Fig. 2. Single-molecule analysis of the dsRNA cleavage reaction by Dcr-2 tethered on the glass.

a Schematic representation of the Dcr-2-anchored single-molecule observation. b–e Representative single-molecule images of surface-tethered Dcr-2 and co-localized dsRNAs (wild-type and BLT [b n = 9 independent experiments], wild-type and 3′ovr [c, n = 20 independent experiments], the G31R helicase mutant and BLT [d n = 2 independent experiments], and the G31R helicase mutant and 3′ovr [e, n = 2 independent experiments]). Co-localized spots were indicated by yellow circles. dsRNAs were frequently co-localized with wild-type Dcr-2, but rarely with the helicase mutant. Scale bar, 2 μm; inset. f–h Representative traces of the “3-steps” (f), “2-steps” (g), and “1-step” (h) events. The x axis shows the time after starting the observation. i Representative trace of 3× Cy3-labeled BLT dsRNAs co-localized with the RNase III mutant. Three repetitive PIFE events were observed without cleavage. j Binding rate of dsRNAs, calculated from the total number of binding events (3-steps, 2-steps, and 1-step). BLT dsRNAs bind Dcr-2 more frequently than 3′ovr dsRNAs. Loqs-PD enhances the binding frequency for both BLT and 3′ovr dsRNAs. Source data are provided as a Source Data file.