Abstract
BackgroundGenetic transformation of the malaria mosquito Anopheles gambiae has been successfully achieved in recent years, and represents a potentially powerful tool for researchers. Tissue-, stage- and sex-specific promoters are essential requirements to support the development of new applications for the transformation technique and potential malaria control strategies. During the Plasmodium lifecycle in the invertebrate host, four major mosquito cell types are involved in interactions with the parasite: hemocytes and fat body cells, which provide humoral and cellular components of the innate immune response, midgut and salivary glands representing the epithelial barriers traversed by the parasite during its lifecycle in the mosquito.FindingsWe have analyzed the upstream regulatory sequence of the An. gambiae salivary gland-specific apyrase (AgApy) gene in transgenic An. gambiae using a piggyBac transposable element vector marked by a 3xP3 promoter:DsRed gene fusion. Efficient germ-line transformation in An. gambiae mosquitoes was obtained and several integration events in at least three different G0 families were detected. LacZ reporter gene expression was analyzed in three transgenic lines/groups, and in only one group was tissue-specific expression restricted to salivary glands.ConclusionOur data describe an efficient genetic transformation of An. gambiae embryos. However, expression from the selected region of the AgApy promoter is weak and position effects may mask tissue- and stage- specific activity in transgenic mosquitoes.
Highlights
Genetic transformation of the malaria mosquito Anopheles gambiae has been successfully achieved in recent years, and represents a potentially powerful tool for researchers
We reported that a short region (~800 bp) from the An. gambiae An. gambiae salivary glandspecific apyrase (AgApy) promoter was able to drive stageand tissue-specific expression of the reporter gene in transgenic An. stephensi
We describe here the in vivo analysis of a large 5' regulatory region taken from the AgApy gene of An. gambiae
Summary
As far as the main African malaria vector An. gambiae is concerned, after the initial successful transformation [8] only one additional study with transgenic An. gambiae has been reported so far [9]. In both cases, low transformation efficiencies were observed. It was clear that the putative An. gambiae promoter needed to be examined directly in this species before more firm conclusions could be drawn In this context, we should mention that while this work was in progress, robust salivary gland specific expression of a reporter gene in An. stephensi was reported using a promoter fragment from the An. stephensi aapp gene [14]. All the reactions were performed at least twice using different batches of RNA preparations
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