钙整合素结合蛋白CIB真核表达载体的构建及在NIH3T3细胞中的表达  

Construction of eukaryotic expression of CIB vectors and its expression in NIH3T3 cells

作　　者：惠玲[1] 王晓辉[1] 袁碧波[2] 杨金开[1] 吕同德[1] 惠斌[3] 蔡文琴[2] 

机构地区：[1]兰州军区兰州总医院医学实验中心,兰州730050 [2]第三军医大学基础医学部神经生物学教研室,重庆市神经科学研究所,重庆400038 [3]复旦大学上海医学院药理研究中心,上海200032

出　　处：《第三军医大学学报》2007年第3期216-218,共3页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目青年基金(30400125)~~

摘　　要：目的构建钙整合素结合蛋白(calcium-andintegrin-bindingprotein,CIB)的真核表达载体,了解它在真核细胞中的表达及定位。方法从人脑组织分离总RNA,采用RT-PCR获得CIB开放读框序列,克隆至真核表达载体pCMV-myc,构建表达带myc表达标签的真核表达载体pCMV-myc-CIB,将其转染至NIH3T3细胞中,免疫组化鉴定其在细胞中的表达及定位。结果序列分析表明克隆的CIB序列与GeneBank报道的序列一致,脂质体转染入NIH3T3细胞中,用myc抗体检测到目的蛋白myc-CIB的表达。结论pCMV-myc-CIB表达载体构建成功,并在真核细胞中得到了正确表达,CIB蛋白主要定位于细胞质中。Objective To construct the expression vector of calcium-and integrin-binding protein （ CIB）, and study its localization as a fusion protein in NIH3T3 cell. Methods Total RNA was extracted from human brain tissue for CIB sequence with RT-PCR, and then the obtained fragment was ligated into pCMV-myc vector. After confirmation with sequence analysis, the expression vector pCMV-myc-CIB was transformed into the NIH3T3 cell by Lipofectamin 2000. The protein expression was observed by immunofluorescent staining. Re- suits DNA sequence analysis showed that the CIB cDNA had been successfully inserted into pCMV-myc expression vector, myc-CIB was expressed in the NIH3T3 cells transfected with pCMV-myc-CIB. Conclusion The eukaryotic expression vector containing CIB is successfully constructed and expressed in the cytoplasma of NIH3T3 cells.

关 键 词：CIB 基因克隆 真核表达 

分 类 号：Q513[生物学—生物化学] Q782