马铃薯甲虫气味结合蛋白的序列和基因表达谱分析  

Sequence analysis and gene expression profiling of odorant binding proteins in the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)

作　　者：张方梅[1,2] 刘杨[2] 李祥瑞[2] 张云慧[2] 程登发[2] 郭文超[3] 吐尔逊.阿合买提 ZHANG Fang-Mei;LIU Yang;LI Xiang-Rui;ZHANG Yun-Hui;CHENG Deng-Fa;GUO Wen-Chao;TURSUN Ahmat(Henan Provincial South Henan Crop Pest Green Prevention and Control Academician Workstation,Xinyang Agriculture and Forestry University,Xinyang,Henan 46400,China;State Key Laboratory for Biology of Plant Diseases and Insect Pests,Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,China;Institute of Plant Protection,Xinjiang Academy of Agricultural Sciences,Urumchi 830091,China)

机构地区：[1]信阳农林学院河南省豫南农作物有害生物绿色防控院士工作站,河南信阳464000 [2]中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室,北京100193 [3]新疆农业科学院植物保护研究所,乌鲁木齐830091

出　　处：《昆虫学报》2019年第4期428-441,共14页Acta Entomologica Sinica

基　　金：国家重点研发计划(2018YFD0200800);公益性行业(农业)科研专项(201103026-2);信阳农林学院重点学科培育学科建设项目(ZDXK201701);河南省高等教育教学改革研究与实践项目(2017SJGLX135)

摘　　要：【目的】昆虫气味结合蛋白(OBPs)在昆虫嗅觉行为中发挥着重要作用。马铃薯甲虫Leptinotarsa decemlineata是马铃薯上一种最主要的毁灭性害虫。为阐明该虫嗅觉识别分子机制,本研究对马铃薯甲虫26个OBP基因序列特征及组织表达谱进行研究。【方法】基于马铃薯甲虫触角转录组测序数据,利用生物信息学方法及qRT-PCR技术,分别对马铃薯甲虫26个LdecOBPs (LdecOBP1-LdecOBP26)的系统进化及基因的组织表达谱进行分析。【结果】除LdecOBP26基因外,其余25个LdecOBPs基因序列均具有完整的开放阅读框,编码120～255个氨基酸残基,预测的蛋白分子量为13.66～29.38 kD,等电点为4.12～8.42,它们属于两个亚家族,其中13个为Classical-C OBPs, 12个为Minus-C OBPs。除LdecOBP3和LdecOBP26外,其他24个LdecOBPs的N端均由16～23个氨基酸组成的信号肽序列。不同的OBPs亚家族均具有各自典型保守的Cys残基。LdecOBPs之间高度分化,氨基酸序列一致性在3.20%～41.91%。系统进化树分析表明,LdecOBPs与沙葱萤叶甲Galeruca daurica的GdauOBPs亲缘关系最近。基因表达谱分析显示,26个LdecOBPs基因在马铃薯甲虫的不同组织中表达,其中有12个LdecOBPs基因(LdecOBP2,LdecOBP4,LdecOBP6,LdecOBP9,LdecOBP10,LdecOBP12,LdecOBP13,LdecOBP16,LdecOBP20-22和LdecOBP24)在触角中高表达,2个LdecOBPs基因(LdecOBP5和LdecOBP17)在足中高表达,其他12个LdecOBPs基因(LdecOBP1,LdecOBP3,LdecOBP7,LdecOBP8,LdecOBP11,LdecOBP14,LdecOBP15,LdecOBP18,LdecOBP19,LdecOBP23,LdecOBP25和LdecOBP26)在触角、头(去除触角)、胸、腹、足和翅这些组织中均表达。【结论】本研究结果为进一步研究马铃薯甲虫嗅觉识别分子机制奠定了基础。【Aim】 Odorant-binding proteins (OBPs) play a fundamental role in insect olfaction. The Colorado potato beetle (CPB),Leptinotarsa decemlineata (Say),is one of the most notorious insect pests of potato. The sequence properties and tissue expression profiles of 26 putative OBP genes of L. decemlineata were identified to clarify the molecular mechanisms of its olfactory perception.【Methods】 Based on antennal transcriptome sequencing data of L. decemlineata,the phylogeny and tissue-specific gene expression profiles of 26 LdecOBPs (LdecOBP1-LdecOBP26) in L. decemlineata were analyzed by bioinformatic methods and qRT-PCR,respectively.【Results】 Except LdecOBP 26,the genes of the other 25 LdecOBPs have the full-length open reading frames,encoding 120-255 amino acid residues with the predicted molecular weights of 13.66-29.38 kD and isoelectric points of 4.12-8.42. The 25 LdecOBPs belong to two subfamilies,with 13 Classical-C OBPs and 12 Minus-C OBPs. Except LdecOBP3 and LdecOBP26,each of the other 24 LdecOBPs contains a signal peptide at the N-terminus consisting of 16-23 amino acid residues. These OBPs in each subfamily have their own typical conservative Cys residues. LdecOBPs are highly divergent and their amino acid sequence identities range from 3.20% to 41.91%. Phylogenetic analysis showed that LdecOBPs and GdauOBPs of Galeruca daurica are most closely related. Gene expression profiling showed the 26 LdecOBPs genes were expressed in different adult tissues of L. decemlineata,with 12 LdecOBPs genes ( LdecOBP2,LdecOBP4,LdecOBP6,LdecOBP9,LdecOBP10,LdecOBP12,LdecOBP13,LdecOBP16,LdecOBP 20-22 and LdecOBP 24) highly expressed in antennae,two genes ( LdecOBP 5 and LdecOBP 17) highly expressed in legs,and the other 12 ( LdecOBP1,LdecOBP3,LdecOBP7,LdecOBP8,LdecOBP11,LdecOBP14,LdecOBP15,LdecOBP18,LdecOBP19,LdecOBP23,LdecOBP 25 and LdecOBP 26) expressed in tissues antennae,head (with antennae removed),thorax,abdomen,legs and wings.【Conclusion】 These results establish a foundation for further research on the mol

关 键 词：马铃薯甲虫 气味结合蛋白 序列分析 组织表达谱 系统发育 

分 类 号：Q966[生物学—昆虫学]