结核分枝杆菌Rv2994基因重组穿梭质粒的构建与鉴定  

Construction and Identification of Recombinant Shuttle-plasmid Bearing Rv2994 Gene from Mycobacterium Tuberculosis

作　　者：赵明才[1] 鲍朗[2] 吴悦涵[2] 张会东[2] 

机构地区：[1]川北医学院附属医院风湿免疫研究所,四川南充637007 [2]四川大学华西医学中心感染与免疫研究室,四川成都610041

出　　处：《川北医学院学报》2006年第3期211-214,共4页Journal of North Sichuan Medical College

基　　金：国家自然科学基金(30271172)

摘　　要：目的构建结核分枝杆菌假想药物外排泵蛋白编码基因Rv2994的重组穿梭质粒pMV-2994,并对其进行鉴定。方法以结核分枝杆菌H37Rv基因组为模板,应用PCR技术扩增Rv2994基因编码序列,定向克隆入融合蛋白原核表达载体pGEX-1λT,获得重组表达质粒pGEX-2994,测序鉴定。酶切该基因片段,定向重组入大肠杆菌-分枝杆菌穿梭载体pMV261,构建重组质粒pMV-2994,通过限制性内切酶酶切及PCR鉴定。结果从结核分枝杆菌H37Rv株基因组DNA中扩增出的Rv2994基因与GenBank公布的序列一致,经过酶切及PCR鉴定,表明Rv2994基因成功插入了pMV261穿梭载体。结论成功构建了重组穿梭质粒pMV-2994,为进一步研究Rv2994基因功能奠定了基础。Objective To construct and identify a recombinant shutfle-plasmid bearing Rv2994 gene of Mycobacterium tuberculosis. Methods The Rv2994 gene of Mycobacterium tuberculosis H37Rv strain was amplified by PCR and cloned into prokaryotic expression vector pGEX-1λT. The recombinant plasmid pGEX-2994 was transformed into E. coli JM109 to sequence. The recombinant shuttleplasmid was constructed by correctly inserting Rv2994 gene cut from pGEX-2994 into pMV261 and identified by restriction endonuclease and PCR. Results The Rv2994 gene was amplified accurately from the genome DNA of H37Rv. A recombinant shuttle-plasmid bearing Rv2994 was constructed. Conclusion A recombinant shuttle-plasmid bearing Rv2994 was successfully constructed. It provided the basis for the further study of the gene Rv2994.

关 键 词：结核分枝杆菌 Rv2994基因 穿梭载体 

分 类 号：R52[医药卫生—内科学]