蜂毒素-SP94杂合基因原核表达载体的构建及蛋白纯化  

Construction of Prokaryotic Expression Vector of Melittin-SP94 and Purification

作　　者：张红绪[1] 曹静[2] 周庆峰[2] 赵鹏[1] 金白洁[1] 王亚威[1] 刘琦[1] 李成伟[2] 

机构地区：[1]河南师范大学生命科学学院,河南新乡453007 [2]商丘师范学院生命科学系,河南商丘476000

出　　处：《河南师范大学学报（自然科学版）》2011年第3期122-125,共4页Journal of Henan Normal University(Natural Science Edition)

基　　金：国家自然科学基金(30901877);河南省教育厅自然科学研究项目(2009B180021);河南省动物学重点学科资助

摘　　要：用可以特异结合到肝癌细胞膜上的小肽SP94,与蜂毒素连接,构建重组表达载体pET32a-蜂毒素-SP94.转入大肠杆菌BL21(DE3),IPTG诱导表达,SDS-PAGE检测表达结果,产物经Ni-NTA Agrosc亲和层析柱纯化后进行Tricine-SDS-PAGE检测.重组蛋白在大肠杆菌中高效表达,约占细胞总蛋白的30%,超声破碎后50%的目的蛋白以可溶性形式存在,纯化后获得高纯度蛋白.本实验为肝癌的靶向治疗研究与临床应用提供了新思路.A small gene SP94 that could specific ally bind to the membrane of hepatoma cell is connected to Melittin,obtaining a fusion gene melittin-SP94.pET32a is chosen as the recombinant vector and pET32a-Melittin-SP94 is constructed,the recombinant protein Trx-Melittin-SP94 is expressed into E.coli BL21(DE3).The recombinant protein is identified by Tricine-SDS-PAGE and purifed by Ni2+ affinity chromatography.The resulting expression of fusion protein Trx-Melittin-SP94 could reach up to 30% of the total cell proteins.More than 50% of the target protein is in a soluble form.High quatified purity protein is obtained by purification.The study has provided a new path of targeted therapy of liver cancer research and clinical applications.

关 键 词：肝癌 蜂毒素-SP94 载体构建 表达 纯化 

分 类 号：N55[自然科学总论] G658.3[文化科学—教育学]