Contrasting Inducible Knockdown of the Auxiliary PTEX Component PTEX88 in P. falciparum and P. berghei Unmasks a Role in Parasite Virulence
Fig 1
Generation of a PTEX88 knockdown line in Plasmodium falciparum.
A: Schematic of targeting construct designed to integrate into the endogenous ptex88 locus by single crossover recombination. Arrows indicate binding sites for diagnostic PCR primers. SM, selectable marker; HA, haemaglutinin epitope tag; glmS, glmS ribozyme. B: Diagnostic PCR on wildtype (WT) and transgenic genomic DNA using the indicated primer combinations to test for integration of the targeting construct. Absence of a product using primer combination A/B in PTEX88-HA and PTEX88-glmS gDNA indicates these parasite lines are clonal. C: Representative Western blot showing levels of tagged PTEX88 in PTEX88-HA control parasites and PTEX88-glmS parasites after incubation with the indicated concentration of glucosamine. EXP2 was used as a loading control. GlcN, glucosamine. D: Densitometry performed on Western blots to quantify the protein levels of tagged PTEX88 in the control PTEX88-HA line and the PTEX88-glmS line. The protein level of PTEX88 does not decrease in the control PTEX88-HA line but decreases in the PTEX88-glmS with increasing GlcN concentrations by up to 90% when compared to the sample without GlcN (n = 4).