人细小病毒B19 XA株VP1蛋白原核表达及初步鉴定  

Expression and verification of VP1 protein of human parvovirus B19-XA strain in E. coli

作　　者：李小青[1] 张国成[1] 许东亮[1] 李志宏[1] 

机构地区：[1]第四军医大学西京医院儿科,陕西西安710032

出　　处：《医学研究生学报》2008年第11期1124-1127,共4页Journal of Medical Postgraduates

基　　金：国家自然科学基金资助项目(批准号:30571948)

摘　　要：目的:通过DNA重组技术,构建B19病毒XA株VP1全长基因表达载体,诱导重组VP1融合蛋白表达。方法:自B19病毒感染患者血清中提取病毒DNA为模板,用PCR法扩增编码VP1蛋白的全长基因片段,以pET28(a)为表达载体,构建重组表达质粒pET28(a)-VP1,转化E.coli.BL21(DE3),获得重组工程菌株。经IPTG诱导培养获得高效表达的VP1融合蛋白,SDS-PAGE、免疫印迹鉴定表达产物;并以该重组融合蛋白为抗原,免疫家兔,ELISA法检测所获抗体效价。结果:①获得了含重组表达质粒pET28(a)-VP1的正相阳性工程菌株,经IPTG诱导能高效表达VP1融合蛋白。②经ELISA法检测抗VP1多克隆抗体效价为1∶12 800。结论:重组工程菌可表达VP1融合蛋白,且该蛋白具有良好的免疫原性,对诊断试剂制备及疫苗研制具有重要意义。Objective： To construct a recombinant expression vector containing the VP1 whole gene of human parvovirus （HPV） B19-XA strain by DNA recombinant technology and to induce the expression of the VP1 fusion protein in E. coll. Methods ： The gene of interest was amplified by PCR from the viral DNA genome extracted from the patient infected with B19 virus, and inserted into the pET28 （a） expression vector. The positive reeombinants pET28 （a） -VP1 were transformed into E. coli BI221 （ DE3）, and then the VP1 proteins were expressed after induced by IPTG and analyzed by SDS-PAGE and immunoblotting. In addition, rabbits were immunized with the recombinant VP1 fusion proteins as antibodies, and their valence was detected by ELISA. Results： The positive bacteria strains containing the recombinant expressive vector pET28（ a）-VP1 were constructed successfully, and IPTG induced a high expression of the VP1 fusion proteins. The valence of muhiclone antibodies of anti-VP1 proteins was 1：12 800. Conclusion ： The VP1 fusion proteins of the HPV B19-XA strain can be expressed efficiently in E. coli, and these proteins possess satisfactory immunogenicity and play an important role in the preparation of diagnostic reagents and the development of vaccines.

关 键 词：人细小病毒B19 VP1蛋白 表达载体 免疫原性 

分 类 号：Q786[生物学—分子生物学]