【摘要】 大多数枯草杆菌载体在表达外源基因时会出现分离或结构不稳定,而将目的基因整合至染色体上的基因整合方法,近年来得到人们的普遍关注。通过构建一套整合载体,将β-环状糊精葡基转移酶基因随机整合至枯草杆菌1A289的染色体上,从表达氯霉素抗性及β-CGTase阳性株中选得菌株Bs2,其氯霉素抗性为5γ/ml,β-CGTase酶活为266u/ml。后又经多次整合及更高浓度氯霉素抗性的选择,获得一株Bs16-7-7菌株,在无选择压力下,表现出极为稳定的遗传表型,氯霉素抗性为125γ/ml,β-CGTase酶活达13000u/ml以上。更多还原

【Abstract】 Most expression vectors for B. subtlis may occour segrigational and/or structural instability in the expression of heterologous genes. A new alternative, integration of production genes into the genome, is gained more attention.By constructing a set of integration vectors,the β-CGTase gene was randomly integrated into the genome of B.subtilis 1A289. The integration strain Bs2 which showed 266u/ml β-CGTase activity and Cmr phenotype was selected. The number of integrated β-CGTase gene copies was further increased by repeating the process of integration and by amplifying the gene with Cm selection. Strain Bs16-7-7 tolerating up to 125μg/ml of Cm showed a stable phenotype when grew without selection pressure.Its β-CGTase activity reached to 13000u/ml.更多还原