简套式荧光RT-PCR检测患者血清样本中SARS冠状病毒

目的　建立一种快速、特异、灵敏的SARS冠状病毒(SARS CoV)核酸检测方法。方法　根据GenBank中SARS CoV基因序列,自行设计引物、荧光探针,在PE 770 0扩增仪上探讨工作参数,形成试剂盒,并用研制的试剂检测76份临床SARS样本。结果　简套式荧光RT PCR方法对血清样本、漱口液样本、正常人样本检出率分别为33.3% (12 36 )、6 7.5 % (2 7 4 0 )、0 (0 / 16 0 ) ,与经典套式检测结果一致。而传统一步法荧光RT PCR对样本的检出率分别为13 9% (5 36 )、5 2 5 % (2 1 4 0 )、0 (0 / 80 )。结论　简套式荧光RT PCR核酸检测法是SARS临床早期诊断快速、特异、灵敏的方法。

Determination of SARS-CoV by simplified nested fluorescent RT-PCR

Objective To establish a simple rapid and sensitive nested RT-PCR method for detection of SARS coronavirus RNA by designing the specific primers for SARS and optimizing the parameters for PCR. Methods Primers and fluorescent probes were designed according to the sequences of SARS coronavirus genes available from GenBank. The optimization of the parameters for PCR was performed in PE 7700 thermal cycler. The 36 serum samples and 40 mouthwash of SARS patients and 80 samples of healthy people were tested. Results The positive rate of patient serum and mouthwash was 33.6%(12/36)and 67.5%(27/40), respectively, while the positive rate of healthy people was zero (0/160). Conclusion The simple nested RT-PCR method was a rapid, efficient and sensitive one for SARS early diagnosis.