论文部分内容阅读
双酰基肼类杀虫剂模拟天然蜕皮激素作用影响幼虫蜕皮。昆虫蜕皮激素受体的高度敏感性和专一性要求必须建立新的杀虫活性检测技术,以适应快速准确和大批量筛选的要求。本研究采用RT-PCR技术,获取斜纹夜蛾Spodoptera litura蜕皮激素受体(EcR)与超气门蛋白(USP)功能域目的基因,构建EcR、USP功能区基因原核表达载体(pEHISEGFPTEV-EcRcde和pEHISEGFPTEV-USPcde)。载体经诱导表达和蛋白纯化,获得EcR和USP功能区纯化蛋白。在蛋白浓度l mg/mL,3H-PonA终浓度8 nmol/L的条件下,采用放射性配基受体结合分析测定了4种药剂(虫酰肼、呋喃虫酰肼、抑食肼和灭幼脲)不同浓度下的放射性比活的变化。结果显示:随着药剂浓度的逐渐增大,前3种药剂的放射性比活都有不同程度的降低,其中虫酰肼的放射性比活降低程度最大,其次是呋喃虫酰肼和抑食肼,灭幼脲的放射性比活基本无变化。这些结果表明相同条件下虫酰肼比呋喃虫酰肼和抑食肼有更高的杀虫活力,本研究的方法可对双酰基肼类杀虫剂或者先导化合物进行初步筛选。
Bisacylhydrazine insecticides mimic natural ecdysone effects on larval ecdysis. The high sensitivity and specificity of insect ecdysone receptors has necessitated the establishment of new insecticidal activity detection techniques to meet the requirements for rapid, accurate and high-volume screening. In this study, the target genes of ecdysteroid (ERR) and the USP domain of Spodoptera litura were obtained by RT-PCR and the prokaryotic expression vectors of EcR and USP functional regions (pEHISEGFPTEV-EcRcde and pEHISEGFPTEV- USPcde). The vector was induced to express and protein purified to obtain EcR and USP purified protein. Under the conditions of 1 mg / mL protein and 8 nmol / L 3H-PonA, the radiopharmaceutical receptor binding assay was used to determine the inhibitory effects of four insecticides (tebufenozide, furazolidone hydrazide, Urea) at different concentrations of radioactive specific activity changes. The results showed that with the increasing of the concentration of the drug, the specific radioactivity of the first three kinds of medicaments decreased to some extent, the tebufenozide had the highest specific activity of radioactivity, followed by furazolidone hydrazide, The radiobiological activity of diflubenzuron remained almost unchanged. These results indicate that tebufenozide has higher insecticidal activities than furazolidone and imazapyr under the same conditions. The method of this study can be used for preliminary screening of biacylhydrazine insecticides or lead compounds.