【摘要】 通过计算机分析设计,人工合成了寡核苷酸引物,并优化了翻译起始区,应用PCR及重组DNA技术,获得了重组人白细胞介素6(IL—6)高效表达工程菌。从全菌SDS—PAGE考马斯亮蓝染色后薄层密度扫描分析表达产率为38.6％,分子量为2.1×10~4。以IL—6依赖性的小鼠杂交瘤细胞株B9用MTT法测定表达重组人IL—6的HGF活性为2×10~7U/L菌液。不同菌株表达重组人IL—6研究表明,以E.coli DH5α/pBV IL—6表达产率最高。更多还原

【Abstract】 By means of PCR amplification,site-specific mutagenesis,DNA recombination in vitro and optimization of translation initiation, we have constructed a recombinant expression plasmid pBVIL-6 which highly produces biologically active human Interleukin-6 (rhIL-6) in E. coli . The expressed rhIL-6 with molecular weight 21kD accounted for 38. 6% of the E. coli proteins. The biological activity of the expressed rhIL-6 was 2 X 107 U/L bacterial culture and 1 X 107 U/L bacterial culture in E. coli DH5a and in E. coli RRI,respectively. They were measured with an IL-6-de-pendent B-cell hybridoma line B9. E. coli DH5a,one of the three E. coli in our experiments, was the most suitable one for high expression of the rhIL-6.更多还原