Analytical comparisons of SARS-COV-2 detection by qRT-PCR and ddPCR with multiple primer/probe sets
Analytical comparisons of SARS-COV-2 detection by qRT-PCR and ddPCR with multiple primer/probe sets
复制标题
qRT-PCR 和 ddPCR 使用多个引物/探针组检测 SARS-COV-2 的分析比较
DOI：
10.1080/22221751.2020.1772679
复制
发表时间：
2020-01-01
影响因子：
13.2
通讯作者：
Lan, Ke
中科院分区：
文献类型：
--
作者：
Liu, Xinjin;Feng, Jiangpeng;Lan, Ke
摘要 世界各地已开发出不同的引物/探针组，用于通过定量实时聚合酶链反应（qRT-PCR）作为标准方法来检测 SARS-CoV-2 的核酸。在我们最近的研究中，我们探讨了使用中国疾病预防控制中心 (CDC) 发布的针对病毒 ORF1ab 或 N 基因的相同引物/探针组的 qRT-PCR 与 qRT-PCR 相比，液滴数字 PCR (ddPCR) 用于临床 SARS-CoV-2 核酸检测的可行性，结果表明 ddPCR 可以在很大程度上减少 qRT-PCR 造成的假阴性报告 [Suo T, Liu X, Feng J, et al. 2017]。 ddPCR：一种更灵敏、更准确的工具，用于低病毒载量样本中的 SARS-CoV-2 检测。 medRxiv [互联网]。 2020;2020.02.29.20029439。可从：https://medrxiv.org/content/early/2020/03/06/2020.02.29.20029439.abstract]。在这里，我们进一步严格比较了具有相同临床样本和条件的 8 个引物/探针组的 qRT-PCR 和 ddPCR 的性能。结果显示，qRT-PCR 中使用的 8 个引物/探针组均无法在低病毒载量（10−4 稀释度）下显着区分真阴性和阳性。此外，还观察到使用 UCDC-N1、N2 和 CCDC-N 引物/探针组进行 qRT-PCR 的假阳性报告。相比之下，与 qRT-PCR 相比，ddPCR 对于低病毒载量样品总体表现出明显更好的性能。值得注意的是，ddPCR 的背景读数相对较低，这可以有效减少假阳性报告的产生。
ABSTRACT Different primers/probes sets have been developed all over the world for the nucleic acid detection of SARS-CoV-2 by quantitative real time polymerase chain reaction (qRT-PCR) as a standard method. In our recent study, we explored the feasibility of droplet digital PCR (ddPCR) for clinical SARS-CoV-2 nucleic acid detection compared with qRT-PCR using the same primer/probe sets issued by Chinese Center for Disease Control and Prevention (CDC) targeting viral ORF1ab or N gene, which showed that ddPCR could largely minimize the false negatives reports resulted by qRT-PCR [Suo T, Liu X, Feng J, et al. ddPCR: a more sensitive and accurate tool for SARS-CoV-2 detection in low viral load specimens. medRxiv [Internet]. 2020;2020.02.29.20029439. Available from: https://medrxiv.org/content/early/2020/03/06/2020.02.29.20029439.abstract]. Here, we further stringently compared the performance of qRT-PCR and ddPCR for 8 primer/probe sets with the same clinical samples and conditions. Results showed that none of 8 primer/probe sets used in qRT-PCR could significantly distinguish true negatives and positives with low viral load (10−4 dilution). Moreover, false positive reports of qRT-PCR with UCDC-N1, N2 and CCDC-N primers/probes sets were observed. In contrast, ddPCR showed significantly better performance in general for low viral load samples compared to qRT-PCR. Remarkably, the background readouts of ddPCR are relatively lower, which could efficiently reduce the production of false positive reports.