入侵害虫红棕象甲核转录因子RfRelish编码基因的克隆与表达特征分析

发布时间：2018-06-07 16:19

本文选题：红棕象甲 + 昆虫免疫　；参考：《福建农林大学》2017年硕士论文

【摘要】：红棕象甲Rhynchophorus ferrugineus Olivier是我国的一种高风险性外来有害生物,是棕榈植物的一种世界性毁灭害虫。该害虫危害隐蔽,田间种群已产生抗药性而且生物防治的控制效率一直较低。因此,寻找针对该害虫新的、有效的可持续控制策略迫在眉睫。昆虫免疫是昆虫对抗病原天敌的最主要途径,然而对于该害虫的免疫防御机理知之甚少。因此,本文通过红棕象甲核转录因子RfRelish的克隆和定量表达分析等手段解析其潜在的免疫功能角色。研究发现,该核转录因子RfRelish基因的全长cDNA序列为2707 bp,开放阅读框为2148 bp。其中,5'非编码区为87 bp,3'非编码区为472 bp,包含AAAAAAA加尾信号并含有长为12bp的poly(A)尾。该基因编码一个由714个氨基酸组成的蛋白质。该蛋白质的等电点为5.46,其分子量为81.5K Da。该编码蛋白具有N末端的Rel homology domain(RHD)、IPT domain(Ig-like,plexins,transcription factors)和由四个 ankyrin repeats组成的IκB-like domain等三个NF-κB家族蛋白所具有的典型保守结构域。同时,该蛋白具有核转位定位信号,该信号由第210-213位氨基酸残基KRPR组成。这些保守结构域的存在说明红棕象甲RfRelish基因编码的蛋白质是一类具有重要功能的转录因子。荧光定量分析发现,该基因在红棕象甲不同组织中的表达量存在极显著差异;其中,在免疫相关组织脂肪体、血淋巴、嗉囊和中肠中的表达量显著高于非免疫相关组织。注射大肠杆菌后,红棕象甲脂肪体中RfRelish基因的表达量显著提高。而注射金黄色葡萄球菌6h后,仅红棕象甲肠道中的RjtRelish基因表达量明显高于对照组。已有证据显示,红棕象甲的肠道中栖居着丰富的细菌群落。因此,这说明RfRelish基因的编码蛋白不仅在红棕象甲的系统免疫中发挥重要作用,而且在肠道菌群稳态的调控中也扮演关键角色。
[Abstract]:Red brown weevil (Rhynchophorus ferrugineus Olivier) is a high risk alien pest in China and a worldwide destructive pest of palm plants. The pest is concealed, the field population has developed resistance and the control efficiency of biological control has been low. Therefore, it is urgent to find a new and effective sustainable control strategy for the pest. Insect immunity is the most important way for insects to fight natural enemies, but little is known about the immune defense mechanism of insect pests. Therefore, the potential role of RfRelish was analyzed by cloning and quantitative expression analysis of its transcription factor. It was found that the full-length cDNA sequence of RfRelish gene was 2707 BP and the open reading frame was 2148 BP. Among them, the non-coding region of 5'is 87bp / 3'and the non-coding region is 472bp. it contains AAAAAAA signal with tail and 12bp's long polycaudal tail. The gene encodes a protein consisting of 714 amino acids. The isoelectric point of the protein is 5.46 and its molecular weight is 81.5 K Da. The encoding protein has the typical conserved domain of the N-terminal Rel homology domain Rel homology domain like Ig-like transcription factors and three NF- 魏 B family proteins, I 魏 B-like domain, composed of four ankyrin repeats. At the same time, the protein has nuclear translocation signal, which is composed of 210-213 amino acid residues KRPR. The existence of these conserved domains suggests that the proteins encoded by the RfRelish gene of the red brown weevil are a class of transcription factors with important functions. Fluorescence quantitative analysis showed that there were significant differences in the expression of the gene in different tissues of red brown weevil, and the expression of the gene was significantly higher in immune related tissues than in non-immune related tissues, such as fat body, hemolymph, crop and midgut. After the injection of E. coli, the expression of RfRelish gene in the fat body of red brown weevil was significantly increased. However, the expression of RjtRelish gene in the intestine of red brown weevil was significantly higher than that of the control group 6 h after injection of Staphylococcus aureus. There is evidence that red brown elephant beetles live in abundant bacterial communities in their intestines. Therefore, the encoding protein of RfRelish gene not only plays an important role in the systemic immunity of red brown weevil, but also plays a key role in the regulation of intestinal flora homeostasis.
【学位授予单位】：福建农林大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S763.38

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