华山松大小蠹和共生真菌P450基因差异性表达与解毒功能研究

发布时间：2018-03-11 21:39

  本文选题：华山松大小蠹　切入点：秦岭细粘束孢　出处：《西北农林科技大学》2016年博士论文　论文类型：学位论文

【摘要】：细胞色素P450酶作为昆虫重要的解毒酶系,不仅与昆虫信息素和激素合成密切相关,而且在降解寄主次生代谢毒素和杀虫剂等方面发挥着重要的作用。小蠹虫在入侵寄主树木,以及在寄主树木韧皮部和木质部组织内生存繁殖的整个生命过程中必须具备克服寄主树木原生性和诱导性抗性,降解寄主树木树脂(萜类物质)毒害的能力。本研究选择在我国秦岭巴山林区持续发生的重大森林害虫——华山松大小蠹(Dendroctonus armandi)为研究对象,旨在通过对华山松大小蠹细胞色素P450基因系统发育,华山松大小蠹细胞色素P450基因在不同发育阶段与组织部位表达的差异与分布,华山松大小蠹成虫对华山松挥发性物质的耐受性,以及华山松大小蠹和共生真菌秦岭细粘束孢(Leptographium qinlingensis)P450基因对萜类物质的响应模式等研究,揭示华山松大小蠹P450基因的多样性,华山松大小蠹克服寄主华山松多元抗性相关的P450基因类群,华山松大小蠹和共生真菌P450基因在克服寄主华山松抗性和降解寄主华山松萜类物质毒害中的作用等重大科学问题,取得以下研究结果:1.华山松大小蠹成虫对寄主华山松5种单萜类化合物((±)-α-Pinene、(-)-α-Pinene、(-)-β-Pinene、(+)-3-Carene、(±)-Limonene)和华山松萜类提取物耐受性存在显著差异,寄主华山松萜类物质对华山松大小蠹具有明显的抗性和致死阈值,当萜类物质浓度超过阈值(600 ppm)死亡率迅速上升。2.华山松大小蠹至少具有64条P450基因,且在家族上存在多样性。在进化分枝CYP3中存在小蠹虫特异性P450基因,它们的功能与华山松大小蠹适应特殊生境、克服寄主华山松多元抗性和华山松大小蠹特有的行为与生活史密切相关。3.华山松大小蠹43条P450基因在不同发育阶段表达的反映了华山松大小蠹P450基因时间表达的差异性与功能的多样性,其中在华山松大小蠹幼虫向蛹和成虫的发育过程中P450基因表达差异显著,成虫阶段随着羽化、扩散和入侵,华山松大小蠹成虫CYP3、CYP4进化分枝的P450基因表达差异显著,这说明在华山松大小蠹幼虫、蛹和成虫的整个发育过程中有多种P450酶的参与,华山松大小蠹P450基因在时间和发育阶段的表达差异性与华山松大小蠹不同时间和发育阶段所需要克服寄主华山松萜烯类物质抗性的差异具有密切的关系,而且与华山松大小蠹寄主选择有关。4.华山松大小蠹43条P450基因在雌雄成虫的触角、消化道和生殖器中表达差异显著,且具有性别差异性,这说明华山松大小蠹P450基因在性别或组织中具有功能的特异性。华山松大小蠹消化道特异表达的P450基因与降低华山松树脂毒害密切相关,触角中特异性表达的P450基因与降解气味分子和寄主选择密切相关。5.华山松大小蠹关键P450基因和秦岭细粘束孢(Leptographium qinlingensis)P450基因对寄主华山松萜类物质具有特殊的响应模式,华山松大小蠹和共生真菌通过调控自身P450酶的表达水平,降低寄主华山松树脂等对华山松大小蠹的毒害。6.华山松大小蠹共生真菌秦岭细粘束孢CYP51F1基因具有典型的14-α-脱甲基酶蛋白活性位点和膜锚定结构域,对华山松萜烯类物质中的三萜(甾醇类化合物)具有显著的降解作用,从而构成华山松大小蠹和共生真菌协同作用克服寄主树木抗性的机制。
[Abstract]:Cytochrome P450 enzyme as an importantdetoxification enzyme of insects, and insects not only pheromones and hormones are closely related, but also play an important role in the degradation of primary and secondary students send metabolic toxins and pesticides etc. bristletail in the invasion of host trees, and the survival of the entire life in the process of reproduction must have overcome original and host trees induced resistance in the host tree phloem and xylem tissues, degradation of host tree resin (terpenoids) poison ability. This study continues to occur in China's Qinling Mountains Bashan forests of the major forest pests in Huashan pine beetle (Dendroctonus armandi) as the research object, through the development of Huashan pine beetle cells cytochrome P450 gene system, differential expression of Dendroctonus cytochrome P450 gene in different developmental stages and tissues and the distribution of Huashan pine, Huashan pine beetle into Tolerance to insect volatile substances of Huashan pine, and Huashan pine beetle and symbiotic fungi in Qinling Mountains fine beam (Leptographium qinlingensis) spore of P450 gene on the terpenoid response pattern, diversity revealed Huashan Dendroctonus P450 gene, Huashan pine beetle overcome host Huashan P450 gene related to resistance to multiple groups of pine the Huashan pine beetle and symbiotic fungus P450 gene in host resistance and degradation of Huashan pine to overcome the host role of Huashan pinene toxicity in some important scientific problems, and achieved the following results: 1. Huashan pine beetle