FIGURE 3. Generation of pb22 knockout and restoration parasites.
A. Schematic representation of the WT pb22 locus, the transfection construct, the recombined locus and reconstructed locus. The construct contains a dhfr expression cassette, directed by the pbeef1aa promoter, for selection of the pb22 knockout (KO) parasites with pyrimethamine and yfcu expression cassette for selection of the pb22-restored (RE) parasites with 5-fluorocytosine. Primers used to detect pb22 deletion and restoration are marked. B. Diagnostic PCR of WT, KO and RE parasites. Lane 1, primers 1 + 2 (1449 bp); Lane 2: primer 1 + 3 (1180 bp).