Expression of a plasmodium gene introduced into subtelomeric regions of Plasmodium berghei chromosomes

Science

Volumn 271, Issue 5249, 1996, Pages 662-665

  Van Dijk, Melissa R ^a   Janse, Chris J ^a   Waters, Andrew P ^a  

^a LEIDEN UNIVERSITY   (Netherlands)

Author keywords

[No Author keywords available]

Indexed keywords

DNA;

ARTICLE; CONTROLLED STUDY; GENE DISRUPTION; GENE EXPRESSION; GENE INSERTION; GENE MUTATION; GENETIC TRANSCRIPTION; MARKER GENE; NONHUMAN; PHENOTYPE; PLASMODIUM BERGHEI; PRIORITY JOURNAL; TELOMERE;

PLASMODIUM BERGHEI; RODENTIA;

EID: 0030049909     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.271.5249.662     Document Type: Article

References (20)

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7. Integration vector pMD223 was described previously (2) On digestion of P. berghei genomic DNA with Hind III, the following relevant fragments are generated; 2.3-kb-sized copies of the 2 3-kb repeat and 5- and 12-kb fragments of the genomic DHFR-TS locus. Insertion of linear pMD223 in the tandem repeat cluster leads to the introduction of a second copy of the genomic 5-kb Hind III DHFR-TS fragment and generation of a new fragment of 3 ko that will hybridize to the probe specific for the DHFR-TS locus. The hybridization to the 5-kb fragment therefore appears more intense. In addition, Hind III digestion released another band of 5 kb, consisting largely of pBSKS vector sequences (Fig. 2A). Circularization of Hind III-digested genomic DNA containing pMD223 integrated by homologous recombination into a genomic array of 2 3-kb repeats will result in the formation of circular DNA elements that can be rescued by transformation back into E. coli. Digestion of rescued plasmid with Hind III and Eco RI demonstrates the rescue of the Hind III site adjacent to the site of integration and distinguishes between integrated and episomally maintained vector because the latter contains an additional 1 2-kb fragment (Fig 2B). Sequence analysis confirmed the rescue of the Hind III site, which delineates the termini of the 2.3-kb repeat (5).
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19. During the course of this work we became aware of similar work with asexual bloodstage forms of the human malaria parasite, P. falciparum Plasmid DNA has been successfully introduced into the P. falciparum genome in a site-specific manner with DHFR-TS genes from either P. falciparum or Toxoplasma gondil. A manuscript describing this work is now in press (Y. Wu, L. A. Kirkman, T. E. Wellems, Proc. Natl. Acad Sci U S A., in press
20. We thank J. Ramesar and J. Kos for technical assistance, the yeast genetics group in the Institute of Molecular Plant Sciences at Leiden for consideration of our work, R Vinkenoog and R E. Sinden for discussions, G van Dam for curve fitting, and H A del Portillo for critical reading of the manuscript. This work was supported by grants from the European Union program "Science and Technology for Development" (contract TS3*-CT92-0116) and the United Nations Development Program-World Bank-World Health Organization Special Programme in Research and Training in Tropical Diseases (contract 940351)