全长人雌激素受体cDNA的克隆及酵母表达载体的构建

Cloning of Full-length Human Estrogen Receptor and Construction of the Expression Vector in Yeast

目的 建立环境雌激素基因重组酵母测评系统 ,将全长人雌激素受体 c DNA在酵母中克隆并表达。方法 用Eco RI和 Sac I从 p SG5 /HEG0质粒上切下 h ER c DNA,定向插入 p U C18克隆载体中 ,构建成重组质粒 p UC- h ER;再用Eco RI和 Bam HI切下 h ER c DNA,将其连接到 p GAD42 4酵母表达载体上。结果 构建成功重组酵母表达载体 p GAD-h ER,序列分析表明插入处接头和读框完全正确。结论 本文构建成功全长人雌激素受体 c DNA的酵母表达载体p GAD- h ER。

Objective To establish the recombinant yeast estrogen system(RYES),cloning and expression of full length human estrogen receptor(hER) cDNA in yeast Methods hER cDNA was cut down from pSG5/HEG0 with EcoRI/SacI,and then was inserted into the cloning plasmid pUC18 From the recombinant cloning plasmid pUC hER obtained,hER cDNA was cut down again with EcoRI/BamHI,and was inserted into the yeast expression vector pGAD424 Results The recombinant yeast expression vector pGAD hER was constructed successfully Sequence analysis indicated that sequence and ORF of the joint where hER cDNA was inserted into was correct exactly Conclusion The recombinant yeast expression vector of full length human estrogen receptor cDNA,pGAD hER,was constructed successfully,which was the very important groundwork for further establishment of the recombinant yeast estrogen system(RYES)