Obtaining the stably transfected <i>T. vivax TvLrDNA-Luc</i> strain.

<p>(A). <i>T. vivax</i> axenic epimastigotes were transfected with circular or <i>AfeI</i> linearized p<i>TvL</i>rDNA-Luc using the ×006 Amaxa program and luciferase light emission (RLU) was measured 24 h post-transfection in the supernatants. (B) Schematic representation of p<i>Tv</i>LrDNA-Luc linearized with <i>AfeI</i> and its expected recombination in the parasite ribosomal DNA region. Hatched boxes show ribosomal promoter-containing regions; Black crosses represent recombination events; grey arrows indicate primers used to verify plasmid integration into <i>T. vivax</i> genome. 18 S, 5.8 S and 28 S, rRNA genes. (C) Verification of the 5′- and 3′- integration of the vector. PCR using respectively upF/upR (upper panel) and downF/downR (lower panel) primers. PCR was performed on genomic DNA from 2 independent cultures stably transfected with p<i>TvL</i>rDNA-Luc (1, 2) or from WT strain (3). (D) Changes in luminescence (RLU) emitted per p<i>TvL</i>rDNA-Luc parasite after G418 selection. (E) Graphic representation of bioluminescence expressed as luciferase light emission (RLU) of increasing numbers of <i>TvLr</i>DNA-luc parasites.