Analysis of apyrase 5' upstream region validates improved Anopheles gambiae transformation technique.
Analysis of apyrase 5' upstream region validates improved Anopheles gambiae transformation technique.
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DOI：
10.1186/1756-0500-2-24
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发表时间：
2009-02-19
影响因子：
1.8
通讯作者：
Arca, Bruno
中科院分区：
文献类型：
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作者：
Lombardo, Fabrizio;Lycett, Gareth J;Arca, Bruno
背景技术背景：近年来，冈比亚按蚊的遗传转化已经成功实现，这对研究人员来说是一个潜在的强大工具。组织，阶段和性别特异性启动子是必不可少的要求，以支持新的应用程序的转化技术和潜在的疟疾控制策略的发展。在无脊椎动物宿主中的疟原虫生命周期中，四种主要的蚊子细胞类型参与与寄生虫的相互作用：血细胞和脂肪体细胞，提供先天免疫反应的体液和细胞成分，中肠和唾液腺代表寄生虫在其生命周期中穿越的上皮屏障在mosquito.FINDINGS：我们已经分析了上游调控序列的An。冈比亚唾液腺特异性腺苷三磷酸双磷酸酶（AgApy）基因在转基因安. gambiae，使用由3xP 3启动子：DsRed基因融合体标记的piggyBac转座因子载体。有效的胚系转化在An。获得了冈比亚蚊子，并在至少三个不同的G 0家族中检测到几个整合事件。LacZ报告基因的表达进行了分析，在三个转基因株系/组，只有一个组是组织特异性表达限于唾液腺。结论：我们的数据描述了一个有效的遗传转化的安。冈比亚胚胎。然而，从所选区域的AgApy启动子的表达是弱的，位置效应可能掩盖转基因蚊子中的组织和阶段特异性活性。
BACKGROUND: Genetic 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.FINDINGS: We 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.CONCLUSION: Our 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.