Transgenic engineering of Aedine mosquitoes

伊丁蚊的转基因工程

• 批准号: 6738965
• 负责人: Malcolm J. FRASER
• 金额: $ 40.95万
• 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 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6738965
• 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（由申请人提供）：蚊媒传染病 继续对许多人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 通过对这些基因和生理学有更深入的了解 昆虫。蚊子基因研究进展将大大加强 通过开发有效且可重复的转基因方法 操纵。这种能力的一个关键组成部分是识别和 操纵能够有效载体基因的昆虫转座子 进入蚊子基因组，允许基因标记，并保持基因处于活跃状态 状态。该领域研究的融合已取得成功 使用多个昆虫转座子对蚊子进行转基因操作，每个转座子 具有自己独特的属性。昆虫转座子piggyBac已被 被证明对多种昆虫的基因操作有效 物种，包括蚊子。最近，我们展示了改造 使用朱砂眼睛颜色标记基因的 PiggyBac 埃及伊蚊。这 提案寻求大量资金来了解和利用这些能力 用于建立和维持转基因菌株的piggyBac转座子 蚊子种类。其他基于piggyBac的转基因载体 含有受普遍存在的昆虫控制的理想基因 将构建、测试启动子或组织特异性蚊子启动子， 并与用于转基因的选择标记组合。存在和 转座子和载体基因的表达将在以下检测 在Gi代选择共转化标记基因，以及 使用多种技术进行分析，包括 Southern 杂交、PCR 扩增和测序、Northern 和 Western 分析，以及 免疫荧光，视情况而定。稳定性的遗传表征 并将检查蚊子基因组内转基因序列的移动 经过几代人的努力。这项研究将提供令人兴奋且多 显着加强基础研究工作所需的能力 蚊子遗传学和生理学。