Figure 3.

RNAi-mediated depletion of TbMLP has a delayed effect on growth of bloodstream-form parasites.A. Northern blot showing loss of the TbMLP transcript on induction of RNAi. Cultures of the parental cell line (2T1) and two clones transformed with the RNAi construct were split in two. One flask of each had tetracycline added (1 μg ml⁻¹, indicated by +). The other did not (indicated by −). After 24 h of induction, total RNA was extracted, blotted and hybridized with the TbMLP ORF. α-Tubulin mRNA is shown as loading control.B. Growth curves of RNAi clones in the presence (broken black line) or absence (solid black line) of tetracycline. Cells were seeded at 10⁵ ml⁻¹ and counted after 24 h. They were then diluted back to 10⁵ ml⁻¹ every 24 h to maintain exponential growth. For longer counting intervals, cells were diluted appropriately. Each clone was plated in triplicate and each well counted twice. P-values for the final time point are indicated to the right of the curves. The growth of the parental inducible cell line used to generate the RNAi transfectants is shown in grey (solid line without tetracycline, dashed line with tetracycline).