Key Laboratory for Molecular Genetic Mechanisms and Intervention Research on High Altitude Disease of Tibet Autonomous Region, School of Medicine, Xizang Minzu University, Xianyang, China.
Department of Basic Medical Sciences, Taizhou University, Taizhou, China.
J Clin Lab Anal. 2022 Jan;36(1):e24137. doi: 10.1002/jcla.24137. Epub 2021 Dec 3.
Coronavirus disease 2019 (COVID-19) has had a devastating impact on public health services worldwide. Currently, there are no standard remedies or therapies for COVID-19. it is important to identify and diagnose COVID-19 to control the spread. But clinical symptoms of COVID-19 are very similar to those of other respiratory viruses.
As a result, the diagnosis of COVID-19 relies heavily on detecting pathogens. We established a bunch of triplex new TaqMan real-time PCR assays. Three sets of primers and probes (targeting the ORF1ab, N, and E genes, respectively) are poorly consistent with other human coronaviruses and the human influenza virus. The sensitivity of established PCR assays notices as few as 100 copies per PCR of the ORF1ab, N, and E genes. Meanwhile, standard curves concluded from constant PCR reaction all showed glorious linear correlations between Ct values and the polymer loading copy variety (correlation coefficient (R ) of ORF1ab, N, and E genes is 0.996, 0.991, and 0.998, respectively). Surveillance of RNA-based pseudovirus demonstrated that they were identified to be positive with respect to SARS-CoV-2 and that established PCR assays are achievable.
The assays established provide a smaller reaction volume for diagnosing COVID-19.
2019 年冠状病毒病（COVID-19）对全球公共卫生服务造成了毁灭性影响。目前，尚无针对 COVID-19 的标准治疗方法或药物。识别和诊断 COVID-19 对于控制其传播非常重要。但是，COVID-19 的临床症状与其他呼吸道病毒非常相似。
因此，COVID-19 的诊断主要依赖于检测病原体。我们建立了一系列三重 TaqMan 实时 PCR 检测方法。三套引物和探针（分别针对 ORF1ab、N 和 E 基因）与其他人类冠状病毒和人流感病毒的一致性较差。建立的 PCR 检测方法的灵敏度可低至每个 PCR 反应中的 ORF1ab、N 和 E 基因的 100 个拷贝。同时，从恒定 PCR 反应得出的标准曲线均显示 Ct 值与聚合酶加载拷贝数之间具有极好的线性相关性（ORF1ab、N 和 E 基因的相关系数（R）分别为 0.996、0.991 和 0.998）。基于 RNA 的假病毒监测表明，它们可被鉴定为 SARS-CoV-2 阳性，并且建立的 PCR 检测方法是可行的。
建立的检测方法为诊断 COVID-19 提供了更小的反应体积。
