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Science 2 June 1995:
Vol. 268. no. 5215, pp. 1358 - 1362
DOI: 10.1126/science.7761856
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Articles

Science, Vol 268, Issue 5215, 1358-1362
Copyright © 1995 by American Association for the Advancement of Science

articles

Stable transfection of malaria parasite blood stages

MR van Dijk, AP Waters, and CJ Janse

Department of Parasitology, University of Leiden, Netherlands.

Genetic manipulation of malaria parasites would revolutionize the study of this group of pathogens and have implications for vaccine and drug development. This report describes the stable, drug-selectable genetic transformation of the clinically relevant intracellular blood stages of a malaria parasite. A plasmid transfection vector carrying the gene locus that encodes a drug-resistant form of the bifunctional enzyme dihydrofolate reductase-thymidylate synthase from the rodent malaria parasite Plasmodium berghei was constructed. Derivatives of this vector were introduced into merozoites of P. berghei by electroporation, and parasites were selected for successful transformation in the rodent host on the basis of resistance to pyrimethamine. The plasmids were present in a circular, unrearranged form that replicated episomally to an observed maximum of 15 copies per cell in drug-resistant populations.


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Plasmodium knowlesi Provides a Rapid In Vitro and In Vivo Transfection System That Enables Double-Crossover Gene Knockout Studies.
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Infect. Immun. 70, 655-660
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Topology and replication of a nuclear episomal plasmid in the rodent malaria Plasmodium berghei.
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Physiol Rev 81, 495-537
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PNAS 96, 8716-8720
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Molecular Basis of In Vivo Resistance to Sulfadoxine-Pyrimethamine in African Adult Patients Infected with Plasmodium falciparum Malaria Parasites.
L. K. Basco, R. Tahar, and P. Ringwald (1998)
Antimicrob. Agents Chemother. 42, 1811-1814
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Precise Timing of Expression of a Plasmodium falciparum-derived Transgene in Plasmodium berghei Is a Critical Determinant of Subsequent Subcellular Localization.
C. H. M. Kocken, A. M. van der Wel, M. A. Dubbeld, D. L. Narum, F. M. van de Rijke, G.-J. van Gemert, X. van der Linde, L. H. Bannister, C. Janse, A. P. Waters, et al. (1998)
J. Biol. Chem. 273, 15119-15124
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Transformation with human dihydrofolate reductase renders malaria parasites insensitive to WR99210 but does not affect the intrinsic activity of proguanil.
D. A. Fidock and T. E. Wellems (1997)
PNAS 94, 10931-10936
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A novel alternate secretory pathway for the export of Plasmodium proteins into the host erythrocyte.
M. F. Wiser, H. N. Lanners, R. A. Bafford, and J. M. Favaloro (1997)
PNAS 94, 9108-9113
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Transfection of the Primate Malaria Parasite Plasmodium knowlesi Using Entirely Heterologous Constructs.
A. M. v. d. Wel, A. M. Tomas, C. H.M. Kocken, P. Malhotra, C. J. Janse, A. P. Waters, and A. W. Thomas (1997)
J. Exp. Med. 185, 1499-1504
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Antifolate-resistant mutants of Plasmodium falciparum dihydrofolate reductase.
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Functional Equivalence of Structurally Distinct Ribosomes in the Malaria Parasite, Plasmodium berghei.
R. M. L. van Spaendonk, J. Ramesar, A. van Wigcheren, W. Eling, A. L. Beetsma, G.-J. van Gemert, J. Hooghof, C. J. Janse, and A. P. Waters (2001)
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