槲皮素和球孢白僵菌对小菜蛾酚氧化酶基因翻译表达的影响

发布时间：2018-06-30 11:02

本文选题：酚氧化酶 + 槲皮素　；参考：《山东农业大学》2016年硕士论文

【摘要】：酚氧化酶(phenoloxidase,简称PO,EC.1.14.18.1)在昆虫的生长发育和内源性免疫过程中具有重要功能,如具有黑化或包被入侵的病原物、昆虫伤口的愈合等功能。酚氧化酶可以作为害虫控制中一个潜在的作用靶标,通过开发酚氧化酶抑制剂,对于害虫绿色防控和环境生态安全提供新思路。本论文利用植物源的槲皮素和微生物农药球孢白僵菌处理小菜蛾,通过制备小菜蛾两个酚氧化酶基因的多克隆抗体,研究不同处理对酚氧化酶蛋白表达水平的影响。酚氧化酶及其抑制剂也一直是国内外科学家研究的热点。本论文以小菜蛾为研究对象,在制备获得小菜蛾PxPPO1和PxPPO2基因抗体的基础之上,利用蛋白质印迹技术(Western Blot)研究了槲皮素和球孢白僵菌(Beauveria bassiana)处理小菜蛾后,小菜蛾酚氧化酶蛋白翻译表达水平的相对变化,从而来探讨外来效应物对酚氧化酶基因在翻译水平上的影响。主要结果如下:1.成功制备获得小菜蛾PxPPO1和PxPPO2基因的多克隆抗体。2.分别对内参基因β-actin以及目的基因PxPPO1和PxPPO2进行了 Western Blot曝光饱和分析,根据上样量与信号饱和点出现的时间,确定Western Blot试验中上样量为 10μg。3.采用微量点滴法,用不同浓度槲皮素处理小菜蛾3龄幼虫,处理不同时间后,PxPPO1和PxPPO2基因翻译相对表达量随槲皮素浓度升高而明显减少。4.采用浸渍法,用球孢白僵菌侵染小菜蛾3龄幼虫,成功侵染不同时间后,PxPPO1和PxPPO2基因翻译相对表达量在处理2h时比对照处理提高,在6h、12h、24h和48h时蛋白表达量均比对照处理减少。
[Abstract]:Phenoloxidase (POC. 1.14.18.1) plays an important role in the growth and development of insects and endogenous immune process, such as melanization or encapsulation of invading pathogens, wound healing of insects and so on. Phenoloxidase can be used as a potential target in pest control. Through the development of phenoloxidase inhibitors, it provides a new idea for pest green control and environmental ecological safety. In this paper, quercetin from plant and microbial pesticide Beauveria bassiana were used to treat diamondback moth. The polyclonal antibody of two phenoloxidase genes of Plutella xylostella was prepared to study the effect of different treatments on the expression of phenoloxidase protein. Phenoloxidase and its inhibitors have also been the focus of scientists at home and abroad. Based on the preparation of PxPPO1 and PxPPO2 gene antibodies from diamondback moth (Plutella xylostella), the effects of quercetin and Beauveria bassiana on diamondback moth (Plutella xylostella) were studied by Western blot. The relative changes of phenoloxidase protein translation and expression level in Plutella xylostella were used to explore the effect of foreign effectors on the translation level of phenoloxidase gene. The main results are as follows: 1. The polyclonal antibodies against PxPPO1 and PxPPO2 genes of Plutella xylostella were successfully prepared. The 尾 -actin gene and the target genes PxPPO1 and PxPPO2 were analyzed by Western Blot. According to the amount of sample and the time of signal saturation, it was determined that the amount of sample was 10 渭 g 路3 in Western Blot. The third instar larvae of Plutella xylostella were treated with quercetin at different concentrations by microdrop method. The relative expression of PxPPO1 and PxPPO2 genes decreased significantly with the increase of quercetin concentration. The third instar larvae of Plutella xylostella were infected with Beauveria bassiana by dipping method. The relative expression of PxPPO1 and PxPPO2 genes was increased at 2 h after successful infection, and decreased at 6 h, 12 h, 24 h and 48 h, respectively.
【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S433.4

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