Skp2过表达载体的构建及表达  

Construction of Skp2 Eukaryotic Expression Vector

作　　者：曲璇[1] 张芮[2] 李军[1] 左艳[1] 万文涛[1] 马晓军[1] 袁银娜 韩洛川[1] 史海龙[1] 申亮亮[3] 

机构地区：[1]陕西中医药大学,陕西咸阳712046 [2]青岛大学附属医院,山东青岛266003 [3]第四军医大学,陕西西安710032

出　　处：《生物技术通讯》2016年第6期830-833,共4页Letters in Biotechnology

摘　　要：目的:在HEK293细胞中获得Skp2全长基因,构建其高效真核表达载体。方法:提取人HEK293细胞总RNA,反转录合成c DNA,以特异性引物扩增Skp2全长基因并克隆入p Babe载体,筛选阳性克隆进行序列测定后,鉴定其表达情况。结果:PCR扩增得到特异性的约1300 bp的片段,序列测定结果表明与国外已发表的序列完全一致;将构建的p Babe-Skp2质粒转入HEK293细胞,与空质粒对照相比,Skp2的m RNA和蛋白表达水平均提高,并且p27蛋白水平降低。结论:构建获得Skp2基因高效真核表达载体,将用于Skp2的功能研究。Objective： To clone the Skp2 full length gene from human HEK293 cells and construct its eukaryoticexpression vector. Methods： Total RNA was extracted from cultured human HEK293, and the Skp2 c DNA frag-ment was obtained by RT-PCR with two specific gene primers, and then it was inserted into p Babe vector to con-struct the expression vector. Results： The 1300 bp specific fragment was obtained, and the sequence of whichwas consistent with published details. Compared with the empty plasmid control, the increased m RNA and proteinlevel of Skp2, and the decreased p27 were detected in the plasmid p Babe-Skp2 transformed HEK293 cells. Conclusion： The Skp2 expression vector will be used in the further studies of the roles of Skp2.

关 键 词：SKP2 基因扩增 真核表达载体 

分 类 号：Q78[生物学—分子生物学]