杀虫剂辛硫磷诱导家蚕脂肪体蛋白的表达特征及SerB基因的组织转录活性分析  

Protein Expression Profiles of Fat Body and Tissue-specific Transcriptional Activity of SerB Gene in Bombyx mori Induced by Pesticide Phoxim

作　　者：葛君[1] 王举梅[1] 李兵[1,2] 沈卫德[1,2] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123 [2]现代丝绸国家工程实验室,江苏苏州215123

出　　处：《蚕业科学》2010年第3期428-434,共7页ACTA SERICOLOGICA SINICA

基　　金：国家重点基础研究发展计划"863"项目(No.2006AA10-A118);江苏省高校自然科学基金项目(No.07KJB230102)

摘　　要：为了从蛋白质水平探讨家蚕对有机磷杀虫剂的抗性机制,采用双向电泳结合基质辅助激光解析电离飞行时间质谱技术,分析家蚕5龄中期幼虫在添食杀虫剂辛硫磷前后脂肪体蛋白的表达特征。经ImageMaster6.0软件分析表明,对照组共检测到465个蛋白点,实验组检测到396个蛋白点,其中能匹配的蛋白点有348对,匹配率为73.54%。对特征蛋白进行质谱分析,已鉴定的蛋白包括磷酸丝氨酸转氨酶(SerB)和5'-甲硫腺苷磷酸化酶(MTAP)。采用半定量RT-PCR分析家蚕5龄幼虫添食辛硫磷后脑、中肠、丝腺、脂肪体、精巢、马氏管等6个组织器官中SerB基因mRNA的转录水平,结果与对照组相比均呈现上调趋势,其中在脑组织的转录水平上调尤为明显,在脂肪体中的转录水平上调与其蛋白的表达上调一致。受辛硫磷诱导的影响,家蚕5龄中期幼虫脂肪体的蛋白数目减少,SerB基因在各组织器官增量表达,推测可能与蚕体对农药的代谢解毒作用有关。To study the resistant mechanism of Bombyx mori against organophosphorus pesticides at the protein level, high resolution two-dimensional polyacrylamide gel electrophoresis followed by matrix-assisted laser desorption ionization and time of flight mass spectrometry was employed to analyze the protein expression profiles of fat body from the middleaged 5th larvae before and after treatment with phoxim. The ImageMaster 6.0 analysis showed that 465 protein spots were detected in the control group and 396 ones in the experimental group. A total of 348 protein spots were matched to each other, showing a matching rate of 73.54%. Mass spectrometry analysis revealed that the differential protein spots included phosphoserine aminotransferase （SerB） and 5＇-methylthioadenosine phosphorylase （MTAP）. Semi-quantitative RT-PCR analyses on transcriptional levels of SerB gene in six tissues, namely brain, midgut, silk gland, fat body, testis, and Malpighian tubules, of the 5th instar larvae treated with phoxim showed that the expressions of SerB gene were all up-regulated, among which the up-regulation in brain tissue was especially obvious. The transcriptional level of SerB gene was consistent with the up-regulation of its protein expression in fat body. Being affected by phoxim＇s induction, the number of proteins expressed in fat body of the 5th instar larvae was reduced, and the expression levels of SerB gene in six tissues were increased, sug-gesting an Key words nvolvement of this protein Bombyx mori; Fat body Semi-quantitative RT-PCR n the detoxification of pesticide in Bombyx mori larvae.

关 键 词：家蚕 脂肪体 磷酸丝氨酸转氨酶基因 辛硫磷 双向电泳 半定量RT—PCR 

分 类 号：S884.96[农业科学—特种经济动物饲养] Q78[农业科学—畜牧兽医]