野桑蚕细胞色素P450氧化酶活性测定及CYP9家族基因的诱导转录

Cytochrome P450 oxidase activity determination and induced-transcription of CYP9 family genes in Bombyx mandarina

采用酶标板酶活性动力学法检测了氯氰菊酯诱导苏大野桑蚕中肠和脂肪体的PNOD活性。结果表明:氯氰菊酯诱导24 h,脂肪体、中肠PNOD活性分别增加了18.7%,12.2%。根据野桑蚕CYP9A20基因(GenBank登录号:FJ378716)、CYP9A21基因(GenBank登录号:FJ265741)、CYP9A22基因(GenBank登录号:EF535806)和CYP9G3基因片段设计引物,用半定量RT-PCR方法分析了氯氰菊酯诱导野桑蚕CYP9家族基因的表达水平。结果表明,氯氰菊酯诱导24 h,CYP9A21基因在中肠的mRNA表达量是对照的2.1倍,CYP9A20,CYP9A21,CYP9G3基因在脂肪体的mRNA表达量是对照的1.9,3.5,1.4倍。推测野桑蚕CYP9A21等基因的过量表达可能导致野桑蚕对氯氰菊酯氧化解毒代谢的增强。

Enzyme linked immunosorbent assay was used to determine the activity of PNOD in the midgut and fat body of Bombyx mandarina under cypermethrin treatment.The results showed that after 24 h induction by cypermethrin,the PNOD activity of fat body and midgut increased by 18.7% and 12.2%,respectively.According to the sequence of CYP9A20（GenBank No.FJ378716）,CYP9A21（GenBank No.FJ265741）,CYP9A22（GenBank No.EF535806） and CYP9G3 gene segments of Bombyx mandarina,the specific primers were designed.The expression levels of CYP9 family genes of Bombyx mandarina induced by cypermethrin were determined by semi-quantitive RT-PCR.The results indicated that after 24 h induction,the relative mRNA level of CYP9A21 in midgut was 2.1 times of the control,and the relative mRNA levels of CYP9A20,CYP9A21,CYP9G3 in fat body were 1.9,3.5 and 1.4 times of the control,respectively.The over expression of these cypermethrin P450 genes might enhance the oxidation and detoxification capacity of Bombyx mandarina to cypermethrin.