气味结合蛋白在印度谷螟重复感染行为中的功能研究

The Indian meal moth, Plodia interpunctella, is a major economic pest of stored products, has a peculiar damage habit, easy to repeat infected hosts, but the regulation mechanism of the repeated infection behavior is not clear. In this project, by screening the differences chemicals of volatiles before and after infected host wheat, using EAG technology research the behavior selection to the host odors of P. interpunctella which infected and uninfected the host wheat, identification the key volatile components which caused repeated infection behavior in P. interpunctella; using the technology of transcriptome sequencing and gene expression profiling, the odorant binding protein genes involved in repeat infection behavior were identified from the antennal transcriptome library in the indian meal moth, which infected and uninfected the wheat, and by cloning and expressing the recombinant odorant binding protein, fluorescence competitive binding assays and site directed mutagenesis were performed to examine the odor spectrum of the recombinant protein and binding characteristics with the odor molecules, identify the OBP species that lead to the behavior of repeated infection in P. interpunctella, explore the function of OBP gene in the behavior of repeating infections, and clarify the selection mechanism between P. interpunctella - storage host. The results have great significance and provide new ideas and strategy for interfering the connection between the Indian meal moth and the stored products.

印度谷螟是极具危害性的重要仓储害虫，具有独特的危害习性：易重复感染寄主，但对导致其重复感染行为的调控机制目前尚不明确。本研究通过筛选重复感染前后寄主小麦差异性化学挥发物，利用EAG和Y型嗅觉仪研究印度谷螟对感染前后宿主气味的行为趋性，鉴定引起印度谷螟重复感染行为的关键化学挥发物成分；利用转录组文库和基因表达谱相结合的技术，在重复感染小麦前后印度谷螟触角转录组文库中筛选与印度谷螟重复感染行为相关的气味结合蛋白（OBP）基因，通过原核表达，利用荧光配体竞争结合实验和定点突变技术解析OBP与挥发物气味分子的结合特征和气味识别谱，明确导致印度谷螟重复感染行为的OBP种类，探讨OBP在印度谷螟重复感染行为中的功能。通过以上研究，为阐明印度谷螟-储粮宿主之间的选择机理，发展新的防治方法干扰印度谷螟-储粮宿主之间的联系，从而阻断靶标害虫危害提供新的思路和途径。

嗅觉相关蛋白在昆虫嗅觉信号传导过程中起着重要的作用，其中大部分基因的功能及其参与的信号转导途径仍不清楚。本研究基于印度谷螟对小麦的感染取食行为，利用气相色谱-质谱联用（GC-MS）技术对印度谷螟成虫交配前后特异挥发性化合物以及小麦感染印度谷螟前后特异挥发性化合物进行了鉴定，通过构建印度谷螟转录组，鉴定筛选出136个嗅觉相关基因，包括42个OBPs，12个CSPs，57个Ors，3个ODEs，3个SNMPs和19个IRs，通过Q-PCR方法分析了OBPs在不同组织中的表达情况，通过构建原核表达载体，对PintGOBP2进行了表达纯化，结合昆虫行为学方法，分析研究OBPs在印度谷螟嗅觉感受过程中的作用，探讨了OBPs蛋白相应嗅觉感受信号传导过程的分子机制。该研究为进一步阐明昆虫的嗅觉感受机制、开发嗅觉基因靶标防治印度谷螟技术提供丰富的理论依据。