家蚕谷胱甘肽-S-转移酶基因(BmGSTd1)的诱导表达定量分析  

Induced Expression and Quantitative Analysis of Bombyx mori Glutathione S-transferase Gene BmGSTd1

作　　者：赵国栋[1] 卫正国[1,2] 李兵[1,2] 沈卫德[1,2] 

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

出　　处：《蚕业科学》2010年第1期46-51,共6页ACTA SERICOLOGICA SINICA

基　　金：国家重点基础研究发展计划"973"项目(No.2005CB-121005)

摘　　要：谷胱甘肽-S-转移酶(GST)在机体的解毒代谢和抗氧化中起重要作用。为探究家蚕谷胱甘肽-S-转移酶基因(BmGSTd1)在家蚕体内的解毒和抗性功能,采用实时荧光定量PCR方法,对BmGSTd1基因在正常饲养及添食NaF家蚕5龄幼虫不同组织中的转录水平进行检测,并采用家蚕Actin3、GAPDH、28S rRNA 3种内参照基因对检测结果进行归一化处理。BmGSTd1基因在家蚕5龄幼虫各组织的转录水平存在差异,且采用不同内参照基因结果不一致:分别以家蚕Actin3、GAPDH、28S rRNA基因为内参照,该基因转录水平最高的组织分别为中肠、丝腺、脂肪体。5龄第2天幼虫添食NaF对体内各组织中BmGSTd1基因的转录水平具有诱导作用,且在不同组织中诱导表达的情况不同,采用以上3种内参照基因进行归一化处理的结果数据表明,中肠和脂肪体组织中的诱导转录水平最高,可能与这2种组织是家蚕主要的解毒器官有关。研究认为,检测基因在不同组织中的转录水平,采用合适的内参照基因对保证检测结果的可靠性非常重要。Glutathione S-transferases（GSTs） play important roles in detoxifying insecticides and antioxidation.In order to explore the roles of Bombyx mori glutathione S-transferase gene（BmGSTd1） in detoxification and resistance to insecticides,we used real-time fluorescent quantitative PCR method to detect the transcription levels of BmGSTd1 in different tissues of the 5th instar larvae feeding on both normal mulberry leaves and sodium fluoride treated mulberry leaves.The detection results were normalized by using 3 Bombyx mori internal reference genes namely Actin3,GAPDH and 28S rRNA.The results indicated that the transcriptional levels of BmGSTd1 were different in various tissues of the 5th instar larvae.They were also varied while different internal reference genes were used： the highest transcriptional levels were observed in tissues of midgut,silk gland and fat body while Actin3,GAPDH and 28S rRNA genes were used as the internal reference,respectively.Transcription of BmGSTd1 gene in various tissues could be induced by sodium fluoride applied onto mulberry leaves from the 2nd day of the 5th instar.And the expression varied in different tissues.The normalized data with the above 3 internal reference genes indicated that the expression of BmGSTd1 gene was the highest in mid-gut and fat body,suggesting their involvement in detoxification of sodium fluoride.It is thus concluded that,for detection of gene transcriptional levels,it is very important to choose adequate internal reference genes so as to ensure the reliability of the detection results.

关 键 词：谷胱甘肽-S-转移酶基因 家蚕组织 诱导表达 实时荧光定量PCR 内参照基因 

分 类 号：S881.2[农业科学—特种经济动物饲养] Q78[农业科学—畜牧兽医]