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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、便捷的麻疹病毒诊断试剂
The hemagglutinin gene of the measles virus (strain Nepal) was inserted into the eukaryotic expression vector pIRES EGFP and expressed in HeLa cells, which was truncated to remove the transmembrane region due to its low expression level The truncated HA gene was fused with the green fluorescent protein gene and cloned into the prokaryotic expression vector pET28b.The recombinant plasmid was transformed into E.coli to generate a fusion protein with a molecular weight of about 90 kD.According to ELISA and Western blot The antigenicity of this genetically engineered protein was tested in a series of hemagglutinin-positive or -negative human sera with a positive detection rate of 90% and a negative detection rate of 10% (compared with a commercially available measles Virus diagnostic kit.) Since this HA protein is a prokaryotic expression product, which avoids the complex procedure and expense of the eukaryotic expression system, the HA gene engineering antigen is expected to be a new and convenient Measles virus diagnostic reagent