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为获得高表达人锰超氧化物歧化酶(MnSOD)的工程菌。从肿瘤组织中提取总RNA ,利用逆转录聚合酶链反应扩增人MnSOD的cDNA ,将克隆的基因片段插入表达载体pET2 8a(+ )内的NcoI/EcoRI位点 ,转化大肠杆菌BL2 1(DE3 ) ,用PCR及SDS -PAGE的方法筛选高表达人MnSOD的工程菌。结果构建成功高表达人MnSOD的工程菌 ,表达的人MnSOD相对分子质量约为 2 2 0 0 0 ,表达量约占菌体蛋白的 3 0 %。为大量制备人MnSOD和进一步的应用研究奠定了基础。
To obtain high expression of human manganese superoxide dismutase (MnSOD) engineering bacteria. Total RNA was extracted from the tumor tissue, and cDNA of human MnSOD was amplified by RT-PCR. The cloned gene fragment was inserted into the NcoI / EcoRI site in the expression vector pET2 8a (+) and transformed into E. coli BL21 ), PCR and SDS-PAGE method of screening highly expressed human MnSOD engineering bacteria. Results The recombinant human MnSOD was constructed successfully. The relative molecular mass of human MnSOD was about 20 000 and the expression level was about 30% of the bacterial protein. It laid the foundation for mass preparation of MnSOD and further applied research.