抑制葡萄糖氧化酶基因降低黏虫幼虫对苏云金芽孢杆菌侵染的抗性  

Inhibition of glucose oxidase gene decreases the resistance of Mythimna separata(Lepidoptera:Noctuidae)larvae to Bacillus thuringiensis infection

作　　者：杨航 杨洪佳 张雅男 王晓曦 樊东[1] YANG Hang;YANG Hong-Jia;ZHANG Ya-Nan;WANG Xiao-Xi;FAN Dong(College of Agronomy,Northeast Agricultural University,Harbin 150030,China)

机构地区：[1]东北农业大学农学院,哈尔滨150030

出　　处：《昆虫学报》2022年第7期843-851,共9页Acta Entomologica Sinica

基　　金：现代农业产业技术体系(CARS-04)。

摘　　要：【目的】本研究旨在探讨葡萄糖氧化酶(glucose oxidase,GOX)在黏虫Mythimna separata发育、消化和免疫防御中的作用。【方法】采用转录组测序技术克隆了黏虫GOX基因的cDNA序列。qRT-PCR检测该GOX基因在黏虫不同发育阶段(1-6龄幼虫、蛹和1日龄成虫)和4龄第1天幼虫不同组织(前肠、中肠、后肠、下唇腺、马氏管、脂肪体和表皮)、以及取食用不同浓度(0.01%,0.1%,1%和10%)葡萄糖溶液浸泡10 s的玉米叶片及饥饿和再取食条件下的4龄第1天幼虫中的特异性表达模式。通过RNAi和生物测定研究该GOX基因在黏虫对苏云金芽孢杆菌Bacillus thuringiensis抗性中的功能。【结果】克隆获得了一条新的长2187 bp的黏虫GOX基因MsG OX cDNA序列(GenBank登录号:KY348779),其开放阅读框长1821 bp,编码606个氨基酸,预测分子量为66.4 kD。发育表达谱结果表明MsG OX在黏虫不同发育阶段表达量不同,在4龄幼虫期表达量最高;组织表达谱结果表明MsG OX在黏虫4龄第1天幼虫各组织中均有表达,在下唇腺中表达量最高。饲喂幼虫不同浓度葡萄糖对MsG OX转录的诱导效果不同,在葡萄糖浓度为10%时,该基因的表达水平达到最高。随着饥饿时间的延长,4龄第1天幼虫中MsG OX的表达水平逐渐升高,并在饥饿24 h时达到高峰,饥饿后再用玉米叶片饲喂幼虫,MsG OX的表达水平逐渐回升。注射ds MsG OX 48 h后,MsG OX的表达水平较对照(注射ds EGFP)降低了88.3%;与同一时间点的对照相比,注射ds MsG OX后48和72 h幼虫的体重、体长和消化系数均显著降低,苏云金芽孢杆菌侵染48和72 h提高了幼虫的校正死亡率。【结论】MsG OX可能参与了黏虫中肠的消化和抗菌,研究结果为进一步研究黏虫MsG OX的功能并提出新的黏虫防治策略提供了依据。【Aim】The objective of this study is to explore the roles of glucose oxidase(GOX)in the development,digestion,and immune defense of Mythimna separata.【Methods】The cD NA sequence of GOX gene from M.separata was cloned by transcriptome sequencing technology.The specific expression patterns of this GOX gene in different developmental stages(1st-6th instar larvae,pupae and 1-day-old adults)and tissues(foregut,midgut,hindgut,labial gland,Malpighian tubules,fat body,and integument)of the day-14th instar larvae,and the day-14th instar larvae of M.separata fed with maize leaves soaked in different concentrations(0.01%,0.1%,1%,and 10%)of glucose solution for 10 s and under starvation and refeeding conditions were detected by qRT-PCR.The role of this GOX gene in the resistance of M.separata to Bacillus thuringiensis was explored by RNAi and bioassay.【Results】A 2187 bp cD NA sequence of a novel GOX gene named MsGOX(Gen Bank accession no.:KY348779)was obtained from M.separata.It has the open reading frame of 1821 bp in length,encoding a 606-amino acid polypeptide with the predicted molecular weight of 66.4 kD.Developmental expression profile revealed that MsGOX showed different expression levels in M.separata at different developmental stages,with the highest expression level at the 4th instar larval stage,and tissue expression profile showed that MsGOX was expressed in various tissues of the day-14th instar larvae,with the highest expression level in labial glands.MsGOX transcription could be induced differently by feeding larvae with different concentrations of glucose.The expression level of MsGOX reached the highest when the glucose concentration was 10%.The expression level of MsGOX in the day-14th instar larvae increased gradually with the starvation time increasing,and reached the peak at 24 h after starvation.When the larvae were refed with maize leaves after starvation,the expression level of MsGOX gradually increased.At 48 h after injection of ds MsGOX,the expression level of MsGOX was inhibited by 88.3%

关 键 词：黏虫 葡萄糖氧化酶 表达模式 RNA干扰 苏云金芽孢杆菌 

分 类 号：Q966[生物学—昆虫学]