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转基因玉米褐飞虱 Peregrinus maidis 的表达四环素反式激活蛋白的构建。

Development of transgenic corn planthopper Peregrinus maidis that express the tetracycline transactivator.

机构信息

Department of Entomology and Plant Pathology, North Carolina State University, Raleigh, North Carolina, USA.

Department of Biology, University of Washington, Seattle, Washington, USA.

出版信息

Insect Mol Biol. 2023 Aug;32(4):363-375. doi: 10.1111/imb.12836. Epub 2023 Mar 10.

Abstract

The corn planthopper, Peregrinus maidis, is a vector of several maize viruses and is consequently a significant agricultural pest in many tropical and subtropical regions. As P. maidis has developed resistance to insecticides, the aim of this study was to develop transgenic P. maidis strains that could be used for future genetic biocontrol programs. To facilitate the identification of transgenic P. maidis, we isolated and characterized the promoters for the P. maidis ubiquitin-like and profilin genes. Transient expression assays with P. maidis embryos showed that both promoters were active. Transgenic lines were established using piggyBac vectors and fluorescent protein marker genes. The lines carried an auto-regulated tetracycline transactivator (tTA) gene, which has been widely used to establish conditional lethal strains in other insect species. The transgenic lines showed low levels of tTA expression but were viable on diet with or without doxycycline, which inhibits the binding of tTA to DNA. We discuss possible modifications to the tTA overexpression system that could lead to the successful development of conditional lethal strains. To our knowledge, this is the first report of a transgenic Hemiptera. The approach we have taken could potentially be applied to other Hemiptera and, for P. maidis, the technology will facilitate future functional genomics studies.

摘要

褐飞虱,是几种玉米病毒的载体,因此在许多热带和亚热带地区是一种重要的农业害虫。由于褐飞虱对杀虫剂产生了抗性,本研究的目的是开发可用于未来遗传生物防治计划的转基因褐飞虱品系。为了便于鉴定转基因褐飞虱,我们分离并鉴定了褐飞虱泛素样和丝氨酸结合蛋白基因的启动子。用褐飞虱胚胎进行的瞬时表达分析表明,这两个启动子均具有活性。使用 piggyBac 载体和荧光蛋白标记基因建立了转基因品系。这些品系携带一个自动调控的四环素转录激活剂(tTA)基因,该基因已广泛用于在其他昆虫物种中建立条件致死株。转基因品系显示出低水平的 tTA 表达,但在含有或不含有强力霉素的饮食中是存活的,强力霉素抑制 tTA 与 DNA 的结合。我们讨论了可能对 tTA 过表达系统进行的修改,这些修改可能会导致成功开发条件致死株。据我们所知,这是首例关于转 Hemiptera 的报道。我们采用的方法可能适用于其他 Hemiptera,对于褐飞虱来说,该技术将促进未来的功能基因组学研究。

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