1型鸭肝炎病毒E53株VP1基因的原核表达及多克隆抗体的制备
Prokaryotic Expression of VP1 Protein of duck hepatitis virus type 1 and preparation of polyclonal antibody
利用RT-PCR技术扩增1型鸭肝炎病毒(DHV-1)E53株VP1基因序列，将其连入pET-30a载体中，构建原核表达载体pET-30a-VP1，并转化至宿主菌Rosetta(DE53)plysS中，经IPTG诱导后，SDS-PAGE分析表明VP1蛋白在大肠杆菌中获得了表达，western blot表明表达产物具有良好的反应原性。将表达产物纯化后免疫新西兰大白兔，制备的多克隆抗体可特异性识别DHV-1，间接ELISA测定多克隆抗体效价达1:12800，中和实验测定多克隆抗体中和抗体效价达1:32。 本研究表达的蛋白和制备的多克隆抗体为VP1蛋白功能的进一步研究和诊断制剂的开发提供了重要数据和材料。
ccording to the genome sequence of duck hepatitis virus type 1 E53, VP1 gene was amplified by RT-PCR and cloned into prokaryotic expression vector pET-30a. After sequencing, the recombniant expression vector pET-30a-VP1 was transformed into the hostRosetta(DE53)plysS. The fusion protein His-VP1 was expressed after induced by IPTG. Western blot analysis showed that the fusion protein was expressed correctly. The rabbits were immunized with purified recombinant VP1 protein， western blot analysis showed that the sera from immunized rabbits had specific reaction with DHV-1. In addition, the antibody titer was 1:12800 by i-ELISA and neutralization test showed its neutralization antibody titer was up to 1:32. In this study, the expressed recombinant protein and polyclonal antibodies provided important data and material for the further study of VP1 protein function and the development of diagnostics.
王君伟、李鑫、刘琰、付培芬、马波
基础医学生物科学研究方法、生物科学研究技术微生物学
1型鸭肝炎病毒VP1原核表达免疫原性中和活性
duck hepatitis virus type 1VP1prokaryotic expressionimmunogenicityneutralization
王君伟,李鑫,刘琰,付培芬,马波.1型鸭肝炎病毒E53株VP1基因的原核表达及多克隆抗体的制备[EB/OL].(2012-03-14)[2025-04-28].http://www.paper.edu.cn/releasepaper/content/201203-472.点此复制
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