RNA Interference in Schistosoma mansoni Schistosomula: Selectivity, Sensitivity and Operation for Larger-Scale Screening (original) (raw)
Figure 6
RNAi of SmCC is accompanied by decreased translation product measured as proteolytic activity.
(A) Activity in extracts of schistosomula that had been co-incubated for 6 days with 30 µg/ml mCherry (Ch) or SmCC (CC) dsRNA was measured with the cathepsin C-selective peptidyl substrate, GR-AMC. Worm extracts were normalized to 0.4 µg of protein per reaction and the proteolytic activity in extracts from parasites exposed to mCherry dsRNA was set to 100%. (B) Reactivity with the fluorescent cathepsin C-selective activity-based probe (bodipy-labeled FY01; [91]) as assessed by SDS-PAGE. Worm extracts were normalized to 2 µg per labeling reaction. The fluorescent signal at 24 kDa is lost subsequent to exposure to CC-specific dsRNA. Data in (A) are the average of triplicate values (SD values were less the 5% of the mean). Representative data from two experiments are shown.