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利用ＰＣＲ技术和ＤＮＡ体外重组方法，将内皮素Ａ型受体（ＥｎｄｏｔｈｅｌｉｎｔｙｐｅＡｒｅｃｅｐｔｏｒ，ＥＴＡＲ）胞内区ｃＤＮＡ克隆到ｐＵＣ１８载体中进行序列分析。结果表明，ＤＮＡ序列与预期结果完全相符。然后用低熔点琼脂糖回收插入到ｐＵＣ１８载体中的ＥＴＡＲ胞内区ＤＮＡ片段，连接到ｐＧＥＸ２Ｔ融合蛋白表达载体的凝血酶位点下游，转化大肠杆菌ＪＭ１０３，得到的重组质粒命名为２Ｔ／ＥＴＡＲ。ＪＭ１０３（２Ｔ／ＥＴＡＲ）经３０℃培养、ＩＰＴＧ诱导明显表达出ＥＴＡＲ胞内区融合蛋白，表达产物主要以包涵体形式存在。包涵体经变性和复性后，用亲和层析一步纯化法获得了较纯的ＥＴＡＲ胞内区融合蛋白，为进一步研究ＥＴＡＲ胞内区的功能打下了良好的基础
The intracellular region of Endothelin type receptor (ETAR) was cloned into pUC18 vector and sequenced by PCR and DNA in vitro recombination. The results show that the DNA sequence exactly matches the expected result. Then the low melting point agarose was used to recover the intracellular region of ETAR inserted into the pUC18 vector and ligated into the downstream of the thrombin site of pGEX-2T fusion protein expression vector to transform into E. coli JM103. The resulting recombinant plasmid was named as 2T / ETAR . JM103 (2T / ETAR) cultured at 30 ℃, IPTG induced significant expression of ETAR intracellular fusion protein, the expression product mainly in the form of inclusion bodies. After inclusion body denatured and refolded, the pure ETAR intracellular fusion protein was obtained by affinity chromatography one-step purification method, which laid a good foundation for further study on the function of ETAR intracellular region