人促红细胞生成素受体胞外区基因的克隆及其在大肠肝菌中的表达  

Cloning cDNA of Extracellular Domain of Human Erythropoietin Receptor and its Expression in Escherichia coli

作　　者：张映辉[1] 卢一凡[1] 刘一平[1] 邓继先[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071

出　　处：《生物工程学报》2000年第3期328-332,共5页Chinese Journal of Biotechnology

摘　　要：以人胎肝为材料 ,通过RT PCR的方法扩增出人促红细胞生成素受体 (hEpoR)的胞外区基因。将获得的成熟受体胞外区基因起始密码子改构后克隆到原核表达载体pBV2 2 0中 ,进行原核温控诱导表达。表达产物经蛋白N端测序及Western blot实验证实表达产物是hEpoR胞外区蛋白。利用上罐发酵培养获得的包涵体蛋白经复性纯化后 ,体外生物学活性检测表明表达产物可特异地抑制TF 1细胞在Epo刺激下的生长 ,证实了复性表达产物具有人促红细胞生成素受体胞外区结合Epo的生物活性。Human erythropoietin receptor (hEpoR) plays an important role in regulating the red blood cell production by promoting the proliferation and differentiation of RBC from erythroid precursors. hEpoR is a transmembrane protein,and its extrocellular domain (sEpoR) is of great importance in Epo signal transduction pathway.We cloned the gene of sEpoR by RT PCR from the total RNA of human fetal liver and expressed it in E.coli after insertion of the gene in the expression vector pBV220. The cloned gene was confirmed by sequencing analysis and gene product was confirmed by both Western blot and its first 11 amino acid residues sequence of the N\|terminal. In vitro bioassay showed that the purified gene product can repress the growth of TF cells in the presence of Epo.

关 键 词：胞外医基因 hEpoR 基因克隆 基因表达 大肠杆菌 

分 类 号：Q786[生物学—分子生物学]