摘要
为探讨小菜蛾Plutella xylostella对病原真菌入侵的免疫防御机制,利用抑制消减杂交技术构建蝉拟青霉Paecilomyces cicadae侵染的小菜蛾幼虫的抑制性差减文库,并对文库进行鉴定。ESTs序列聚类分析共获得412个独立基因。通过同源比对,28.24%ESTs为未知功能基因,71.76%ESTs与已知功能基因序列相似,包括免疫相关、金属离子结合和加工、核酸和蛋白代谢及加工、细胞信号、细胞结构、形状和流动、能量代谢、胁迫和解毒以及其它基因。鉴定出39个可能参与小菜蛾免疫蝉拟青霉侵染的免疫基因,包括:识别分子、蛋白酶及蛋白酶抑制剂、效应因子及其它免疫基因。qRT-PCR结果表明肽聚糖识别蛋白、器官芽生长因子、葛佬素、溶菌酶、酚氧化酶原激活蛋白酶3以及转铁蛋白基因均可诱导表达。免疫相关基因在不同微生物诱导下的表达模式存在不同。结果表明当病原真菌蝉拟青霉侵染寄主小菜蛾后,小菜蛾体内发生了一系列的复杂反应,小菜蛾抵御病原真菌侵染是一个多途径共同作用的复杂过程。该研究为进一步研究小菜蛾抵御病原真菌入侵的分子机制提供基础信息。
In the present study,the innate immune response of Plutella xylostella against Paecilomyces cicadae challenges was surveyed using a constructed suppression subtractive hybridization library.A total of 825 P.xylostella ESTs were obtained.They were assembled to 412 unigenes（including 107 contigs and 305 unique sequences）.By BLASTN,28.24% of the P.xylostella ESTs had no hit or hitted on unknown gene.71.76% ESTs were subjected to annotated and categorized by their putative functions,including immunity-related genes,metal ion binding and processing,nucleic acid and protein metabolism and process,cell signaling,cell structure,shape and mobility,energy metabolism,stress and detoxification and other genes.GeneOntology revealed that 39 immunity-related genes potentially involved in innate immunity and were subdivided into 4 categories： recognition molecules（such as peptidoglycan recognition protein,gram negative bacteria-binding protein）,proteases and protease inhibitors（such as trypsin-like serine proteinase and serpin）,antimicrobial effectors（antimicrobial peptide,lysozyme and prophenoloxidase-activating proteinase,etc） and other genes.qRT-PCR results showed that the diamond-back moth peptidoglycan recognition protein gene,imaginal disk growth factor gene,gloverin gene,lysozyme gene,prophenoloxidase-activating proteinase 3 gene and transferrin gene were up-regulated after immunization with bacteria and fungus.The expression profiles of these genes showed difference after being induced by bacteria and fungus,indicating they play different biological roles in immunity.These results showed that P.xylostella infected by P.cicadae could express serials of special genes and the response of P.xylostella to P.cicadae infection was a complex process.ESTs categorization analyses allowed us to identify 39 considered candidate immune genes,providing new insights into the molecular mechanisms during the innate immunity of P.xylostella against P.cicadae infection.
出处
《中国生物防治学报》
CSCD
北大核心
2012年第1期32-40,共9页
Chinese Journal of Biological Control
基金
浙江省自然科学基金(Y3110597)
浙江省农业科学院创新提升工程项目(2010R26Y01D02)
浙江省科技厅重大科技专项农业项目(2008C12044-3)
温州市科技局重大项目(N2005A018)
国家公益性行业(农业)科研专项(201103021)
关键词
小菜蛾
蝉拟青霉
天然免疫
SSH文库
表达序列标签
诱导表达
Plutella xylostella
Paecilomyces cicadae
innate immunity
SSH library
expressed sequence tags（ESTs）
induced expression
