【摘要】 目的在大肠杆菌中表达重组1-4型登革病毒外膜蛋白,为研制登革病毒抗体快速检测试剂提供原材料。方法根据基因序列设计、合成引物并经PCR方法扩增1-4型登革病毒部分外膜蛋白基因片段,克隆至表达载体pET30a(+)并在大肠杆菌中诱导表达。小量表达以及纯化后的重组蛋白做SDS-PAGE分析并经WB实验验证其免疫活性。结果酶切消化及电泳结果表明4种重组质粒均构建成功。经IPTG诱导后,4种重组大肠杆菌均可表达相应的重组蛋白。纯化前后的重组蛋白经WB实验表明,4种重组蛋白均能与对应型别的病毒感染者血清反应。结论利用大肠杆菌表达系统,成功表达重组1-4型登革病毒外膜蛋白,4种型别的重组蛋白可被各自血清型的抗体结合,均具有免疫反应活性。更多还原

【Abstract】 In this study,we expressed the recombinant envelope(env) proteins of dengue virus serotype 1-4 in E.coli,which expected to be as the raw material for developing rapid dengue viral antibody detection kit.According to the sequences of viral env genes,primer pairs were designed and synthesized.By using PCR method,the truncated env fragments of four serotypes were amplified and inserted into the pET30a(+) vector.The recombinant plasmids were confirmed by restriction reaction and induced with IPTG in E.coli,to express the recombinant proteins in small scale.The two forms of recombinant env proteins of four serotypes,in bacterial lysates or purified,were analyzed by SDS-PAGE and the immunological reactivities were tested by western blotting.Each type of recombinant protein,which could be recognized and bound by the antibodies of distinct serotype,possessed the immunological reactivity.更多还原