Figure 2.

(A) Map of the P. falciparum transfection vector pHD22Y expressing the highly MTX-resistant human L22Y dhfr allele under regulatory control of the P. falciparum elements 5′-hrp3 and 3′-hrp2 (28). (B) Detection of pHD22Y DNA replicated as episomes in transfected P. falciparum. T1+MTX genomic DNA (prepared 2 months posttransfection) and DNA from a representative plasmid rescued from T1+MTX were restricted with ScaI (S; which cuts once in the vector) or ScaI + DpnI (S+D; with over a dozen recognition sites) and hybridized with a probe corresponding to the pBluescriptII SK(+) vector backbone. Although P. falciparum-replicated episomes were resistant to DpnI digestion, digestion of E. coli-replicated plasmids resulted in detection of bands of the expected sizes 2.3, 1.0, 0.7, and 0.3 kb (the largest being fainter as the majority of this fragment contains 5′-hrp3 sequences).