猪链球菌乳酸脱氢酶基因的原核表达及重组蛋白的反应原性研究  

Prokaryotic Expression of Lactate Dehydrogenase Gene of Streptococcus suis Serotype 7 and Reactionogenicity of Recombinant Protein

作　　者：高明明[1] 王淑杰[2] 王俊 金家民[2] 张璐[2] 刘永刚[2] 李爱东[2] 陶冶[2] 李莉[2] 姜成刚[2] 王刚[2] 申红[1] 

机构地区：[1]石河子大学动物科技学院,石河子832003 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/动物疫病诊断及技术服务中心,哈尔滨150001 [3]北京市畜牧总站,北京100107

出　　处：《石河子大学学报（自然科学版）》2015年第2期188-191,共4页Journal of Shihezi University(Natural Science)

基　　金：国家自然科学基金项目(31060281);国家科技重大专项(2012ZX10004214)

摘　　要：为了验证双向电泳筛选出的7型猪链球菌WH分离株免疫相关蛋白乳酸脱氢酶的反应原性,本实验采用克隆表达的方法对其进行研究。根据质谱鉴定的NCBI登录号查找乳酸脱氢酶基因序列,并设计特异性引物,对乳酸脱氢酶基因进行扩增,克隆入p ET30a载体,构建重组质粒p ET30a-LDH,测序后转化入大肠埃希氏菌中。用IPTG进行诱导重组菌,SDS-PAGE和Western blot分析表达产物。SDS-PAGE分析结果显示表达的重组融合蛋白相对分子量为41ku;Western blot分析结果显示该重组蛋白可与7型猪链球菌WH分离株多克隆抗体发生特异性血清学反应。这表明表达的重组LDH蛋白具有良好的反应原性,为预防和控制猪链球菌病提供有效的疫苗候选分子奠定基础。To verify the reactogenici ty of immune-related protein lactate dehydrogenase （LDH） which was screened from Streptococcus sois type 7 WH isolate by two-dimensional electrophoresis.According to LDH gene sequences accessed in NCBI, a pair of specific primers was desigend to amplify LDH gene.The amplified product was cloned into expression vector pETB0a for expressing.Recombinant plasmid pET30a-LDH was transformed into E.coli.The expression products were analysed by SDS-PAGE and Western blot after IPTG induction.SDS-PAGE results confirmed that the recombinant fusion protein was expressed with a relative molecular weight of 41 ku;western blot analysis showed that the recombinant protein could specifically be reactive to S.suis type7 WH polyclonal antibody,which confirmed that the recombinant LDH protein was endowed with good reactionogenicity.The study provides an effective vaccine candidate molecule to prevent and control S.sois disease.

关 键 词：猪链球菌 乳酸脱氢酶 反应原性 原核表达 

分 类 号：S852.61[农业科学—基础兽医学] R378.1[农业科学—兽医学]