adults to host Huashan pine 5 monoterpenes ((+) - alpha -Pinene. (-) -Pinene (-) - alpha, beta -Pinene (+) -3-Carene (+) -Limonene) and Huashan pine terpenoids extract tolerance are significantly different, the host Huashan pine terpenes of Huashan pine beetle has obvious resistance and lethal threshold When the value of the terpenoid concentration exceeds the threshold (600 ppm) mortality rose rapidly.2. Huashan pine beetle has at least 64 P450 genes, and there exists diversity in the family. There are bristletail specific P450 gene in clade CYP3, their functions and the Huashan pine beetle adapt to special habitat, overcome host Huashan pine and Huashan pine beetle resistance multiple unique behavior and life history of.3. is closely related to the Huashan pine beetle of 43 P450 genes in different developmental stages of expression reflects the diversity of function and expression of P450 gene of Huashan pine beetle time, the pine beetle larvae to pupae in Huashan P450 development process the adults and the gene expression in the adult stage had significant difference, with the emergence, proliferation and invasion, CYP3 size of Huashan pine beetle, the differential expression of P450 gene CYP4 evolutionary branching significantly, which shows in Huashan pine beetle larvae There are a variety of insects, the P450 enzyme involved in the whole developmental process of pupae and adults in Huashan, difference of Dendroctonus P450 gene in time and developmental expression difference and Huashan pine beetle in different time and stage of development to overcome host Huashan pine terpenes resistance has close relationship with Huashan, and the size of pine choose the.4. Huashan host beetle Dendroctonus P450 43 gene in male and female adults, differential expression of digestive tract and genital significantly, and have gender differences, indicating the specificity of Huashan pine beetle has the function of P450 gene in sex or tissues. The P450 gene of Huashan pine beetle digestive tract specific expression closely associated with the reduction of Huashan pine resin toxicity, specific expression of P450 gene in the antennae and degradation of odorants and host selection of.5. is closely related to the Huashan pine beetle key gene and P450 in Qinling Mountains Fine beam (Leptographium qinlingensis) from P450 gene has a special response pattern to host Huashan pine terpenes, Huashan pine beetle and its symbiotic fungi by regulating the expression levels of the P450 enzyme, reduce the toxicity of.6. beetle host Huashan pine resin on Huashan pine and Huashan pine beetle symbiotic fungi in Qinling Mountains fine stick bundle spore the CYP51F1 gene has typical 14- alpha demethylase protein active site and membrane anchoring domain of Huashan pine, three terpene terpenes in (sterols) has significant degradation effect, which constitute the Huashan pine beetle and symbiotic fungus synergy mechanism to overcome host tree resistance.

【学位授予单位】：西北农林科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S763.38
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本文编号：1600006