真核表达质粒pIRES_2-eGFP-Annexin A2的构建及其在293T细胞中的表达  

Construction of eukaryotic expression vector pIRES_2-eGFP-Annexin A2 and expression of Annexin A2 in 293T cells

作　　者：李娜[1] 周红[1] 俞颖[2] 王婷[1] 黄宏亮[1] 石文霞[1] 王海波[1] 

机构地区：[1]江苏大学基础医学与医学技术学院,江苏镇江212013 [2]浙江省中医院检验科,浙江杭州310006

出　　处：《江苏大学学报（医学版）》2009年第1期46-49,F0003,共5页Journal of Jiangsu University：Medicine Edition

基　　金：国家自然科学基金资助项目(30670907);浙江省自然科学基金资助项目(Y206830)

摘　　要：目的:构建含有人Annexin A2基因的真核表达质粒pIRES2-eGFP-Annexin A2,并在真核细胞293T中表达。方法:将质粒pCMV5-Annexin A2上Annexin A2基因酶切后与真核表达质粒pIRES2-eGFP连接,酶切鉴定及测序正确后,脂质体介导法转染293T细胞,荧光定量PCR及蛋白质印迹法检测其基因和蛋白的表达。结果:构建的质粒pIRES2-eGFP-Annexin A2转染293T细胞后,目的基因mRNA和蛋白表达均明显增高。结论:成功构建pIRES2-eGFP-Annexin A2质粒并表达蛋白,为研究Annexin A2的生物学功能奠定了基础。Objective： To construct pIRES2-eGFP-Annexin A2 eukaryotic expression vector carrying human Annexin A2 gene and express Annexin A2 protein in 293T cell.Methods： Annexin A2 gene digested from pCMV5-Annexin A2 was ligated into eukaryotic expression vector pIRES2-eGFP.After restriction analysis and sequencing,the recombinant plasmid pIRES2-eGFP-Annexin A2 was transfected into 293T cells in the mediation of liposome.The expression of Annexin A2 was analyzed by western blot and real-time PCR. Results： The expression of Annexin A2 at both mRNA and protein levels were significantly increased when transfected with pIRES2-eGFP-Annexin A2 in 293T cells. Conclusion： The eukaryotic expression vector pIRES2-eGFP-Annexin A2 was correctly constructed and the Annexin A2 protein was successfully expressed in 293T cells.This will facilitate the following study on Annexin A2.

关 键 词：ANNEXIN A2 真核表达 抗磷脂综合征 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学] Q343[医药卫生—基础医学]