棉铃虫β-catenin的多克隆抗体制备及在蛹脑中的表达研究

Preparation of polyclonal antibody and expression analysis of β- catenin in pupal brains from Helicoverpa armigera

β-catenin是Wnt/β-catenin信号通路的重要成员。本研究构建β-catenin的原核表达载体,成功地在大肠杆菌Escherichia coli中进行表达。镍柱纯化重组蛋白后免疫兔子,制备了抗棉铃虫β-catenin的多克隆抗体。ELISA和Western blot的方法检测抗体效价和抗体特异性,结果表明抗体的效价较高,特异性较好。最后应用该抗体调查了滞育和发育蛹脑中β-catenin的表达情况,从蛋白水平证实了发育蛹脑中β-catenin高于滞育蛹脑。本研究为后续研究棉铃虫β-catenin的功能以及Wnt/β-catenin信号通路在棉铃虫发育中的作用奠定了基础。

β- catenin is a key component of Wnt / β- catenin signaling pathway. In this study,β-catenin was cloned into prokaryotic expression vector and the recombinant protein was expressed successfully in Escherichia coli. After purified by Ni column,the recombinant protein was used to immune New Zealand rabbit. The antibody titer was estimated as high as 1 ∶ 256000 dilution ration detected by ELISA,and this antibody was of good specificity. Finally,the expression levels of Har- β-catenin were investigated by Western blot using this antibody,and the results confirmed that Har- β-catenin was significantly higher expressed in non- diapause- pupal brains than in diapause- destined brains. This study lays a foundation for further investigation the function of β- catenin and Wnt / β-catenin in H. armigera pupal development.