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为探讨SARS CoV的M蛋白的免疫学特性以及M蛋白作为SARS CoV病毒疫苗组分的可行性和必要性。分别用pET 15b和pET 22b在大肠杆菌中表达SARS CoV的M蛋白,亲和层析纯化后作为抗原应用。同时,将M蛋白的编码基因克隆进分泌型真核表达载体pSecTagB中得到重组质粒pSecM作为DNA疫苗,免疫BALB/c小白鼠、制备SARS CoV M蛋白的抗血清。并用纯化后的M蛋白建立的SARS CoV M抗体ELISA检测技术研究所构建的M DNA疫苗的免疫效果。结果表明:两种重组M蛋白在大肠杆菌中均以可溶性形式得到高效表达,经与华大产的用灭活SARS全病毒制备的SARS CoV抗体ELISA检测试剂盒比较实验,证明该原核表达的重组M蛋白能与SARS确诊病人血清以及M DNA免疫鼠血清发生特异性抗原抗体反应。这两种重组M蛋白有可能作为抗原组分用于临床SARS CoV检测中;所构建的SARS CoV的M基因核酸疫苗能在小鼠体内产生特异性抗体,提示M蛋白在SARS CoV疫苗尤其是组分疫苗的研制中应加以考虑,为DNA疫苗的开发提供了依据。
To investigate the immunological properties of M protein of SARS CoV and the feasibility and necessity of using M protein as vaccine component of SARS CoV virus. The M protein of SARS CoV was expressed in E. coli using pET 15b and pET 22b, respectively, and was purified by affinity chromatography and used as an antigen. At the same time, the gene encoding M protein was cloned into secretory eukaryotic expression vector pSecTagB to obtain the recombinant plasmid pSecM as a DNA vaccine and BALB / c mice were immunized to prepare anti-SARS CoV M protein antiserum. The immune effect of M DNA vaccine constructed by purified M protein was tested by ELISA assay of SARS CoV M antibody. The results showed that both of the recombinant M proteins were efficiently expressed in soluble form in E. coli. Compared with the ELISA kit for detection of SARS CoV antibody produced by China SARS virus produced by China, it was proved that the recombination of prokaryotic expression M protein can react with the serum of patients with SARS and the serum of M DNA immunized mice to produce specific antigen and antibody. The two recombinant M proteins may be used as antigen components in clinical SARS CoV detection; the constructed SARS CoV M gene nucleic acid vaccine can produce specific antibodies in mice, suggesting that the M protein in the SARS CoV vaccine, especially the group Sub-vaccine development should be considered for the development of DNA vaccine provided the basis.