酵母双杂交系统筛选FOXP3的相互作用蛋白  

Identification of interacting protein of FOXP3 by yeast two-hybrid system

作　　者：周丽娜[1] 吴军[1] 贺伟峰[1] 罗高兴[1] 陈希炜[1] 柏干苹[1] 袁顺宗[1] 张小容[1] 胡晓红[1] 

机构地区：[1]第三军医大学西南医院全军烧伤研究所,创伤,烧伤与复合伤国家重点实验室,疾病蛋白质组重庆市市级重点实验室,重庆400038

出　　处：《第三军医大学学报》2008年第24期2258-2261,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金(30500447)~~

摘　　要：目的利用酵母双杂交系统筛选与人FOXP3相互作用的蛋白。方法应用酵母双杂交系统,将成功构建的酵母诱饵表达载体pGBKT7-FOXP3,与转化了成人肝cDNA文库的酵母菌Y187接合,筛选与FOXP3相互作用的蛋白,通过一对一的回复性杂交实验排除假阳性,并对阳性克隆进行序列测定和生物信息学分析。结果经过酵母双杂交共获得最终确认3个阳性克隆,测序后经生物信息分析为肿瘤蛋白D52、分裂因子3b及未知蛋白。结论成功应用酵母双杂交系统筛选出与FOXP3相互作用的3个蛋白,它们可能与Treg细胞的发育分化及其功能有关。Objective To screen the proteins interacting with FOXP3 in yeast two-hybrid system. Methods The ＂bait plasmid＂ pGBKT7 （named as pGBKT7-FOXP3 ） was constructed successfully. Using FOXP3 as bait, a human liver eDNA library was screened and the proteins interacting with FOXP3 were searched. The false positive clones were discarded by one to one yeast two-hybrid system, and the positive elones were sequenced and analyzed by bioinformatic methods. Results The bait plasmid pGBKT7-FOXP3 was constructed successfully and there was no self-activation or toxicity in AH109. Three proteins had been found in our system to be able to interact with FOXP3. They were tumor protein D52, splicing factor 3b subunit 1 and one hypothetical protein. Conclusion FOXP3 interacts with tumor protein D52, splicing factor 3b subunit 1 and one hypothetical protein, all of which may interfere in cell metabolism and function of T cell.

关 键 词：基因 克隆 FOXP3 酵母双杂交 

分 类 号：R341[医药卫生—基础医学] R392-33