本文選題：黃曲條跳甲 + 化學感受基因��； 參考：《仲愷農業(yè)工程學院》2015年碩士論文

【摘要】：黃曲條跳甲Phyllotreta striolata(Fabricius)是十字花科蔬菜的主要害蟲之一,在中國南方地區(qū)嚴重危害十字花科蔬菜的生產。目前黃曲條跳甲的防治主要依靠化學防治,但農藥的過度使用導致抗藥性產生和食品安全隱患。因此,需要發(fā)展新型的防治策略。應用引誘化合物或驅避化合物是害蟲防治的發(fā)展方向之一,研究黃曲條跳甲的化學感受基因可以加深對化學感受基因的認知,同時也為將來對其功能研究奠定科學基礎。本論文對本實驗室前期構建的黃曲條跳甲轉錄組數據進行生物信息學分析,通過同源性分析和c DNA末端快速擴增技術(RACE)從黃曲條跳甲轉錄組數據庫中鑒定化學感受相關基因并獲得基因全長;應用生物信息學方法分析候選基因的結構和特征,并構建系統(tǒng)發(fā)育進化樹;應用實時熒光定量PCR技術,研究化學感受基因在不同性別的嗅覺器官與非嗅覺器官的表達差異。通過以上研究,獲得以下結果:1)黃曲條跳甲化學感受基因鑒定和生物信息學分析應用生物信息學方法和RACE技術共鑒定16個化學感受基因,其中氣味結合蛋白(Odorant-binding proteins,OBPs)基因3個,化學感受蛋白(Chemosensory proteins,CSPs)基因2個,氣味受體(Odorant receptors,ORs)基因1個,離子型受體(Ionotropic receptors,IRs)基因7個,感受神經膜蛋白(Sensory neuron membrane proteins,SNMPs)基因2個,味覺受體(Gustatory receptors,GRs)基因1個。通過選取部分昆蟲的化學感受基因與鑒定所得的黃曲條跳甲化學感受基因構建系統(tǒng)發(fā)育進化樹,分析顯示黃曲條跳甲化學感受基因多與赤擬谷盜等鞘翅目昆蟲的化學感受基因的進化關系較為接近。(1)Pstr OBP1、Pstr OBP2、Pstr OBP3分別與松墨天牛Monochamus alternatus的Malt OBP4、Malt OBP3和赤擬谷盜Tribolium castaneum的Tcas OBP19d同源性最高,分別為52%、41%、42%。(2)Pstr CSP1與Pstr CSP2分別與與黃粉甲Tenebrio molitor的Tmol CSP3和赤擬谷盜T.castaneum的Tcas CSP6同源性最高,分別為57%和37%。(3)Pstr SNMP1和Pstr SNMP2分別與松山甲蟲Dendroctonus ponderosae的Dpon SNMP(AFI45066.1)和赤擬谷盜T.castaneum的Tcas SNMP2-like(XP_008198962.1)最高,分別為49%和46%;系統(tǒng)進化分析顯示,2個SNMP分在SNMP1和SNMP2兩類的分支上。(4)Pstr IR1、Pstr IR2、Pstr IR5、Pstr IR6、Pstr IR7均與glutamate receptor ionotropic,kainate類基因同源性較高,Pstr IR3與赤擬谷盜T.castaneum的glutamate receptor 2的同源性最高,同源性為59%,Pstr IR4與赤擬谷盜glutamate[NMDA]receptor subunit 1同源性最高,同源性達到87%。(5)Pstr OR2與赤擬谷盜T.castaneum的Tcas OR37同源性最高,同源性為34%。(6)Pstr GR1與Tcas GR10同源性最高同源性為69%,與Dmel GR21a為直系同源。2)嗅覺感受基因在黃曲條跳甲兩性嗅覺和非嗅覺的組織的表達分析采用q RT-PCR方法,分別選取黃曲條跳甲不同性別的觸角和腹端,研究了其雌雄蟲Pstr OBP1、Pstr OBP2、Pstr OBP3、Pstr CSP1、Pstr IR1、Pstr IR2、Pstr IR3、Pstr SNMP1和Pstr SNMP2在這2個部位的表達情況。(1)OBPs的表達研究顯示:Pstr OBP1與Pstr OBP2為觸角特異表達,Pstr OBP1在雄蟲觸角表達量是雌蟲觸角表達量的2倍,Pstr OBP2在雄蟲和雌蟲觸角均是特異表達,且在兩性觸角的表達量無明顯差異,Pstr OBP3在雄蟲腹部的表達量高于觸角,同時雄蟲觸角與雌蟲觸角表達量相差不大,因此推測Pstr OBP1可能在雄蟲感知異性性信息素的過程中起作用;推測Pstr OBP2可能參與感知普通氣味;推測Pstr OBP3可能參與雄蟲聚集信息素的釋放和對聚集信息素的感知;(2)CSPs的表達研究顯示:Pstr CSP1在觸角和腹部均有表達,在雌蟲腹部表達最低,其余部位表達差異不顯著;(3)IRs表達研究顯示:Pstr IR1、Pstr IR2、Pstr IR3在觸角和腹部都有表達,總體上雄蟲表達量高于雌蟲;4)SNMPs表達研究顯示:Pstr SNMP1與Pstr SNMP2在觸角和腹部都有表達,但表達差異不明顯。
[Abstract]:Phyllotreta striolata (Fabricius) is one of the main pests of cruciferous vegetables and seriously endangering the production of cruciferous vegetables in the south of China. At present, the prevention and control of the beetle is mainly based on chemical control, but the excessive use of pesticides leads to the potential of resistance and food safety. Therefore, a new type of food is needed to develop a new type. The application of attractant compounds or repellent compounds is one of the development directions of pest control. The study of the chemoreceptor genes of the Yellow curved strip can deepen the cognition of the chemoreceptor genes and lay a scientific foundation for the study of its function in the future. According to bioinformatics analysis, using homology analysis and C DNA terminal rapid amplification technique (RACE) to identify the chemoreceptor related genes and obtain the whole length of the gene from the database of the Yellow curved strip armor transcriptome, the structure and characteristics of the candidate genes were analyzed by bioinformatics, and the phylogenetic tree was constructed; real time fluorescence determination was applied. PCR technique was used to study the difference between