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目的:构建真核表达质粒pcDNA3/gD,并在体外COS-7细胞中进行表达。方法:体外PCR扩增出gD糖蛋白基因,克隆入真核表达质粒pcDNA3中。PCR、酶切和测序证实插入的gD基因正确后,转染COS-7细胞,用原位ELISA鉴定表达蛋白。结果:构建了pcDNA3/gD真核表达质粒,经原位ELISA证实,转染COS-7细胞表达的gD蛋白能和gD单克隆抗体ID3和DL11结合,证明该蛋白具有天然gD蛋白的抗原性。结论:构建的重组真核表达质粒pcDNA3/gD能够在COS-7细胞中表达。
Objective: To construct eukaryotic expression plasmid pcDNA3 / gD and express it in COS-7 cells in vitro. Methods: gD glycoprotein gene was amplified by in vitro PCR and cloned into eukaryotic expression plasmid pcDNA3. PCR, digestion and sequencing confirmed that the correct inserted gD gene was transfected into COS-7 cells, and the expressed protein was identified by in situ ELISA. Results: The pcDNA3 / gD eukaryotic expression plasmid was constructed and confirmed by in situ ELISA. The gD protein expressed in COS-7 cells was able to bind with gD monoclonal antibody ID3 and DL11, which proved that the protein has the antigenicity of natural gD protein. Conclusion: The recombinant eukaryotic expression plasmid pcDNA3 / gD can be expressed in COS-7 cells.