利用Red重组工程技术解决外源基因在大肠杆菌染色体lac操纵子中的表达.pdfVIP

生 物 工 程 学 报 Chin J Biotech 2008, April 25; 24(4): 576-580 journals.im.ac.cn Chinese Journal of Biotechnology ISSN 1000-3061 cjb@im.ac.cn © 2008 Institute of Microbiology, CAS CSM, All rights reserved 研究简报 Red lac 李山虎, 施庆国, 黄翠芬, 周建光 , 100850 : 为了应用 Red 重组工程技术实现外源基因在大肠杆菌染色体上的表达, 寻找染色体上外源蛋白的稳定高效表 达位点, 使用Red 重组工程系统和kan/sacB 无痕迹修饰技术, 将易于定量分析的荧光素酶报告基因替换DY330 染色体 lac 操纵子中的lacZ 基因。检测该位点的表达效率结果显示: 大肠杆菌染色体上lac 操纵子能够高效稳定表达外源基因, 初步证明了染色体可以作为外源蛋白或抗原的表达载体, 不会影响细菌的生长繁殖。 : 重组工程, 同源重组, 荧光素酶报告基因, 原核表达 Heterologous Genes Expression on Escherichia coli Chromosome lac Operon Using Red Recombination Shanhu Li, Qingguo Shi, Cuifen Huang, and Jianguang Zhou Institute of Biotechnology , Academy of Military Medical Sciences , Beijing 100850, China Abstract: To achieve efficient and stable expression of heterologous exogenetic protein or antigen in E. coli chromosome, the luciferase report gene was knocked in lacZ site of chromosome lac operon by using Red recombination system and selection-counterselection kan/sacB technology. The quantitative analysis of exogenous gene expression indicated that the target gene could be efficiently expressed at lacZ site of lac operon. The results confirmed the efficient screening and stable expression of heterologous protein or antigen on chromosome by using the recombinant engineering technique. This study demonstrated that the chromosome could be used as a vector for heterologous protein or antigen and the stable expression of exogenous gene on E. coli chromosome had no side effect on the bacterial growth and propagation. Keywords: recombination, homologous recombination, luciferase report