SARS冠状病毒多聚酶区抗原在感染诊断中的作用探讨

Prokaryotic expression and application of RNA polymerase of SARS coronavirus

目的原核表达严重急性呼吸综合征冠状病毒(SARS-CoV)非结构蛋白RNA多聚酶表位区,了解其在SARS-CoV感染诊断及作为病毒复制指标方面的意义。方法通过PCR扩增获得RNA多聚酶表位区基因,与原核表达载体pET32a+连接,转化大肠杆菌BL-21表达获得重组融合蛋白。经切胶透析纯化后,应用ELISA检测SARS患者及动物模型血清标本的IgG抗体。结果获得原核表达的RNA多聚酶表位区。ELISA检测正常人血清均阴性,SARS患者血清阳性率为95%;检测SARS病毒感染实验动物血清阳性率100%,灭活纯化病毒接种恒河猴均为阴性。结论RNA多聚酶表位区具有很好的免疫原性,可用于感染诊断的检测及作为病毒复制的指标。

Objective To study the antigenicity of RNA polymerase and the relationship between the protein and the SARS-CoV replication. Methods The fragment was amplified by PCR, ligated with the prokaryotic expression vector pET32a＋ and transformed into E. coli BL-21 . The expressed fusion protein identified by Western-blot and ELISA was used to detect the anti-SARS CoV IgG in different sera. Results The fusion protein was expressed successfully in E. coli BL-21. Detected by ELISA, the positive percentage of the anti-SARS IgG in the healthy donors, patients, infected animals and the rhesus administrated with the inactivated-virus were 0%, 95%, 100% and 0%, respectively. Conclusion Good antigenicity was shown in the expressed RNA polymerase. It can be used to diagnose the infection and to demonstrate the replication of SARS-CoV virus.