哺乳动物细胞高效表达载体的优化  

Optimization of High Expression Vectors in Mammalian Cells

作　　者：张国强[1] 刘志刚[1] 刘珊[1] 俞炜源[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071

出　　处：《生物技术通讯》2006年第5期692-695,共4页Letters in Biotechnology

基　　金：国家高技术研究发展计划项目(2001AA215351)

摘　　要：目的:优化哺乳动物细胞表达系统,提高目的基因的表达效率。方法:以组织型纤溶酶原激活剂(tPA)为报告基因,利用本实验室建立的CHOfrt/dhfr-细胞定点整合表达系统,对多种表达调控元件(包括hCMV和hEF-1α启动子、hCMV增强子、hEF-1α1st内含子及翻译增强子H213和V163等)及其多种组合的表达效率进行了系统的比较和评价。结果:hCMV启动子与H213组合以及hEF-1α启动子与V163组合的表达效率分别是仅含hCMV启动子的156.6%和139.5%。结论:该研究为构建高效的哺乳动物细胞表达载体奠定了基础。Objective： To increase the expression level of foreign gene by the optimization of the expressional regulatory elements in the cassette of interesting gene. Methods： A series of expression vectors containing diverse combinations of expressional regulatory elements such as hCMV, hEF-1α promoter, hCMV enhancer, hEF-1α1st intron, translational enhancer H213 and V163 were constructed. And then the expression efficiency of these regulatory elements were, based on the assay of k2tPA （a mutant of tPA） activity by monitoring fibrin clearance, valuated by the use of the targeted integration system. Results： The results showed that the two new hybrid regulatory elements hCMV/H213 and hEF-1α/V163, in comparison with the only hCMV promoter, increased the expression efficiency by more than 55% and 39%, respectively. Conclusion： This finding revealed that the high expression of introduced gene in mammalian cells can be realized through the use of proper combinations of regulatory elements.

关 键 词：CHO-dhfr^-细胞 表达调控元件 优化 表达载体 

分 类 号：Q782[生物学—分子生物学]