根癌农杆菌介导的烟曲霉几丁质合成酶chsC基因缺失突变体的构建  

The construction of the chsC-chitin synthase gene mutant of Aspergillus fumigatus by Agrobacterium tumefaciens-mediated transformation

作　　者：龙朝钦[1] 邓军[1] 李芹芥[1] 周村建[1] 郝飞[1] 

机构地区：[1]第三军医大学西南医院,重庆400038

出　　处：《中国感染控制杂志》2008年第2期84-87,共4页Chinese Journal of Infection Control

摘　　要：目的利用根癌农杆菌介导的基因转化定点插入突变几丁质合成酶chsC基因,实现基因失活而构建几丁质合成酶chsC基因缺失烟曲霉突变体。方法构建根癌农杆菌二元载体pDHt/SK,利用根癌农杆菌介导的转化插入突变烟曲霉几丁质合成酶chsC基因,用潮霉素抗性基因筛选转化子,并进行实时定量逆转录聚合酶链反应(RT-PCR)鉴定。结果潮霉素抗性基因通过同源重组成功插入烟曲霉基因组中;经RT-PCR鉴定,烟曲霉几丁质合成酶基因chsC表达失活。结论利用根癌农杆菌介导的基因转化可以定点插入突变并失活目的基因,此方法有可能成为烟曲霉基因转化和功能基因组研究的有力工具。Objective To construct the chitin synthase gene chsC- mutant of Aspergillus fumigatus by targeted disrupting and inactivating of chsC of Aspergillus fumigatus through Agrobacteriurn tume faciens-mediated transformation. Methods Binary vector pDHt/SK of Agrobacterium tumefaciens was constructed, chitin synthase gene chsC of Aspergillus fumigatus was disrupted by Agrobacterium tumefaciens-mediated transformation, and the positive transformant was selected by hygromycin resistance cassette and identified by RT-PCR. Results The hygromycin resistance cassette was integrated into genome of Aspergillus fumigatus by homologous recombination; the result of RTPCR showed that chitin synthase gene chsC was clearly inactivated. Conclusion Agrobacterium tumefaciens mediated transformation can disrupt and inactivate the target gene of Aspergillus furnigatus. This method would be an efficient tool for studying the gene transformation and function genome of Aspergillus fumigatus.

关 键 词：烟曲霉 根癌农杆菌 chsC基因 潮霉素抗性 

分 类 号：R379.6[医药卫生—病原生物学]