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目的 探讨γ干扰素在大肠杆菌中的表达及纯化方法 ,为下一步批量生产提供实验依据。方法 应用DNA重组技术把中国人淋巴细胞mRNA反转录产物IFN -γcDNA克隆到原核表达质粒 pBV2 2 0上 ,构建出pBVIFN高效表达载体 ,转化大肠杆菌 plyss。通过温度诱导 ,高效表达IFN -γcDNA。然后 ,经过包涵体溶解、复性、浓缩及DEAE离子交换层析 ,使γ干扰素纯化。结果 IFN -γcDNA表达水平占菌体可溶性蛋白的 40 4% ,包涵体纯度为 70 % ,总生物活性为 1 2 7× 10 9U ,比活性达 1.31× 10 9U/L蛋白 ,活性回收率为 77.7%。结论 本方法为γ干扰素的批量生产奠定了基础
Objective To investigate the expression and purification of IFN - γ in Escherichia coli and provide experimental evidence for mass production in the next step. Methods The recombinant human IFN-γ cDNA of lymphocyte mRNA was cloned into the prokaryotic expression vector pBV220 using DNA recombinant technology to construct pBVIFN highly efficient expression vector and transformed into E. coli plyss. IFN-γ cDNA is highly expressed by temperature induction. Then, the interferon is purified by inclusion body lysis, renaturation, concentration and DEAE ion exchange chromatography. Results The expression level of IFN-γcDNA was 40 4%, the purity of inclusion body was 70%, the total bioactivity was 12 7 × 10 9 U, the specific activity was 1.31 × 10 9 U / L, the activity recovery was 77.7 %. Conclusion This method laid the foundation for the mass production of IFN-γ