra-active form of recombinant human Dicer, capable of cleaving more than 95% of dsRNA template into siRNAs within 2 h (Genlantis), in an in vitro cleavage assay. In brief, ³²P-labeled dsRNA (700 bp, 30 ng) and recombinant human Dicer (0.5 U) were incubated in the buffer solutions provided by the manufacturer for 2 h at 37 °C, and then analyzed on a 15% PAGE gel. The indicated concentrations of recombinant His-YFC protein, His-NS1 protein (Bioclone), 6× His-tag peptide (Abcam), or recombinant GFP (Kerafast) were coincubated in the dicing assay.