Sheng X Y, Li Y Y
Institute of Genetics, Fudan University, Shanghai, China.
Chin J Biotechnol. 1991;7(1):15-23.
Phage Blp7 DNA was digested with restriction enzyme and ligated to the restriction enzyme digested vector pTG402. The ligated mixture was used to transform competent cells of E. coli MC1061. Plasmid DNA was extracted from pooled transformants and competent cells of B. subtilis were transformed. By selecting yellow colonies upon spraying with catechol solution, 22 clones containing DNA fragments with promoter function were obtained. The promoter activity of 15 clones was determined by the color reaction of catechol-2,3-dioxygenase. The inserted fragment of the most potent promoter was mapped with restriction enzymes. CatO2 ase activity of two clones was measured in cells of B. subtilis of all growth phases and was found to increase rapidly at the end of the log-phase. It is inferred that these two promoters might be recognized by sigma 37.
噬菌体Blp7 DNA用限制酶消化，然后连接到经限制酶消化的载体pTG402上。连接混合物用于转化大肠杆菌MC1061的感受态细胞。从汇集的转化子中提取质粒DNA，并转化枯草芽孢杆菌的感受态细胞。通过用儿茶酚溶液喷雾后选择黄色菌落，获得了22个含有具有启动子功能的DNA片段的克隆。通过儿茶酚-2,3-双加氧酶的颜色反应测定了15个克隆的启动子活性。用限制酶对最强启动子的插入片段进行图谱分析。在枯草芽孢杆菌所有生长阶段的细胞中测量了两个克隆的CatO2酶活性，发现其在对数期末迅速增加。推测这两个启动子可能被σ37识别。
