Stable germline transformation of the malaria mosquito Anopheles stephensi

Abstract

Anopheline mosquito species are obligatory vectors for human malaria, an infectious disease that affects hundreds of millions of people living in tropical and subtropical countries. The lack of a suitable gene transfer technology for these mosquitoes has hampered the molecular genetic analysis of their physiology, including the molecular interactions between the vector and the malaria parasite. Here we show that a transposon, based on the Minos element1 and bearing exogenous DNA, can integrate efficiently and stably into the germ line of the human malaria vector Anopheles stephensi , through a transposase-mediated process.

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Figure 1: The Minos element integrates into the A. stephensi genome.

References

  1. 1

    Franz, G. & Savakis, C. Minos, a new transposable element from Drosophila hydei, is a member of the Tc1-like family of transposons. Nucleic Acids Res. 19, 6646 (1991).

  2. 2

    Loukeris, T. G., Livadaras, I., Arca, B., Zabalou, S. & Savakis, C. Gene transfer into the medfly, Ceratitis capitata , with a Drosophila hydei transposable element. Science 270, 2002–2005 ( 1995).

  3. 3

    Coates, C. J., Jasinskiene, N., Miyashiro, L. & James, A. A. Mariner transposition and transformation of the yellow fever mosquito, Aedes aegypti. Proc. Natl Acad. Sci. USA 95, 3748–3751 (1998).

  4. 4

    Jasinskiene, N. et al. Stable transformation of the yellow fever mosquito, Aedes aegypti, with the Hermes element from the housefly. Proc. Natl Acad. Sci. USA 95, 3743– 3747 (1998).

  5. 5

    Berghammer, A. J., Klingler, M. & Wimmer, E. A. A universal marker for transgenic insects. Nature 402, 370–371 ( 1999).

  6. 6

    Catteruccia, F. et al. Toward Anopheles transformation: Minos element activity in anopheline cells and embryos. Proc. Natl Acad. Sci. USA 97, 2157–2162 ( 2000).

  7. 7

    Chalfie, M., Tu, Y., Euskirchen, G., Ward, W. W. & Prasher, D. C. Green fluorescent protein as a marker for gene expression. Science 263, 802–805 (1994).

  8. 8

    Cormack, B. P., Valdivia, R. H. & Falkow, S. FACS-optimized mutants of the green fluorescent protein (GFP). Gene 173, 33–38 (1996).

  9. 9

    Klinakis, A. G., Loukeris, T. G., Pavlopoulos, A. & Savakis, C. Mobility assays confirm the broad host range activity of the Minos transposable element and validate new transformation tools. Insect Mol. Biol. (in the press).

  10. 10

    Miller, L. H. et al. Stable integration and expression of a bacterial gene in the mosquito Anopheles gambiae. Science 237, 779–781 (1987).

  11. 11

    Loukeris, T. G., Arca, B., Livadaras, I., Dialektaki, G. & Savakis, C. Introduction of the transposable element Minos into the germ line of Drosophila melanogaster. Proc. Natl Acad. Sci. USA 92, 9485–9489 (1995).

  12. 12

    Pinkerton, A. C., Michel, K., O'Brochta, D. A. & Atkinson, P. W. Green fluorescent protein as a genetic marker in transgenic Aedes aegypti . Insect Mol. Biol. 9, 1– 10 (2000).

  13. 13

    Tamura, T. et al. Germline transformation of the silkworm Bombyx mori L. using a piggyBac transposon-derived vector. Nature Biotechnol. 18, 81–84 ( 2000).

  14. 14

    Kumar, V. & Collins, F. H. A technique for nucleic acid in situ hybridization to polytene chromosomes of mosquitoes in the Anopheles gambiae complex. Insect Mol. Biol. 3, 41–47 (1994).

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Acknowledgements

We thank A. Richman at EMBL for the initial efforts for anopheline transformation and D. Prager and the MacArthur Vector Biology Network for the collegial interactions that greatly stimulated the project. The research was supported by a Network grant of the training and Mobility Program of the European Community, by Implyx Ltd, by WHO-TDR, by the John D. and Catherine T. MacArthur Foundation, the SFB 544 of the Deutsche Forchungsgemeinschaft and an individual fellowship of the Biotechnology Program of the EU (T.G.L.)

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Correspondence to Andrea Crisanti.

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