铜绿假单胞菌外膜蛋白F与HSV-1VP22融合DNA疫苗的构建与表达

目的:构建铜绿假单胞菌外膜蛋白F与Ⅰ型单纯疱疹病毒(herpes simplex virus 1,HSV-1) VP22融合表达的DNA疫苗,并研究其在真核细胞中的表达情况.方法: 构建重组质粒 pVAX1-OprF、 pVAX1-VP22、 pVAX1-VP22-OprF、 pVAX1-OprF-VP22.原核表达VP22 的碳端蛋白,以电洗脱的方法得到纯化蛋白,免疫BALB/c小鼠,制备VP22蛋白的抗血清,West-ern blot鉴定抗体特异性.最后,以脂质体法将重组质粒转染真核细胞,利用Western blot检测其在真核细胞中的表达情况.结果: 成功制备了抗VP22的抗血清,重组质粒在真核细胞中得到了表达.结论: 铜绿假单胞菌OprF与 HSV-1VP22融合表达的重组质粒能够在真核细胞中表达,这为进一步的动物实验打下了基础.

Construction and expression of fusion vaccine DNA of Pseudomonas aeruginosa outer membrane protein F and HSV-1VP22

Objective:To construct the DNA vaccine for the outer membrane protein F from Pseudomonas aerufinosa and VP22 from herpes simplex virus 1 fusion gene, and to study its express in eukaryotic cells. Methods: The eukaryotic expression vectors pVAX1-OprF、pVAX1-VP22、pVAX1-VP22-OprF and pVAX1-OprF-VP22 were constructed. The VP22 carboxy-terminal was expressed in prokaryotic cells and purified by electroelution. Polyclonal antibody was developed by immunizing BALB/c mice with purified recom-binant protein. The specificity of BALB/c mice antiserum was measured by Western blot. The recombinant plasmids were transfected in-to eukaryotic cells by LipofectamineTM. The expressions of recombinant eukaryotic vectors were investigated by Western blot. Results:The polyclonal antibody against VP22 protein was successfully prepared. The recombinant vectors could be expression in eukaryotic cells. Conclusion:The eukaryotic expression vector for the outer membrane protein F from Pseudomonas aerufinosa and VP22 from her-pes simplex virus 1 fusion gene can express in eukaryotic cells,which may paves a way for further studies in animals.