构建多顺反子表达载体系统的新策略  

New Strategy for Construction of Polycistron Expression Vector

作　　者：沈俊杰[1] 单娟娟[1] 骆菁菁[1] 刘立[1] 钱程[1] 

机构地区：[1]浙江理工大学生命科学学院,杭州310018

出　　处：《浙江理工大学学报（自然科学版）》2009年第4期561-566,共6页Journal of Zhejiang Sci-Tech University(Natural Sciences)

基　　金：863项目(2006AA02Z126);973计划(2004CB518804);浙江省科技厅重点项目(No.2006C23006)

摘　　要：利用pSM155-GFP载体为骨架质粒,构造IRES-2A联用结构,成功实现该结构向2A结构的简单切换,得到可以同时表达shRNA和多基因、可切换多功能的真核表达载体。克服目前在单基因或多基因功能机制的研究中,普遍需要构建多个真核表达载体才能完成的研究过程。并进一步将p53基因克隆至载体中,通过多种方法证实构建得到的IRES-2A和2A元件均能有效介导多顺反子的表达,且表达的蛋白具有完好的生物活性。In this study, using pSM155-GFP vector as a backbone, construct a mammalian expression vector, employing IRES-2A combined structure which can switch to 2A structure easily. This multifunetional and switchable vector can co-express shRNA and multiple genes, and avoid constructing several eukaryotie expression vectors involving in a series of subcloning procedures. Employed p53 as model gene and its expression is detected by appropriate methods including fluorescenee microscopy and Western blot. The results show that IRES-2A and 2A elements can mediate efficient functional expression of polycistron.

关 键 词：多顺反子 IRES 2A SHRNA 

分 类 号：R73-362[医药卫生—肿瘤]