the olfactory organ and the non olfactory organ in different sexes. Through the above study, the following results were obtained: 1) the identification of the chemoreceptor gene and the bioinformatics analysis, the application of bioinformatics and the RACE technology to identify 16 chemoreceptor genes. There are 3 Odorant-binding proteins (OBPs) genes, 2 Chemosensory proteins (CSPs) genes, 1 Odorant receptors (ORs) genes, 7 ionic receptors (Ionotropic receptors, IRs) genes, and 2 receptor genes. Ptors, GRs) gene 1. The phylogenetic tree was constructed by selecting the chemoreceptor genes of some insects and the identification of the chemoreceptor genes of the Yellow curved strip armor. The analysis showed that the evolution of chemoreceptor genes of Coleoptera and other Coleoptera insects, such as hellfish, was close to that of the chemoreceptor genes of the beetle. (1) Pstr OBP1, Pstr OBP2, P STR OBP3 is the highest homology with Malt OBP4, Malt OBP3 and Tribolium castaneum of Monochamus alternatus, respectively, 52%, 41%. 37%. (3) Pstr SNMP1 and Pstr SNMP2 are respectively the highest of Dpon SNMP (AFI45066.1) and the T.castaneum of Matsuyama Kousa Dendroctonus ponderosae, respectively, 49% and 46%, respectively. The homology of IR7 is higher than that of glutamate receptor ionotropic and kainate gene. Pstr IR3 has the highest homology with the glutamate receptor 2 of the ash T.castaneum, and the homology is 59%. The homology of OR37 is the highest, the homology is 34%. (6) Pstr GR1 and Tcas GR10 homology with the highest homology, and the expression of the olfactory receptor gene with the Dmel GR21a as the direct homologous.2) in the olfactory and non olfactory tissues of the stripe dive is analyzed by Q RT-PCR method, respectively, to select the antennae and the abdominal end of the different sex of the stripe diaphora. The expression of Pstr OBP1, Pstr OBP2, Pstr OBP3, Pstr CSP1, Pstr IR1, Pstr IR2, and Pstr IR2 are expressed as the specific expression of the antennae, and the expression of the male antennae is 2 times the female's antennae expression. The expression of Pstr OBP3 is higher than the antennae in the abdomen of the male, and the difference between the male antennae and the female antennae is not significant. Therefore, it is suggested that Pstr OBP1 may play a role in the process of male perception of heterosexual pheromone; Pstr OBP2 may be involved in the perception of common odor. It is speculated that Pstr OBP3 may be involved in the release of the aggregation pheromone of the male and the perception of the aggregation pheromone. (2) the expression of CSPs shows that Pstr CSP1 is expressed in the antennae and abdomen, and the expression in the abdomen is the lowest, and the expression of the rest is not significant. (3) the expression of IRs shows that Pstr IR1, Pstr IR2, Pstr IR3 have a table in the antennae and abdomen. In general, the expression level of male was higher than that of female. 4) SNMPs expression study showed that Pstr SNMP1 and Pstr SNMP2 were expressed in antennae and abdomen, but the difference was not obvious.
【學位授予單位】：仲愷農業(yè)工程學院
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：S433.5

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1 程乘;尹姣;曹雅忠;李克斌;彭宇;王倩倩;;昆蟲氣味結合蛋白研究進展[A];“創(chuàng)新驅動與現代植保”——中國植物保護學會第十一次全國會員代表大會暨2013年學術年會論文集[C];2013年

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本文編號：1985786