根癌农杆菌介导的赭曲霉遗传转化体系的建立  

Establishment of Agrobacterium tumefaciens-mediated Transformation System of Aspergillus ochraceus

作　　者：崔倩 李洁[1] 刘晓光[1] 

机构地区：[1]天津市工业微生物重点实验室、工业发酵微生物教育部重点实验室、天津科技大学生物工程学院,天津300457

出　　处：《生物技术通报》2014年第6期199-204,共6页Biotechnology Bulletin

摘　　要：赭曲霉在工业上用于甾体C-11α羟基化。为了对现有赭曲霉生产菌株进行遗传改造,以农杆菌LBA4404作为侵染菌株,以赭曲霉(TCCC41060)作为受体菌株,以潮霉素B基因作为筛选标记,成功建立了根癌农杆菌介导的赭曲霉遗传转化体系并对其转化效率的主要影响因素,如农杆菌浓度、孢子浓度、共培养时间和温度等进行了分析。在最佳条件下,转化效率可以达到57个转化子/108个分生孢子。随机挑选的8个阳性转化子10代连续培养结果表明所获得的转化子能稳定遗传。该遗传转化体系的建立为赭曲霉菌株的定向遗传改造提供了重要的操作平台。Filamentous fungus Aspergillus ochraceus TCCC41060 is an industrial strain used for microbial steroid C11-αhydroxylation. In order to create genetic tools for A. ochraceus, the transformation system of A. ochraceus was established using Agrobacterium tumefaciens-mediated system（ATMT）. A. tumefaciens LBA4404 was used as the infective strain and hygromycin B gene as the selection marker. The major factors affecting the transformation efficiency including the concentrations of A. tumefaciens cell and A. ochraceus spores, co-culture time, and co-culture temperature were investigated. Under the best conditions, the transformation efficiency of 57 transformants/107 spores was obtained. A. ochraceus transformants were confirmed by PCR amplification . Randomly selected 8 transformants were shown to be genetically stable after 10 rounds of successive cultivation. Results indicated this ATMT transformation system would provide important means for the directed genetic manipulation of this important industrial strain A. ochraceus TCCC41060.

关 键 词：赭曲霉 根癌农杆菌 转化体系 

分 类 号：Q93[生物学—微生物学]