摘要
利用PCR方法从幽门螺旋杆菌(Helicobacter pylori,HP)基因组DNA中获得α1,2-岩藻糖基转移酶(α1,2-fuco-syltransferase,α1,2-fuct)基因,得到大小为906 bp的目的基因,将其定向插入到原核表达载体pET-22b(+)中,得到重组表达载体pET-fuct。将重组表达载体转化到大肠杆菌BL21(DE3)中,25℃,0.1 mmol/L异丙基硫代半乳糖苷(IPTG)诱导表达4 h,并用SDS-PAGE分析目的蛋白的表达情况。结果表明,可表达出相对分子质量为33 kD的蛋白,与预期分子量一致,说明α1,2-岩藻糖基转移酶在大肠杆菌BL21中实现表达,应用HPLC法进行酶活检验,酶活达到了13.21 pmol/(mgPr.h)。
A prokaryotic expression vector pET-fuct containing the gene of alpha 1,2-fucosyltransferase was constructed and expressed in E.coli BL21（DE3）.The fuct gene was amplified from the extracted total DNA of Helicobacter pylori by PCR,and was inserted into the prokaryotic expression vector pET-22b（＋） to construct a recombinant expression vector pET-fuct.The expression of fuct gene was induced by 0.1 mmol/L IPTG at 25℃ for 4 hours,and SDS-PAGE analysis showed that protein with molecular weight of 33 kD was expressed in E.coli BL21.This protein had a specific activity of 13.21 pmol/（mgPr·h） which was detected by HPLC.
出处
《生物技术通报》
CAS
CSCD
北大核心
2011年第3期185-190,共6页
Biotechnology Bulletin
基金
国家自然科学基金青年科学基金项目(20806060)
天津市教委项目(20080601)
天津科技大学引进人才科研启动基金项目(20080435)
作者简介
贾红红，女，硕士，研究方向：微生物与生化药学；E-mail：jiahonghong850@163．com
通讯作者：李玉，女，博士，副教授，E-mail：liyu@tust．edu．cn
