7株不同原核质粒表达丙型肝炎核心蛋白的差异研究  

Differential research of the seven different prokaryotic expression plasmid expressing the CORE protein of hepatitis C

作　　者：汪明珊[1] 颜学兵[2] 周林福[3] 李杨霞[3] 

机构地区：[1]浙江省人民医院感染科,浙江杭州310014 [2]徐州医学院附属医院感染病科,江苏徐州221000 [3]浙江大学医学院附属第一医院传染病研究所,浙江杭州310000

出　　处：《中国现代医生》2012年第33期9-12,共4页China Modern Doctor

基　　金：江苏省自然科学基金(BK2007031)

摘　　要：目的通过比较不同原核质粒表达丙型肝炎核心蛋白(CORE)的差异,筛选出能高效表达CORE的原核质粒。方法用PCR扩增并获得CORE基因,扩增产物分别构建到7种不同原核表达质粒中,重组质粒用IPTG诱导表达,聚丙烯凝胶电泳(SDS-PAGE)初步鉴定表达产物,并经Western blot验证。结果 PCR及双酶切验证表明CORE基因成功构建到7种原核质粒中,SDS-PAGE验证各融合质粒表达目的蛋白存在差异,其中pTrc-CKS-CORE及Pmbp-C-CORE可见相应的融合蛋白表达,以pTrc-CKS-CORE表达量高,表达产物经Western blot验证表达成功。结论成功构建7株不同CORE原核表达质粒,各融合质粒表达CORE存在较大差异,筛选出高表达CORE质粒,为下一步研究奠定基础。Objective Hepatitis C virus （HCV） is closely correlated with hepatic cirrhosis and liver cancer. The core protein （CORE） whice is the structural protein of HCV plays an important role in the pathogenesis of HCV. We constructed a prokaryotic plasmid which could efficiently express the CORE. Methods We amplified and obtained the CORE gene by PCR, than constructed the products into seven different prokaryotic plasmids after restriction enzyme digestion. Selected the positive clones and transformed them into E.coli Top 10F, induced by IPTG to obtained the CORE, identified the expression products by polyacrylamide gel electrophoresis （SDS-PAGE）, and verified by western blot. Results The results of PCR and restriction enzyme digestion Showed that the CORE gene was successfully constructed into seven different kinds of prokaryotic plasmids, the results of SDS-PAGE verified that different plasmids expressed different CORE, pTrc-CKS- CORE expressed high levels of proteins, which were tested by Western-blot. Conclusion We successfully constructed seven CORE expressing plasmids, of which the expression efficiency and the products are quite different. The highly efficient plasmid is selected for the further studies.

关 键 词：丙型 肝炎病毒核心蛋白 质粒构建 

分 类 号：R512.6[医药卫生—内科学]