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目的:研究腺相关病毒载体介导的人红细胞生成素在293细胞的体外表达Ⅰ方法:将人红细胞生成素cDNA克隆至AAV表达载体,采用无辅助病毒共转染法生产重组AAV-EPO病毒,利用RT-PCR和ELISA法检测hEpo的体外表达.结果:成功构建和生产了表达hEpo的重组AAV-EPO病毒.重组AAV-EPO病毒感染293细胞后,RT-PCR结果显示有hEpo基因表达,细胞上清中hEpo的表达水平达500U/L.结论:所制备的重组AAV-EPO病毒可以有效表达hEpo,可以进一步用于动物体内实验.
OBJECTIVE: To study the expression of human erythropoietin in 293 cells induced by adeno-associated virus vector.Methods: Human erythropoietin cDNA was cloned into AAV expression vector and recombinant virus-free AAV-EPO was produced by co-transfection without helper virus. The recombinant adeno-associated virus (AAV-EPO) expressing hEpo was successfully constructed and produced by RT-PCR and ELISA.Results of RT-PCR showed that hEpo gene was expressed in 293 cells, The expression level of hEpo in the supernatant reached 500 U / L. Conclusion: The recombinant AAV-EPO can express hEpo efficiently and can be used in vivo in animals.