本文關鍵詞： 褐飛虱 茚蟲威 氯蟲苯甲酰胺 類胰島素基因 保幼激素酯酶基因 保幼激素環(huán)氧水解酶基因 保幼激素二醇激酶基因　出處：《揚州大學》2017年碩士論文　論文類型：學位論文

【摘要】：褐飛虱 Nilaparvata lugens(Stal)(Hemiptera:Delphacidae)屬半翅目,飛虱科,是水稻上的重要害蟲。茚蟲威和氯蟲苯甲酰胺作為兩種新型殺蟲劑,在生產實踐中被廣泛使用。前期研究表明亞致死濃度的茚蟲威和氯蟲苯甲酰胺能夠抑制褐飛虱的生殖。褐飛虱的生殖受許多因素影響,其中保幼激素對卵巢發(fā)育起重要作用,而保幼激素收受到水解酶的調控。胰島素是昆蟲重要的內分泌激素,對昆蟲生理生化和生殖有調控作用,同時也與保幼激素有密切聯系。本文通過克隆得到了保幼激素二醇激酶Nljhdk基因的全長,分析了該基因在短翅型褐飛虱不同齡期的表達差異,并利用RNAi研究其與保幼激素酯酶Nljhe和環(huán)氧水解酶Nljheh的關系,以及亞致死濃度茚蟲威和氯蟲苯甲酰胺對褐飛虱4個類胰島素基因NlIlp1-4及3種保幼激素水解酶基因mRNA相對表達水平的影響,進一步揭示這兩種藥劑抑制生殖的機理。利用RT-PCR技術克隆了褐飛虱保幼激素二醇激酶基因684 bp cDNA(Nljhdk)。Nljhdk開放閱讀框長558 bp,編碼185個氨基酸,預測蛋白分子質量21.57 kDa,等電點為4.54。多重序列比對發(fā)現Nljhdk與天蠶Antheraea yamamai(KC127671)、桔小實繩Bactrocera dorsalis(JN256054)、家蠶Bombyx mori(AY363308)、馬鈴薯甲蟲 Leptinotarsa decemlineata(KP295467)、煙草天蛾Manduca sexta(AJ430670)、亞洲玉米螟 Ostrinia furnacalis(HM626514)、小菜蛾 Plutella xylostella(JN410823)和斜紋夜蛾Spodoptera litura(KP192493)JHDK基因的氨基酸序列同源性分別為43.65%、55.68%、41.62%、61.08%、51.35%、48.65%、43.78%和45.95%。應用qPCR對Nljhdk表達模式分析發(fā)現,Nljhdk在若蟲和成蟲各個發(fā)育階段都有表達,在4齡時表達水平最低,在5齡時有最高表達,并且在雄成蟲中高于雌成蟲。利用RNAi技術,發(fā)現干擾Nljhdk對Nljhe沒有明顯影響,但能夠提高Nljheh的表達水平。茚蟲威對褐飛虱的亞致死效應研究發(fā)現,茚蟲威亞致死濃度能夠顯著抑制卵巢內NlIlp2-4的表達水平,20 mg/L茚蟲威時能夠顯著上調NlIlp1的相對表達量10.67%;Nljhe表達水平隨著濃度(3,5,10和20 mg/L)的增加,比對照分別顯著上升了 52.92%、38.82%、105.5%和79.44%;Nljheh相對表達量相比對照組被顯著抑制,1、3、5、10和20 mg/L濃度茚蟲威處理后顯著降低了 34.09%、30.49%、48.21%、45.21%和42.49%;Nljhdk在1和3 mg/L濃度時顯著上升了 39.62%和59.32%,其余濃度處理沒有顯著差異。氯蟲苯甲酰胺亞致死濃度處理褐飛虱后,雌蟲卵巢內NlIlp1表達水平在15 mg/L濃度時比對照顯著上升了 26.88%;NlIlp2表達水平在3.75 mg/L處理濃度比對照顯著下降7.9%,但在7.5、30和60 mg/L時比對照顯著上升了 20.13%、28.12%和47.77%;NlIlp3相對表達量只有在最高濃度60 mg/L時有明顯的升高,比對照組高153.27%;NlIlp4相對表達量在3.75 mg/L時比對照上升了 9.62%,而在7.5、15、30和60 mg/L濃度時分別比對照上升了 51.24%、69.77%、52.63%和 95.17%。15、30 和 60 mg/L 的氯蟲苯甲酰胺處理后,褐飛虱卵巢內Nljhe相對表達量分別比對照顯著上升了 25.77%、24.12%和72.45%。Nljheh的相對表達量在3.75、15和60 mg/L氯蟲苯甲酰胺處理后比對照顯著上升了 16.18%、23.09%和65.00%。15和60 mg/L的氯蟲苯甲酰胺處理后,褐飛虱卵巢內Mljhdk的相對表達量分別比對照顯著上升了 48.8%和8.51%,但其他濃度處理后,Nljhdk的相對表達量反而下降。本文深入探究了茚蟲威和氯蟲苯甲酰胺抑制褐飛虱生殖的機制,為稻田合理使用這兩種藥劑提供了理論基礎。
[Abstract]:The brown planthopper Nilaparvata lugens (Stal) (Hemiptera:Delphacidae) belong to Hemiptera, Delphacidae, is an important pest on rice. Indoxacarb and chlorantraniliprole as two kinds of new pesticides, is widely used in the production practice. Previous studies showed that sublethal concentrations of indoxacarb and chlorantraniliprole can inhibit brown planthopper the reproduction of brown planthopper. Reproduction is affected by many factors, including juvenile hormone plays an important role in ovarian development and juvenile hormone to accept hydrolase regulation. Insulin is an important hormone of insects, regulate insect physiology and reproduction, but also closely linked with the juvenile hormone. Is obtained in this paper. The cloning of full-length jhdk gene Nljhdk, analyzes the difference between the expression of the gene in brachypterous BPH at different ages, and the use of RNAi to study with the juvenile hormone esterase and Nljhe The relationship between Nljheh epoxide hydrolase, and sublethal concentrations of indoxacarb and chlorantraniliprole on BPH relative expression level influence NlIlp1-4 of 4 types of insulin gene and 3 juvenile hormone hydrolase gene mRNA, further reveal the mechanism of the two kinds of agents that inhibit reproduction. RT-PCR technology was used to clone BPH juvenile hormone diol 