Mollaei Hamid Reza, Afshar Abass Aghaei, Kalantar-Neyestanaki Davood, Fazlalipour Mehdi, Aflatoonian Behnaz
Tropical and Infectious Diseases Research Center, Kerman University of Medical Sciences, Kerman, Iran.
Medical Mycology and Bacteriology Research Center, Kerman University of Medical Sciences, Kerman, Iran.
Iran J Microbiol. 2020 Jun;12(3):185-193.
The new beta-coronavirus, which caused Severe Acute Respiratory Coronavirus-2 Syndrome (SARS-CoV-2), a major respiratory outbreak in Wuhan, China in December 2019, is now prevalent in many countries around the world. Identifying PCR-based viruses is a well-known and relatively stable protocol. Unfortunately, the high mutation rates may lead to widespread changes in viral nucleic acid sequences, and so using specific primers for PCR can be recommended. In this study, we evaluated the power of a conventional RT-PCR to detect SARS-CoV-2 RNA among the five set primer sets.
The five genomic regions of the Coronavirus SARS-2 virus including Nucleocapsids (N), Envelope (E), RNA depended RNA Polymerase (RdRp), ORF1ab and Spike (S) were selected for primer designing. A conventional RT-PCR was performed to compare sensitivity, specificity and other analytical characteristics of primers designed against two Real Time PCR commercial kits.
The result of the comparative analysis showed that the ORF1ab, N and RdRp primers had a sensitivity, specificity and positive predictive value higher than other primers. A significant difference in the analytical sensitivity between the studied primer sets in RT-PCR kits was observed.
In this study, the ORF1ab, Nucleocapsid and RdRp regions have the best primers for identifying the SARS-CoV-2 RNA between different genes that have been suggested.
新型β冠状病毒引发了严重急性呼吸综合征冠状病毒2（SARS-CoV-2），于2019年12月在中国武汉引发了一次重大呼吸道疫情，目前在世界许多国家广泛传播。识别基于聚合酶链反应（PCR）的病毒是一种广为人知且相对稳定的方案。不幸的是，高突变率可能导致病毒核酸序列发生广泛变化，因此推荐使用针对PCR的特异性引物。在本研究中，我们评估了传统逆转录PCR（RT-PCR）在五组引物中检测SARS-CoV-2核糖核酸（RNA）的能力。
选择严重急性呼吸综合征冠状病毒2（SARS-2）病毒的五个基因组区域，包括核衣壳（N）、包膜（E）、RNA依赖的RNA聚合酶（RdRp）、开放阅读框1ab（ORF1ab）和刺突蛋白（S）用于引物设计。进行传统RT-PCR以比较针对两种实时荧光定量PCR商业试剂盒设计的引物的敏感性、特异性和其他分析特性。
对比分析结果表明，ORF1ab、N和RdRp引物的敏感性、特异性和阳性预测值高于其他引物。观察到RT-PCR试剂盒中所研究引物组之间的分析敏感性存在显著差异。
在本研究中，ORF1ab、核衣壳和RdRp区域在已提出的不同基因中具有用于识别SARS-CoV-2 RNA的最佳引物。
