摘要
目的构建稳定表达由COL1A1启动子驱动的增强型绿色荧光蛋白 (EGFP)的ROS1 7/2 .8细胞株。方法用PCR方法从大鼠基因组DNA中扩增出长为 3.6Kb的COL1A1 (Ⅰ型胶原α1链基因 )启动子 ,并克隆到T载体 ;然后将COL1A1启动子分段酶切 ,获得不同长度的启动子片段 ;与报告基因EGFP相连接 ,构建成真核表达载体 ;随后转染ROS1 7/2 .8细胞系 ,再用G41 8筛选。结果得到了稳定转染COL1A1 EGFP基因的ROS1 7/2 .8细胞株。结论 这些细胞株的建立为研究微重力对COL1A1启动子活性及成骨相关基因表达的影响奠定基础。
Objective To obtain ROS17/2.8 cell lines which were stably expressing EGFP reporter gene drived by COL1A1 promoter. Method A 3.6 Kb COL1A1 promoter from rat was cloned into pMD-18-T vector by PCR. This amplified promoter vector was digested to get several different length fragments which were then fused with EGFP reporter gene to construct eukaryotic expression vectors. ROS17/2.8 cell was stably transfected with these vectors by LipofectAMINE TM and selected by G418. Result The COL1A1-EGFP stably transfected cell lines were established. Conclusion The cell lines will be useful for studying the effects of microgravity on the activity of COL1A1 promoter and expression of gene related with bone form.
出处
《航天医学与医学工程》
CAS
CSCD
北大核心
2004年第2期107-110,共4页
Space Medicine & Medical Engineering
基金
国家高技术研究发展计划 ( 863计划 )资助项目( 863 2 2 10 3 )
