Fig 2. Conditional complementation and mutagenesis of PfMyoA.

A RAP treatment of PfMyoA-comp and PfMyoA-K764E show that the complementing line has no growth defect, while the RAP-treated K764E line grows at around 55% of DMSO-treated control per cycle under static conditions. B Genotyping PCR of the endogenous Pfmyoa locus in PfMyoA-comp and PfMyoA-K764E after RAP treatment shows that the replacement of the unexcised, integrated allele (IN) with the excised allele (EX) is almost complete. C Western blot of WT, PfMyoA-cKO, PfMyoA-comp and PfMyoA-K764E schizonts confirms that PfMyoA-FLAG, expressed from the endogenous Pfmyoa locus, is almost completely lost in favour of truncated PfMyoA-GFP. PfMyoA-3xHA, expressed at the ectopic locus, is unaffected by RAP treatment. D Parasites were treated with DMSO or RAP and cultured under static or suspension conditions. Parasitaemia was then measured in the following cycle by flow cytometry and normalised to DMSO control for each line and condition, showing that suspension conditions partially alleviate the growth defect caused by K764E mutation, from 67% of DMSO-treated to 77%. Bars show mean parasitaemia, N = 4 (or 2 for PfELC-cKO, tested separately), each experiment in triplicate. Significance assessed by paired t test, two tailed.