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为获得高效的H IV诊断试剂,选定H IV-1的外膜蛋白env中469-511 aa,538-674 aa和700-734 aa 3处包含较多抗原位点的区域作为免疫抗原,用PCR的方法从H IV-1全基因扩增编码这3处片段的基因序列,将它们克隆到原核表达载体中,利用大肠杆菌表达系统表达嵌合蛋白.结果发现,3段嵌合基因能在大肠杆菌BL21(Star)中表达,通过N i-sepharose 4B金属N i螯合层析柱分离纯化目的产物,酶联免疫检测结果表明,纯化抗原有较强的抗原特异性.
To obtain an efficient H IV diagnostic reagent, regions containing more antigenic sites 469-511 aa, 538-674 aa and 700-734 aa 3 of the outer envelope protein env of H IV-1 were selected as immunization antigens, PCR was used to amplify the gene sequences encoding the three fragments from the whole H IV-1 gene, and cloned into the prokaryotic expression vector to express the chimeric protein using the E. coli expression system. The results showed that the three chimeric genes could be expressed in Escherichia coli BL21 (Star), purified by Ni-Sepharose 4B metal N i chelate chromatography. The results of enzyme-linked immunosorbent assay showed that the purified antigen has strong antigen specificity.