684 BP cDNA kinase gene (Nljhdk).Nljhdk long open reading frame of 558 BP, encoding 185 amino acids. The predicted molecular mass of 21.57 kDa and isoelectric point of 4.54. multiple sequence comparison showed that Nljhdk and Antheraea Yamamai (KC127671), a small orange solid rope Bactrocera dorsalis (JN256054), Bombyx mori (AY363308), Bombyx mori potato the beetle Leptinotarsa decemlineata (KP295467), Manduca sexta (AJ430670) of Manduca sexta, Asian corn borer (HM626514), Ostrinia furnacalis Plutella xylostella (JN410823) of diamondback moth and oblique Noctuid Spodoptera litura (KP192493) JHDK gene amino acid sequence homology were 43.65%, 55.68%, 41.62%, 61.08%, 51.35%, 48.65%, 43.78% and 45.95%. based on qPCR Nljhdk expression pattern analysis showed that Nljhdk developed in the nymph and adult stages are expressed in 4 at the age of the lowest level of expression, the highest expression at the age of 5, and higher than that of female adults in male adults. The use of RNAi technology, found that the interference of Nljhdk has no obvious effect on Nljhe, but it can increase the expression level of Nljheh. Indoxacarb on Nilaparvatalugens sublethal effects of the indene worm can inhibit the expression of WIA lethal concentration level of NlIlp2-4 in the ovary. 20 mg/L indoxacarb can up regulate the expression of NlIlp1 relative amount of 10.67%; the expression level of Nljhe with concentration (3,5,10 and 20 mg/L) increased than in the control group were significantly increased by 52.92%, 38.82%, 105.5% and 79.44%; Nljheh The relative expression was significantly inhibited compared with the control group, 1,3,5,10 and mg/L 20 concentration of indoxacarb treatment decreased significantly after 34.09%, 30.49%, 48.21%, 45.21% and 42.49%; 1 and 3 Nljhdk in the concentration of mg/L significantly increased by 39.62% and 59.32%, no significant differences in other concentrations. Chlorantraniliprole sublethal concentration. The brown planthopper, the expression level at the concentration of 15 mg/L significantly increased 26.88% NlIlp1 female ovaries; the expression level of NlIlp2 in 3.75 concentration of mg/L significantly decreased by 7.9%, but in 7.5,30 and 60 mg/L significantly increased by 20.13%, 28.12% and 47.77%; the relative expression of NlIlp3 was significantly increased only in the the highest concentration of 60 mg/L, 153.27% higher than the control group; the relative expression of NlIlp4 in 3.75 mg/L higher than the control by 9.62%, and 60 in 7.5,15,30 and mg/L concentration respectively increased by 51.24%, 69.77%, 52.63% 95.17%.15,30 and mg/L and 60 chlorantraniliprole, brown planthopper in the ovaries of the relative expression of Nljhe were significantly increased by 25.77% and 24.12%, the relative expression amount of 72.45%.Nljheh in 3.75,15 and mg/L 60 chlorantraniliprole after treatment significantly increased by 16.18%, 23.09% and 65.00%.15 and 60 mg/L chloride worm benzamide treatment, the relative expression of Mljhdk in ovary of Brown Planthopper were significantly increased by 48.8% and 8.51%, but with other treatments, the relative expression of Nljhdk decreased. This paper explores the mechanism of indoxacarb and chlorantraniliprole inhibiting BPH reproduction, provides a theoretical basis for the in rational use of these two agents.

