Liu Ti, Wang Tianhong, Li Xian, Liu Xuan
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China.
Acta Biochim Biophys Sin (Shanghai). 2008 Feb;40(2):158-65. doi: 10.1111/j.1745-7270.2008.00388.x.
To improve heterologous gene expression in Trichoderma reesei, a set of optimal artificial cellobiohydrolase I gene (cbh1) promoters was obtained. The region from -677 to -724 with three potential glucose repressor binding sites was deleted. Then the region from -620 to -820 of the modified cbh1 promoter, including the CCAAT box and the Ace2 binding site, was repeatedly inserted into the modified cbh1 promoter, obtaining promoters with copy numbers 2, 4, and 6. The results showed that the glucose repression effects were abolished and the expression level of the glucuronidase (gus) reporter gene regulated by these multi-copy promoters was markedly enhanced as the copy number increased simultaneously. The data showed the great promise of using the promoter artificial modification strategy to increase heterologous gene expression in filamentous fungi and provided a set of optional high-expression vectors for gene function investigation and strain modification.
为提高里氏木霉中外源基因的表达水平,获得了一组优化的纤维二糖水解酶I基因(cbh1)启动子。删除了具有三个潜在葡萄糖阻遏物结合位点的-677至-724区域。然后,将修饰后的cbh1启动子的-620至-820区域(包括CCAAT框和Ace2结合位点)重复插入修饰后的cbh1启动子中,获得了拷贝数为2、4和6的启动子。结果表明,葡萄糖阻遏效应被消除,并且随着拷贝数同时增加,受这些多拷贝启动子调控的葡糖醛酸酶(gus)报告基因的表达水平显著提高。数据表明,使用启动子人工修饰策略在丝状真菌中增加外源基因表达具有很大的前景,并为基因功能研究和菌株改良提供了一组可选的高表达载体。