Quantification of viral RNA

SARS-CoV-2 RNA from cell culture supernatant samples was isolated using AVL buffer and the QIAamp Viral RNA Kit (QIAGEN) according to the manufacturer’s instructions. Absorbance-based quantification of the RNA yield was performed using the Genesys 10S UV-Vis Spectrophotometer (Thermo Scientific). RNA was subjected to OneStep qRT-PCR analysis using the Luna Universal One-Step RT-qPCR Kit (New England Biolabs) and a CFX96 Real-Time System, C1000 Touch Thermal Cycler. Primers were adapted from the WHO protocol (^{Corman et al., 2020}) targeting the open reading frame for RNA-dependent RNA polymerase (RdRp): RdRP_SARSr-F2 (GTG ARA TGG TCA TGT GTG GCG G) and RdRP_SARSr-R1 (CAR ATG TTA AAS ACA CTA TTA GCA TA) using 0.4 μM per reaction. Standard curves were created using plasmid DNA (pEX-A128-RdRP) harboring the corresponding amplicon regions for RdRP target sequence according to GenBank Accession number {"type":"entrez-nucleotide","attrs":{"text":"NC_045512","term_id":"1798174254","term_text":"NC_045512"}}NC_045512. All quantification experiments have been carried out with biological replicates.