猪瘟E2基因多克隆抗体的制备及其特性分析

Preparation and Characteristics of Polyclonal Antibody Against E2 Gene of Classical Swine Fever Virus

以CSFV FJ-1株为模板对猪瘟E2基因的主要抗原表位进行PCR扩增,得到大小为370bp的目的片段,将其连接到pET-32a表达载体,转化BL21后利用IPTG诱导其表达,SDS-PAGE和Western blot分析表明,目的蛋白成功获得表达,大小约33.5ku融合蛋白。利用Ni-NTA agarose纯化蛋白、重组蛋白与弗氏完全佐剂充分乳化后免疫新西兰兔,多克隆抗体效价经间接ELISA检测可高达1∶6400。Western blot结果表明该重组蛋白具有良好的反应原性,试验结果表明已成功获得猪瘟E2基因的多克隆抗体且能识别CSFV FJ-1毒株。

The DNA fragment of Classical Swine Fever Virus E2 gene was amplified by PCR from the CSFV FJ-1. The fragment was inserted into expression vector p ET32 a. The recombinant expression plasmid was transferred into E. colis strain BL21,and the high level expressed recombinant protein was detected in the inclusion body protein of the expression strain induced by IPTG. The recombinant protein was highly purified by Ni-NTA agarose.Then the purified recombinant protein was injected into rabbits to obtain polyclonal sera. E2-specific antibody titer was up to 1∶ 6400 by indirect ELISA,The result of Western blot analysis showed that the recombinant protein had good antigenicity. The results showed that the CSFV E2 polyclonal antibodies were successfully obtained and can specially identify the FJ-1 strain.