【摘要】 目的:在构建重组真核表达载体pIRESneo2/mIL-2的基础上,建立能够持续稳定表达mIL-2的哺乳类工程细胞。方法:运用分子克隆技术,将由RT-PCR获得的mIL-2cDNA片断插入真核表达质粒pIRESneo2构建成mIL-2重组表达载体pIRESneo2/mIL-2。通过脂质体转染法将pIRESneo2/mIL-2导入C2C12细胞。转染后第30天,用Westernblots检测mIL-2表达情况。结果:经DNA测序证明mIL-2cDNA片断插入方向和碱基组成顺序均准确无误,Westernblots检测转染真核重组表达载体pIRESneo2/mIL-2的C2C12细胞系表达mIL-2。结论:利用pIRESneo2/mIL-2构建的真核表达载体在C2C12细胞系中能够持续稳定表达mIL-2。更多还原

【Abstract】 Objective:To construct a eukaryotic expressive plasmid that codes mIL-2 gene and test its expression in C_ 2 C_ 12 cells.Methods:mIL-2 gene was inserted into eukaryotic expression vector named pIRESneo2 by enzymolysis and ligation, and at the same time, the stop code of mIL-2 cDNA was taken away. It was confirmed by restrictive enzymes (Age I/BamH I) digestion and by analysis of DNA sequencing. The C_ 2 C_ 12 cells were transfected with the recombinant plasmid and the expression of recombinant mIL-2 was examined by Western blot.Result:DNA sequencing shows that the recombinant plasmid pIRESneo2/mIL-2 has been successfully constructed. And the expression of recombinant mIL-2 was testified in C_ 2 C_ 12 cells by Western blot. Conclusion:A eukaryotic engineering cell line expressing mIL-2 steadily has been established successfully.更多还原