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Updated: Jul 25, 2026
CRISPR/Cas9 Gene Editing to Make Conditional Mutants of Human Malaria Parasite P. falciparum
Published on: September 18, 2018
Complementation of Plasmodium berghei TRAP knockout parasites using human dihydrofolate reductase gene as a
A A Sultan1, V Thathy, T F de Koning-Ward
1Department of Pathology, Michael Heidelberger Division of Immunology, New York University School of Medicine, New York, NY 10016, USA. asultan@hsph.harvard.edu
Abstract:
Previously we have used the Plasmodium dihydrofolate reductase thymidylate synthase (DHFR-TS) selectable marker to generate Plasmodium berghei TRAP null mutant parasites. These TRAP null mutants do not glide and they showed a great reduction in their ability to infect mosquito salivary glands and the hepatocytes of the vertebrate host. Thus far, complementation of these knockout parasites was not possible due to the lack of additional selectable markers. Recently, a new selectable marker, based on the human dihydrofolate reductase (hDHFR) gene, has been developed which confers resistance to the antifolate drug WR99210. This drug has been found to be highly active against pyrimethamine-sensitive and -resistant strains of P. berghei. In this study, we have used the hDHFR gene as a second selectable marker for the complementation of P. berghei TRAP null mutant parasites. Restoration of the TRAP null mutant parasites to the wild-type phenotype was achieved in this study via autonomously replicating episomes bearing a wild-type copy of the TRAP gene. This is the first report of complementation of a mutant phenotype in malaria parasites.
