棉铃虫中肠谷胱甘肽S-转移酶(GST)对新疆一枝蒿黄酮提取物和萜类提取物的响应

[目的]研究新疆一枝蒿提取物黄酮及萜类提取物对棉铃虫解毒酶谷胱甘肽S-转移酶(GST)的影响,探明外源有毒物质对该解毒酶表达规律的影响,阐明棉铃虫与植物次生物质相互作用的机制,为发展新型无公害植物源农药提供理论依据.[方法]从新疆一枝蒿中分离提纯黄酮与萜类提取物,以室内饲养棉铃虫(3龄幼虫)为对象,利用生物测定、实时荧光定量PCR及酶联免疫吸附测定法,分别测定新疆一枝蒿中黄酮与萜类提取物对棉铃虫的毒杀效果及其在mRNA水平和蛋白水平上对GST表达的影响.[结果]黄酮和萜类提取物对棉铃虫3龄幼虫在24 h时的半致死浓度LC50分别是21.4和4.1 mg/100 g,在48 h时的半致死浓度LC50分别是19.3和3.1 mg/100 g；实时荧光定量PCR结果表明GST在mRNA水平的表达量受到诱导,时间效应为当黄酮和萜类提取物浓度分别为20和2 mg/100 g时,GST在12 h时能够被两种物质显著诱导表达,而在24 h时表达受到抑制；浓度效应为在18h时,黄酮和萜类提取物浓度分别为20和2 mg/100 g时显著诱导GST的表达,而在40和4 mg/100 g时抑制GST的表达；酶联免疫吸附测定结果表明,萜类和黄酮在时间和浓度效应上都能显著诱导GST蛋白表达,并且蛋白水平在时间与浓度效应中的表达趋势与mRNA水平相类似.[结论]GST对外源物质新疆一枝蒿的黄酮提取物和萜类提取物存在积极响应,低浓度或短时间内黄酮和萜类均能诱导GST的过表达,从而化解外源毒性,而浓度过高或时间过长则抑制其表达.

Response of GST in the Cotton Bollworm(Helicoverpa armigera) to the Flavones Extracts and Terpenes Extracted from Artemisia rupestris

[Objective]In order to elucidate the roles of detoxification enzyme glutathion S - transferase (GST) between cotton boll worm and biochemicals,we investigated the expression patterns of GST in the midgut from the cotton bollworm induced by flavones and terpenes extract from Artemisia rupestris,which will provide a clue to develop a new pollution -free botanical insecticide.[Method]The toxicity of the flavones ex- tracts and terpenes extracts from A.rupestris to the third larvae of cotton bollworm were assayed by bioassay. The GST expression patterns at mRNA level and protein level were detected by real - time quantitative PCR (qPCR) and ELISA,respectively.[Result]1.The LC_(50) at 24 h of flavones extracts and terpenes extracts to bollworm were 20.41 and 4.1 mg/100 g,respectively;the LC_(50) at 48 h were 19.3 and 3.1 mg/100 g;respectively. 2.qPCR results showed that the GST mRNA levels in the midgut of the larvae were induced by these two extracts.For the time effect,when the larvae were exposed to 20 mg/100 g flavones extracts and 2 mg/100 g terpenes extracts respectively for 12 h the relative expression level of GST mRNA in the midgut was over - expressed,while at 24 h the expression was inhibited;For the concentration effect,when the larvae were treated with these two exyracts for 18 h respectively,the relative expression level of GST mRNA was over -expressed by 20 mg/100 g flavones and 2 mg/100 g terpenes extracts,respectively,while the expression was inhibited by 40 mg/100 g flavones and 4 mg/100 g terpenes extrcts in the thet,respectively;3.ELISA results showed similar trends in the GST expression at the protein level to the mRNA.[Conclusion]The GST adopted an active response to the flavones and terpenes extracts from Artemisiar upestris:GST was induced to over - express when the cotton bollworm third larvae were treated with flavones extracts or terpenes extracts from A.upestris either at low concentrations or for short time,but was inhibited at high concentrations or for long time.