Transgenic engineering of Aedine mosquitoes

伊丁蚊的转基因工程

• 批准号: 6880028
• 负责人: Malcolm J. FRASER
• 金额: $ 42.87万
• 依托单位: UNIVERSITY OF NOTRE DAME
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-05-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6880028
• 关键词: Aedes; arthropod genetics; biotechnology; genetic manipulation; genetically modified animals; transfection /expression vector; transposon /insertion element

DESCRIPTION (provided by applicant): Mosquito-borne infectious diseases continue to have significant impact on the health and economy of much of the world. Alternative strategies for control of mosquito vectors could arise through a greater understanding of the genetics and physiology of these insects. Progress in genetic research with mosquitoes will be greatly enhanced by the development of effective and reproducible means of transgenic manipulations. A key component of this capability is the identification and manipulation of insect transposons that are able to efficiently vector genes into the mosquito genome, permit gene tagging, and maintain genes in an active state. The convergence of research in this field has resulted in the successful transgenic manipulation of mosquitoes with several insect transposons, each with its own unique properties. The insect transposon piggyBac has been demonstrated to be effective for genetic manipulation in widely diverse insect species, including mosquitoes. Recently, we demonstrated transformation of Aedes acgypti with piggyBac using the cinnabar eye coloration marker gene. This proposal seeks significant funding to understand and exploit the capabilities of the piggyBac transposon for establishing and maintaining transgenic strains of Aedine mosquito species. Additional piggyBac-based transgene vectors containing desirable genes under the control of either ubiquitous insect promoters or tissue-specific mosquito promoters will be constructed, tested, and combined with the selectable markers for transgenesis. The presence and expression of the transposon and vectored genes will be detected following selection for the co-transforming marker gene at the Gi generation, and analyzed using a variety of techniques including Southern hybridization, PCR amplification and sequencing, Northern and Western analyses, and immunoflourescence, as appropriate. Genetic characterization of the stability and movement of transgene sequences within the mosquito genome will be examined over several generations. This research will provide an exciting and much needed capability for significant enhancement of the basic research effort in mosquito genetics and physiology.

描述(申请人提供)：蚊媒传染病 继续对美国大部分地区的健康和经济产生重大影响 世界。可能会出现控制蚊媒的替代策略 通过对这些生物的遗传学和生理学有更深入的了解 昆虫。蚊子基因研究的进展将大大加强 通过开发有效和可重复性的转基因手段 操纵。这一能力的一个关键组成部分是识别和 操纵能够有效携带基因的昆虫转座子 进入蚊子基因组，允许基因标记，并保持基因处于活跃状态 州政府。这一领域的研究汇聚在一起，导致了 利用几个昆虫转座子对蚊子进行转基因操作 拥有自己独特的特性。昆虫转座子PiggyBac已经 证明对种类繁多的昆虫进行基因操作是有效的 物种，包括蚊子。最近，我们演示了 使用朱砂眼睛着色标记基因的acgypti伊蚊和猪Bac。这 一项提案寻求大量资金来了解和利用这些能力 利用猪转座子建立和保存转基因菌株的研究 伊丁蚊子的种类。额外的基于piggyBac的转基因载体 含有在无处不在的昆虫的控制下的可取的基因 启动子或组织特异性蚊子启动子将被构建、测试 并与转基因的可选择标记相结合。在场和 转座子和载体基因的表达将在下面检测到 GI世代共转化标记基因的选择，以及 使用了多种技术进行分析，包括Southern杂交、PCR 扩增和测序，Northern和Western分析，以及 免疫荧光法，视情况。稳定性的遗传特征 以及转基因序列在蚊子基因组中的移动将被检测 经过几代人的努力。这项研究将提供令人兴奋的和更多的 显著加强基础研究工作所需的能力 蚊子遗传学和生理学。