棉蚜His-CYP6J1融合蛋白的分离纯化及多克隆抗体的制备

发布时间：2018-03-09 05:25

本文选题：P　切入点：CYPJ融合蛋白　出处：《生物技术通报》2017年05期 　论文类型：期刊论文

【摘要】：通过亲和层析法纯化棉蚜His-CYP6J1融合蛋白,制备及鉴定其多克隆抗体。采用镍离子金属螯合亲和层析柱分离纯化棉蚜His-CYP6J1融合蛋白,SDS-PAGE检测纯化产物。用纯化得到的His-CYP6J1融合蛋白免疫小鼠,制备抗棉蚜P450CYP6J1多克隆抗体;采用ELISA法检测抗体效价;免疫组化法检测抗血清特异性。结果表明,纯化的His-CYP6J1融合蛋白免疫小鼠后得到多抗血清,ELISA法检测抗血清效价达1∶200 000。免疫组化结果显示,此多克隆抗体能够与棉蚜P450 CYP6J1天然蛋白特异性结合,却在棉铃虫没有发现相应的结合反应。上述结果对研究棉蚜单一P450蛋白结构、功能及其在棉蚜抗药性形成过程的作用奠定基础。
[Abstract]:His-CYP6J1 fusion protein of cotton aphid was purified by affinity chromatography. The polyclonal antibody was prepared and identified. The purified product of His-CYP6J1 fusion protein of cotton aphid (APHIS gossypii) was separated and purified by nickel ion metal chelate affinity chromatography. The purified His-CYP6J1 fusion protein was used to immunize mice to prepare the polyclonal antibody against APHIS gossypii P450 CYP6J1. The antibody titer was detected by ELISA method, and the specificity of antiserum was detected by immunohistochemical method. The results showed that after immunizing mice with purified His-CYP6J1 fusion protein, the titer of antiserum was 1: 200000 by Elisa. The polyclonal antibody could specifically bind to the natural protein P450 CYP6J1 of cotton aphid, but no corresponding binding reaction was found in cotton bollworm. Function and its role in the formation of resistance to cotton aphids laid the foundation.
【作者单位】：新疆大学生命科学与技术学院新疆生物资源基因工程重点实验室;
【基金】：自治区青年科技创新人才培养工程项目(qn2015yx001) NSFC-新疆联合项目(U1603331)
【分类号】：S433.39

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