Persistence of transcript knockdown during maintenance in vitro.

Following exposure to target long double-stranded (ds)RNA (A, cathepsin B; B, cathepsin L; C, sigma-class glutathione transferase), newly-excysted juveniles (NEJs) were maintained in vitro for up to 25 days. NEJs were collected and assayed by quantitative PCR at 3, 9 and 25 days post dsRNA exposure. Target ΔΔCt (Y-axes) represents ratio of abundance of target transcript to a reference gene (glyceraldehyde phosphate dehydrogenase, GAPDH), in treated samples, relative to the abundance of those transcripts in untreated samples. Statistical significances are indicated relative to effects of negative control dsRNA (dsCTRL, complementary to neomycin phosphotransferase). dsCTRL treatments were performed in parallel with all experimental treatments. Experiments were repeated ≥3 times, employing 20–30 flukes per replicate. *, P<0.05; **, P<0.01; ***, P<0.001. Symbols represent mean±SEM.