HER2胞外区基因的克隆及其在大肠杆菌中的可溶性表达  

Soluble Expression of Extracellular Region of HER2 in Escherichia coli

作　　者：刘阳[1,2] 孙志伟[1] 俞炜源[1] 黄海华[3] 

机构地区：[1]军事医学科学院生物工程研究所 [2]沈阳药科大学生物制药系,辽宁沈阳110016 [3]沈阳药科大学生物制药系

出　　处：《生物技术通讯》2005年第3期252-254,共3页Letters in Biotechnology

摘　　要：采用反转录PCR和PCR方法分别克隆P185HER2n/eu胞外区基因和噬菌体M13KO7g3p-N1结构域基因,然后将二者偶联入pET-22b(+)载体中,在大肠杆菌中进行融合表达。可溶性目的蛋白表达量占细菌可溶性表达产物总量的30%左右,并通过镍亲和层析纯化出目的蛋白。以上结果为从噬菌体抗体库中筛选抗P185HER2/neu的抗体奠定了基础。The extracellular region gene of human P185HER2/neu and the gene of g3p-N1 domain were amplified by RT-PCR and PCR from sk-br-3 cell and M13KO7 phage respectively. Fused to 3'-terminus of the gene of g3p-N1 domain, the extracellular region gene of human P185HER2/neu was high-efficiently expressed in host E.coli by vector pET-22b(+). The fusion protein attaining to the 30% of the total protein which could be soluble expressed in E.coli was conveniently purified by chelating column (HisTrap HP Columns). This research made the basis of screening anti-P185HER2/neu antibody from recombinant phage-display antibody library.

关 键 词：大肠杆菌 胞外区 可溶性表达 基因 克隆 反转录PCR 噬菌体抗体库 目的蛋白 PCR方法 融合表达 层析纯化 表达产物 结构域 表达量 载体 细菌 

分 类 号：Q78[生物学—分子生物学] Q786