Generation of the NF54luctransgenic line. (A) Schematic representing the bxb I-integrase (Int) mediated integration of the Pfpcna-luc plasmid into the cg6-attB locus in NF54attB. BSD and Neo represent the drug selection markers for blasticidin S and neomycin, respectively. The position and orientation of oligonucleotide primers (P1 to P3) used to confirm integration are shown. (B) PCR monitoring of integration of Pfpcna-luc plasmid. The top panel shows results of PCR across the cg6-attB locus (primers P1 and P2), and the lower panel, PCR across the attL junction (primers P1 and P3) resulting from successful integration in NF54luc. Dd2luc is included as control. In the case of Dd2luc, no product is expected across the cg6-attB locus as this is disrupted with the hdhfr drug selection cassette. (C) Stage-specific northern blot using probes to Pfpcna and luc transcripts. The expected sizes of the predominant transcripts of 1.8Kb and 2.6Kb, respectively, were found. T1 to T5 represent timepoints of sampling from early rings, late rings, early trophozoites, late trophozoites and schizonts, respectively. (D) Time-course of luciferase sampling of NF54luc (2% parasitaemia, 2% HCT) over T1–T5. The bars represent mean RLU ± stdev (n = 5) using the standard luciferase substrate.