Viral RNA was isolated from cell-cultured SARS-CoV-2, SARS-1, MERS-CoV, hCoV-NL63, hCoV-OC43, hCoV-229E, and from nasopharyngeal swabs from n=10 patients by MagNA Pure LC (Roche Diagnostics, The Netherlands) using the total nucleic acid isolation kit. The RNA was converted into cDNA using SuperscriptIII (Thermo-Fisher Scientific, USA) and random hexamers. Subsequently, conventional PCR was performed on the cDNA using HotStar Taq DNA polymerase (Qiagen, The Netherlands) with 400 nM forward primer (5′-AG CAC TCT CCA AGG GTG TTC-3′) and 400 nM reverse primer (5′-GCA AAG CCA AAG CCT CAT TA-3′) and the following cycling conditions: 15 min at 95C, followed by 40 cycles of 1 min at 95C, 1 min at 5 C and 1 min at 72C. The PCR products were visualized by electrophoresis. The same RNA was used in a diagnostics reference assay by Corman et al.5 and the Cycle threshold values form this reference assay were used for estimating sensitivity.

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