摘要
目的:研究丙肝病毒非结构蛋白基因3(ns3基因)在体外真核细胞中的诱导表达。方法:以PCR方法从含HCVns3基因的重组载体pBns3中扩增出ns3基因,并将其插入到克隆载体pGEM-T中,再与表达载体pMSG重组,以得到重组表达载体pMSG-ns3。然后采用磷酸钙沉淀法将其转染哺乳动物CHO细胞,经地塞米松(DM)诱导,采用ELISA及Western-blot技术检测NS3蛋白的表达水平。结果:用所构建的诱导型表达载体pMSG-ns3转染CHO细胞,经DM(3×10-8mol/L)诱导后可表达NS3蛋白,其浓度与时间呈正相关。结论:所得到的表达载体在培养细胞中能够有效地表达。
Objective: To study the inducible expression of hepatitis C virus ns3 gene (HCV ns3) in eukaryotic cells. Methods: The ns3 gene was obtained from plasmid pBns3 by polymerase chain reaction and inserted into the cloning vector pGEMT. Then, the ns3 was subcloned into the vector pMSG to generate dexamethasone (DM)inducible expression plasmid pMSGns3. CHO cells were transfected by pMSGns3 by using calcium phosphate precipitated method and cultivated for 12 to 24 h.The transfected cells were induced with DM and the transient expression of NS3 protein was analyzed by using ELISA and Westernblot methods. Results: After treatment with 3×10^(-8) mol/L DM,the expression of NS3 was observed in the transfected CHO cells. A slightly higher level of NS3 was shown with the delay time of DM treatment. Conclusion: The inducible expressing vector pMSGns3 might be helpful for further studies of the characterizations of the ns3 gene in vivo.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
1998年第4期307-310,共4页
Academic Journal of Second Military Medical University
基金
国家自然科学基金