摘要
将麻疹病毒 (Nepal株 )的血凝素 (hemagglutinin)基因插入真核表达载体pIRES EGFP ,并在HeLa细胞中表达 .因其较低的表达量 ,所以将其截短 ,去除跨膜区 .使这个截短的HA基因与绿色荧光蛋白基因融合 ,并克隆至原核表达载体pET 2 8b中 .将重组质粒转入大肠杆菌中表达 ,产生了分子量约为 90kD的融合蛋白 .通过ELISA和Western印迹来检测这个基因工程蛋白的抗原性 .在检测一系列的血凝素阳性或阴性的人血清中 ,这个融合蛋白的阳性检出率为 90 % ,阴性检出率为 10 0 % (与市售麻疹病毒诊断试剂盒相比较 ) .由于此HA蛋白是原核表达产物 ,回避了真核表达系统复杂的操作过程和昂贵的费用 ,所以 ,这个麻疹病毒血凝素基因工程抗原有望成为一种新型。
Hemagglutinin(HA)gene of measles virus(MV)(Nepal strain)was cloned into eukaryotic expression vector pIRES\|EGFP,and the recombinant plasmid was transfected into HeLa cells.Beacause of the low expression of ha gene in HeLa cells, ha gene was truncated and cloned into prokaryotic expression vector pET\|28b which contained egfp gene.Then,the recombinant plasmid was transformed into E.coli BL21DE3,producing a fusion antigenic protein H/EGFP with molecular weight of 90 kD.The engineered HA protein was assayed by using ELISA and Western\|blotting for detection the antibodies to measles virus in a cohort of hemagglutinin\|positive of \|negative human sera.The results showed that the MV hemagglutinin gene engineering antigen might provide a more efficient diagnostic reagent for detection of the MV infection
出处
《中国生物化学与分子生物学报》
CAS
CSCD
北大核心
2001年第6期693-698,共6页
Chinese Journal of Biochemistry and Molecular Biology
基金
国家自然科学基金重点项目 (No .39970 0 32 )资助
武汉晨光计划(No .2 0 0 0 5 0 0 40 2 4)资助项目~~
