Rnadvance viral rna extraction kit

All samples selected for sequencing had nucleic acid freshly extracted from the primary sample source independently of the material extracted for the initial RT-PCR testing. RNA isolation was performed at 4 workplaces—the Public Health Authority of the Slovak Republic in Bratislava (PHA SR), the Regional Public Health Authority in Banská Bystrica (RPHA BB), the Regional Public Health Authority in Trenčín (RPHA TN) and the Regional Public Health Authority in Košice (RPHA KE). Samples for the Comenius University Science Park in Bratislava (CU SP) and the Jessenius Faculty of Medicine of Comenius University (JFM CU) in Martin sequencing centers were isolated at PHA SR in Bratislava. The workplaces extracted viral RNA from nasopharyngeal swab samples using RNAdvance Viral RNA Extraction Kit (Beckman Coulter, CA, USA), QIAamp Viral RNA Mini Kit (QIAGEN GmbH, Hilden, DE, USA) and Quick-RNA™ Viral Kit (Zymo Research Corp., Irvine, CA, USA). They followed the manufacturer’s protocol with the following small exception to the Quick-RNA™ Viral Kit: for the first step, the addition of 100 μL of DNA/RNA Shield™ was skipped. RNA isolation was performed on a plate and then stored in a 96-well plate at −80 °C. Subsequently, the plates with isolated RNA were processed by the appropriate sequencing center.

Genomic RNA from SARS-CoV-2 USA WA1/2020 (wild type, WT), viral culture stocks of SARS-CoV-2 hCoV-19/England/204820464/2020 (B.1.1.7 variant, 501-MT) and SARS-CoV-2 Isolate hCoV-19/South Africa/KRISP-K005325/2020 (B.1.351 variant, 484-MT) were obtained from BEI Resources, NIAID (Manassas, VA). RNA was isolated from both the variant strains in a BSL3 laboratory, using RNAdvance viral RNA extraction kit (Beckman Coulter, Indianapolis, IN).