SARS-CoV-2 Detection by qRT-PCR

Severe Acute Respiratory Syndrome Coronavirus 2 genomic RNA was detected by qRT-PCR using SARS-CoV-2-specific primers and probes for the E, N, and RdRP genes (31 ). In brief, total RNA was extracted from 140 μL swab-UTM using the QIAamp Virus RNA mini kit (QIAGEN, Germany) following the manufacturer’s instructions. One-step qRT-PCR was performed in a 20-μL mixture containing 5 μL of extracted RNA with an Mx3000P PCR System (Agilent, United States) and a LightCycler Multiplex RNA Virus Master kit (Roche Diagnostics, Germany). A cycle threshold (Ct) value <40 indicates a positive result (32 (link)). Negative (RNAse-free water) and positive (RNA extracted from hCoV-19/Taiwan/4/2020, EPI_ISL_411927 virus culture fluid) controls were included. The primers, probe, mixture, machine, and thermal cycling conditions are listed in Supplementary Tables 1A–C.