【學位授予單位】：揚州大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：S435.112.3

【參考文獻】

相關期刊論文 前10條

1 陳羿渠;向興;貢常委;王學貴;;氯蟲苯甲酰胺亞致死劑量對甜菜夜蛾主要解毒酶活性與生長繁殖的影響[J];中國農業(yè)科學;2017年08期

2 劉鵬;孫麗麗;張琪慧;曹傳旺;;舞毒蛾小分子熱激蛋白基因分析及對甲萘威脅迫的響應[J];北京林業(yè)大學學報;2017年01期

3 尹顯慧;吳青君;張友軍;龍友華;吳小毛;李榮玉;李明;趙贊偉;王英;;低劑量多殺菌素繼代處理對小菜蛾nAChR基因表達的影響[J];昆蟲學報;2016年11期

4 沈登榮;何超;宋文菲;田學軍;袁盛勇;張宏瑞;;高效氯氟氰菊酯亞致死濃度對番石榴實蠅代謝酶的影響[J];西南師范大學學報(自然科學版);2015年08期

5 余慧靈;向興;袁貴鑫;陳羿渠;王學貴;;溴氰蟲酰胺亞致死劑量對甜菜夜蛾生長發(fā)育及體內解毒酶活性的影響[J];昆蟲學報;2015年06期

6 靳三省;孟麗峰;刁青云;;吡蟲啉對意大利蜜蜂乙酰膽堿酯酶的亞致死效應[J];應用昆蟲學報;2015年02期

7 戴瀚洋;孫洋;柏立新;趙靜;肖留斌;譚永安;;亞致死濃度甲維鹽脅迫對甜菜夜蛾幼蟲解毒酶系活力及其相關基因表達量的影響[J];棉花學報;2015年02期

8 王小強;劉長仲;邢亞田;石周;;吡蟲啉、阿維菌素和高效氯氰菊酯亞致死劑量對綠色型豌豆蚜發(fā)育及繁殖的影響[J];草業(yè)學報;2014年05期

9 JIN Min-na;XUE Jian;YAO Yun;LIN Xin-da;;Molecular Characterization and Functional Analysis of Krüppel-homolog 1(Kr-h1) in the Brown Planthopper, Nilaparvata lugens(Stl)[J];Journal of Integrative Agriculture;2014年09期

10 楊洪;王召;金道超;;氯蟲苯甲酰胺對非靶標害蟲褐飛虱實驗種群的亞致死效應[J];應用生態(tài)學報;2013年02期

相關博士學位論文 前1條

1 楊文佳;桔小實蠅幾丁質和保幼激素合成與代謝中重要基因的研究[D];西南大學;2014年

相關碩士學位論文 前5條

1 何叔舫;褐飛虱類胰島素基因分析與功能研究[D];浙江大學;2015年

2 張紅梅;茚蟲威亞致死劑量對褐飛虱和二化螟保幼激素和保幼激素酯酶基因影響的比較研究[D];揚州大學;2015年

3 周云龍;褐飛虱保幼激素環(huán)氧化物水解酶基因jheh功能研究[D];華中農業(yè)大學;2014年

4 楊霞;茚蟲威和氯蟲苯甲酰胺對褐飛虱亞致死效應的研究[D];揚州大學;2013年

5 都振寶;吡蟲啉和噻蟲嗪亞致死劑量對荻草谷網蚜生命表參數及取食行為的影響[D];華中農業(yè)大學;2012年

，

本文編號：1479098