人MASP1和MASP2 C端片段的原核表达  

Prokaryotic expression of the C terminal fragment of mannan-binding lectin-associated serine protease-1 and-2

作　　者：蔡学敏[1] 赵娜[1] 张丽芸[1] 左大明[1] 陈政良[1] 

机构地区：[1]南方医科大学免疫学教研室

出　　处：《免疫学杂志》2009年第5期577-580,共4页Immunological Journal

基　　金：国家自然科学基金资助项目(30371310);广东省自然科学基金研究团队项目(015003)

摘　　要：目的在大肠杆菌中表达人甘露聚糖结合凝集素(MBL)相关丝氨酸蛋白酶1、2(MASP1、MASP2)C端片段。方法采用PCR技术从分别含人MASP1、MASP2cDNA的质粒pGEM-MASP1、pGEM-MASP2中扩增MASP1、MASP2C端基因片段,将其插入原核表达载体pET17b,转化感受态大肠杆菌BL21(DE3),诱导表达MASP1、MASP2C端蛋白,通过Ni2+-NTA亲和层析柱纯化目的蛋白,以SDS-PAGE、Western blot和ELISA进行鉴定。结果分别从pGEM-MASP1和pGEM-MASP2中扩增到约1300bp的基因片段,构建成重组表达载体,经酶切出现约3300bp和1300bp片段,测序结果与预期的完全一致。纯化蛋白经SDS-PAGE可见Mr40000蛋白带,该蛋白可与抗His抗体反应。结论获得了重组人MASP1和MASP2C端片段蛋白及其表达菌株,为MASP分子的进一步研究提供了条件。Objective To express the C-terminal fragment of human mannan-binding lectin(MBL)associated serine proteases-1 and 2(MASP1,2)in E.coli.Methods The target sequence in pGEM-MASP1 and pGEM-MASP2 plasmid containing human MASP1 and MASP2 cDNA were amplified by PCR,and then inserted into prokaryotic expression vector pET17b.After identification by restriction mapping and sequencing,the recombinant expression vectors were transformed into E.coli BL21(DE3)cells.The expressed products were purified by Ni2+-NTA Immobi...

关 键 词：甘露聚糖结合凝集素相关丝氨酸蛋白酶1、2 C端片段 原核表达 

分 类 号：R392.1[医药卫生—免疫学]