Abstract
Malaria is a disease that kills more than a million people each year in tropical and subtropical countries. The disease is caused by Plasmodium parasites and is transmitted to humans exclusively by mosquitoes of the genus Anopheles. The lack of functional approaches has hampered study of the biological networks that determine parasite transmission by the insect vector. The recent discovery of RNA interference and its adaptation to mosquitoes is now providing crucial tools for the dissection of vector–parasite interactions and for the analysis of aspects of mosquito biology influencing the vectorial capacity. Two RNAi approaches have been established in mosquitoes: transient gene silencing by direct injection of double-stranded RNA, and stable expression of hairpin RNAs from transgenes integrated in the genome. Here we describe these methods in detail, providing information about their use and limitations.
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References
van Dijk, M. R., Waters, A. P., and Janse, C. J. (1995) Stable transfection of malaria parasite blood stages Science 268, 1358–1362.
Blandin, S., Moita, L. F., Köcher, T., Wilm, M., Kafatos, F. C., and Levashina, E. A. (2002) Reverse genetics in the mosquito Anopheles gambiae: targeted disruption of the Defensin gene EMBO Rep 3, 852–856.
Handler, A. M., and Harrell, R. A., 2nd (1999) Germline transformation of Drosophila melanogaster with the piggyBac transposon vector Insect Mol Biol 8, 449–457.
Brown, A. E., Bugeon, L., Crisanti, A., and Catteruccia, F.(2003) Stable and heritable gene silencing in the malaria vector Anopheles stephensi Nucleic Acids Res 31, e85.
Brown, A. E., Crisanti, A., and Catteruccia, F.(2003) Comparative analysis of DNA vectors at mediating RNAi in Anopheles mosquito cells and larvae J Exp Biol 206, 1817–1823.
Henschel, A., Buchholz, F., and Habermann, B.(2004) DEQOR: a web-based tool for the design and quality control of siRNAs Nucleic Acids Res 32, W113–W120.
Catteruccia, F., Nolan, T., Loukeris, T. G., Blass, C., Savakis, C., Kafatos, F. C., and Crisanti, A.(2000) Stable germline transformation of the malaria mosquito Anopheles stephensi Nature 405, 959–962.
Lobo, N. F., Clayton, J. R., Fraser, M. J., Kafatos, F. C., and Collins, F. H.(2006) High efficiency germ-line transformation of mosquitoes Nat Protoc 1, 1312–1317.
Frolet, C., Thoma, M., Blandin, S., Hoffmann, J. A., and Levashina, E. A. (2006) Boosting NF-kappaB-dependent basal immunity of Anopheles gambiae aborts development of Plasmodium berghei. Immunity 25, 677–685.
Acknowledgments
The authors first developed these methodologies while working at the EMBL in the group of Professor Fotis C. Kafatos (dsRNA injections) and in the laboratory of Professor Andrea Crisanti at Imperial College London (transgenesis). They further acknowledge members of the London and Strasbourg laboratories for constructive discussions. This work was supported by grants from CNRS, Inserm, Wellcome Trust, BBSRC, Schlumberger Foundation For Education and Research (FSER), and by the 6th European Commission Programme “Networks of Excellence” BioMalPar. F.C. is a MRC Career Development fellow. E.A.L. is an International Scholar of the Howard Hughes Medical Institute.
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© 2009 Humana Press, a part of Springer Science+Business Media, LLC
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Catteruccia, F., Levashina, E.A. (2009). RNAi in the Malaria Vector, Anopheles gambiae . In: Rondinone, C., Reidhaar-Olson, J. (eds) Therapeutic Applications of RNAi. Methods in Molecular Biology™, vol 555. Humana Press. https://doi.org/10.1007/978-1-60327-295-7_5
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DOI: https://doi.org/10.1007/978-1-60327-295-7_5
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