摘要
为获得猪瘟病毒(CSFV)E0重组蛋白,建立CSFV抗体快速检测方法。本研究通过扩增CSFV C株E0基因,亚克隆至原核表达载体pGEX-4T-2,构建重组原核表达质粒pGEX-4T-2-E0,转化至宿主菌Rostta(DE3)后进行诱导表达。SDS-PAGE检测表明与目的蛋白大小一致,其分子质量大小为46ku。优化重组蛋白表达条件,最终获得诱导表达的最佳温度为37℃,最佳IPTG浓度为0.1mmol/L,并且目的蛋白以包涵体形式存在。Western blot表明,目的蛋白能与CSFV兔化高免血清反应,表明该融合蛋白具有良好的抗原性。本研究为CSFV抗体检测试剂盒的研究奠定了基础。
In order to gain the recombinant protein E0 of the classical swine fever virus（CSFV）,to estab-lish CSFV antibody rapid detection method,the CSFV C strain E0 was amplified and sub-cloned into pro-karyotic expression plasmid pGEX-4T-2 vector.The recombinant plasmid named pGEX-4T-2-E0 was transformed into E.coli Rosetta （DE3）and induced by IPTG.The expression products were analyzed by SDS-PAGE,a specific expression band,in form of inclusion body with a molecular weight 46.0 ku was de-tected.The optimum temperature was 18℃,the IPTG was 1.0 mmol/L.Western-blot indicated that the expressed protein was recognized not only by the rabbit antibody to CSFV,but also with the specific mon-oclonal antibody of GST tag.The result suggests that the recombinant protein has good antigenicity and provides substantial base for the CSFV antibody detection kit research.
出处
《动物医学进展》
CSCD
北大核心
2014年第5期31-35,共5页
Progress In Veterinary Medicine
基金
河南省基础与前沿技术研究项目(122300410003)
河南省自然科学基金研究项目(330002)(122300410003)
河南省教育厅科学技术研究重点项目(13A230837)
