FIG 1.

Generation of the Ookluc line. (a) Strategy for generating the Ookluc reporter line. The coding sequence of nLuc (orange box) was cloned downstream the ctrp promoter (arrow) and upstream the 3′ UTR of the thrombospondin-related adhesive protein (TRAP; lollipop) into the plasmid pL0043 (19), containing the homology regions (HRs) for integration in the GIMO locus. Gray boxes indicate the resistant markers yeast fcu (yfcu) and human dihydrofolate reductase (hDHFR). (b) Ookinete conversion assay. Blood from an infected mouse is dispensed in a well of 96-well plate containing ookinete medium at 21°C and incubated for 24 h. Then, nLuc substrate is added, and the relative light units (RLU) were assessed by using a plate luminometer after a 5-min incubation. (c) Kinetics of nLuc activity, expressed as RLU (means + the SD), in Ookluc blood stages (blood) or different time points postactivation in conversion assay. The results are representative of 5 independent experiments. (d) Correlation between the number of ookinetes formed per μl of blood after 24 h conversion and the measured RLU. The RLU was measured without addition of lysis buffer, and the ookinetes were then counted by blood smears (for the image displayed: scale bar, 10 μm). Linear trendline and equation are shown. The results are representative of three independent experiments.