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目的 构建含输血传播病毒 (TTV)部分基因序列的原核表达载体并进行表达及产物活性的鉴定。方法 在计算机模拟基础上 ,采用PCR的方法 ,从一名非甲～庚肝炎患者血清中 ,扩增编码蛋白具有抗原活性的TTVORF2部分基因片段。将该片段插入表达载体pQE - 30 ,在大肠埃希菌中进行表达并利用酶联免疫吸附试验 (ELISA)鉴定表达产物活性。结果 SDS -PAGE电泳后经电脑分析显示 ,表达产物相对分子质量与预计相同 ,表达量占菌体总蛋白量的 2 2 33%。经ELISA检测表明表达产物有结合抗体活性。结论 成功构建了含TTV部分基因的原核表达载体 ,表达产物具有抗原活性
Objective To construct a prokaryotic expression vector containing part of the gene of blood transfusion transmitted virus (TTV) and to express it and identify its activity. Methods Based on the computer simulation, a partial TTVORF2 gene fragment encoding the antigenic activity of the protein was amplified from a non-A ~ G hepatitis patient by PCR. The fragment was inserted into expression vector pQE - 30, expressed in Escherichia coli and the activity of expressed product was identified by enzyme linked immunosorbent assay (ELISA). Results After SDS-PAGE electrophoresis, the relative molecular mass of the expressed product was expected to be the same as that predicted by computer analysis, accounting for 22.33% of the total bacterial protein content. The result of ELISA showed that the expressed product had the bound antibody activity. Conclusion The prokaryotic expression vector containing part of TTV gene has been successfully constructed and the expressed product has antigenic activity