Journal: Malaria Journal

Article Title: Selection of drug resistant mutants from random library of Plasmodium falciparum dihydrofolate reductase in Plasmodium berghei model

doi: 10.1186/1475-2875-10-119

Figure Lengend Snippet: Replacement strategy of mutant Pfdhfr-ts into Pbdhfr-ts by double cross-over homologous recombination . (A) endogenous Pbdhfr-ts gene locus in PbGFP wild-type parasite, (B) linearized plasmid pY005 containing mutant Pfdhfr S108N , (C) Pfdhfr S108N -ts gene locus in transgenic PbPf S108N parasite. Positions of primers used for PCR amplification are indicated by arrows. The expected size of PCR products are described.

Article Snippet: The merozoites were transfected with the circular plasmid DNA harboring Pfdhfr mutant libraries using the standard Amaxa Nucleofector protocol [ ] and re-infected into animals by i.v. injection.

Techniques: Mutagenesis, Homologous Recombination, Plasmid Preparation, Transgenic Assay, Amplification

Journal: Malaria Journal

Article Title: Selection of drug resistant mutants from random library of Plasmodium falciparum dihydrofolate reductase in Plasmodium berghei model

doi: 10.1186/1475-2875-10-119

Figure Lengend Snippet: PCR and RT-PCR analysis of the transgenic mutant parasite . (A) PCR analysis of 5' and 3' integrations on genomic DNA isolated from transgenic mutant parasites, PbPf S108N, are shown in lanes 1 and 4, respectively. Genomic DNA isolated from PbGFP wild-type parasite and distilled water (Neg) served as negative controls as shown in lanes 2, 5 and 3, 6, respectively. (B) RT-PCR analysis of PbPf S108N parasites. RNA from the PbPf S108N parasite was reverse transcribed to cDNA and used as a template to amplify Pfdhfr, Pbdhfr and P. berghei alpha tubulin ( Pbα-tubulin ) genes. The reactions were performed with reverse transcription (lane 1), without reverse transcription (lane 2), P. berghei cDNA derived from PbGFP and distilled water (Neg) were used as negative controls with and without reverse transcription (lanes 3-6).

Article Snippet: The merozoites were transfected with the circular plasmid DNA harboring Pfdhfr mutant libraries using the standard Amaxa Nucleofector protocol [ ] and re-infected into animals by i.v. injection.

Techniques: Reverse Transcription Polymerase Chain Reaction, Transgenic Assay, Mutagenesis, Isolation, Reverse Transcription, Derivative Assay