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将BamHⅠ酶切的人脑髓鞘碱性蛋白(MBP)基因cDNA克隆片段和表达载体pGEX-5T重组后转化大肠杆菌,再筛选、增殖和IPTG诱导。结果显示:阳性克隆10%SDS-PAGE有2条特异区带(49kd和35kd);Western印迹杂交证实该区带具MBP抗原特异性;蛋白质含量测定、免疫斑点杂交和ELISA分析,该膜蛋白表达量占菌体裂解液蛋白质总量6%(50mg/L菌液)。
The cDNA fragment of human MBP gene was digested with BamHI and transformed into Escherichia coli with the expression vector pGEX-5T. The recombinant plasmid was transformed into Escherichia coli and then screened, proliferated and induced by IPTG. The results showed that there were two specific bands (49kd and 35kd) on 10% SDS-PAGE of positive clones. Western blotting confirmed the specificity of MBP antigen in this region; protein content, immunofluorescence and ELISA analysis Amount accounted for 6% of the total bacterial lysate protein (50mg / L bacteria).