FIG. 2.

Characterization of the recombinant P. berghei clones expressing transgenic Pbs21. (A) The non-gametocyte-producing transgenic clone Pb233/221 and the gametocyte-producing clone PbHP/221 contain the same copy numbers of pMD221. DNA was isolated from the recombinant clones digested with ClaI and prepared for Southern analysis. Hybridization with the DHFR-TS-specific probe reveals the genomic single copy of the DHFR-TS gene and the copy carried on the plasmid in the transfected clones. The copy number was estimated to be 2 to 6 per parasite nucleus. (B) Stage-specific expression of the transgenes in PbHP/221. Aliquots of synchronous and parallel cultures of PbHP/221 (B1) or wild-type HP (B2) were removed at the stages shown, RNA was prepared, and equal amounts were subjected to Northern analysis. The blots were simultaneously hybridized with complex probes representing the ORFs of both the DHFR and Pbs21 genes as indicated. The Pbs21 signal in HP wild-type parasites results from gametocytes present in the preparations. R, ring form; T, trophozoite; S, schizont. (C) Relative expression of the Pbs21 and DHFR genes in the different transgenic parasites. RNAs isolated from the same samples as those in panel A were subjected to Northern analysis. Hybridization was performed sequentially with probes for Pbs21 (top), DHFR-TS (middle), and rRNA (bottom). 221, pMD221 isolated from bacteria; HP, untransfected P. berghei ANKA; HP/221, clone PbHP/221; 233/221, clone Pb233/221; 233, untransfected clone 2.33; gam., gametocytes.