Amplicon seq of SARS-CoV-2: nasopharyngeal swab
Aliquots of surplus RNA were provided for sequencing analysis. Briefly, library preparation consisted of converting RNA to cDNA using LunaScript (Thermofisher), then amplified by reverse complement (RC)-PCR amplification (EasySeq SARS-CoV-2 Whole Genome Sequencing kit, Nimagen, Netherlands). This kit barcodes and ligates Illumina adapters in a single PCR reaction, with two separate pools of primers (pools 1 and 2). After amplification, primer pools 1 and 2 for each amplified sample were mixed 1:1 before being cleaned with AmpliCleanTM beads and quantified. All cleaned samples were then pooled together and denatured. Finally, the denatured amplicon library was loaded into the NovaSeq cartridge (2  150 bp run). The sequencing was conducted in two sequencing runs, one for the first 120 patients swab samples, and a second for the final 60 patients swab samples.