人细小病毒B19中国株结构蛋白VP1的克隆及表达  

Clone and expression of VP1 unique region gene segment of Chinese strain of human parvovirus B19

作　　者：陈彩平[1] 张国成[1] 许东亮[1] 汪萍[1] 李茹英[1] 黄莹[1] 盛凯[1] 李飚[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》2004年第5期385-387,共3页Journal of Medical Postgraduates

基　　金：国家自然科学基金资助项目 (批准号 :3 9870 0 2 1)

摘　　要：目的 :构建人细小病毒B1 9中国株结构蛋白VP1基因片段的重组表达载体 ,探索大量表达VP1蛋白的最佳条件。　方法 :从该科保存的人细小病毒B1 9中国株结构蛋白VP1基因片段的重组克隆载体中获得VP1基因片段 ,将该片段亚克隆入表达载体pQE 30 ,构建VP1 pQE 30原核表达载体 ,并转化至感受态大肠杆菌M1 5中 ,给予不同浓度诱导剂异丙基硫代 β D 半乳糖苷 (IPTG)及在不同温度下进行诱导表达。　 结果 :成功构建了融合蛋白VP1 pQE 30的重组表达质粒 ,表达的融合蛋白经 1 5 %SDS PAGE电泳证实其相对分子质量为 2 2 0 0 0。IPTG浓度为 1mmol/L ,诱导表达 5h ,37℃下无表达 ,2 5℃下有表达 ;采用不同的IPTG浓度诱导表达 5h ,0 .0 5mmol/L至 1mmol/L均有表达 ,表达率相近 ,为 (1 6± 1 ) %。　结论 :获得人细小病毒B1 9中国株结构蛋白VP1基因片段的原核表达载体及其产物 。Objective: To construct the recombinant expressive vector of VP1 gene of Chinese strain of human parvovirus B19,and to obtain the expression of the structural protein VP1. Methods:VP1 gene segment was obtained from the recombinant clone vector of VP1 gene kept in our laboratory and sub cloned into expressive vector pQE 30. The recombinant expressive vector of VP1 gene was constructed and transformed into E. coli M15. IPTG was used to induce the expression of protein. Different conditions were taken and SDS PAGE was used to seek out the best way in which the protein was expressed. Results: Recombinant expressive plasmid VP1 pQE 30 was constructed. 15% SDS PAGE showed the molecular weight of the expressed fusion protein was about 22 000. The protein could be expressed under 25℃,but not under 37℃. Different concentrations of IPTG, from 0.05 mmol/L to 1 mmol/L, could induce the expression of the protein and the expressive ratio was similar, which was about (16±1)%. Conclusion: The prokaryotic expressive vector and the fusion protein of human parvovirus B19 VP1 gene of Chinese strain were obtained. It is important in the further purification of human parvovirus B19 VP1 protein and the preparation of the monoclonal antibody against VP1 structural protein and the B19 vaccine.

关 键 词：人细小病毒B19 VP1基因片段 载体构建 诱导表达 

分 类 号：Q754[生物学—分子生物学]