摘要：SummarySevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emerging new type of coronavirus that is responsible for the COVID-19 pandemic and the unprecedented global health emergency. Whole-genome sequencing (WGS) of SARS-CoV-2 plays a critical role in understanding the disease. Performance variation exists across SARS-CoV-2 viral WGS technologies, but there is currently no benchmarking study comparing different WGS sequencing protocols. We compared seven different SARS-CoV-2 WGS library protocols using RNA from patient nasopharyngeal swab samples under two storage conditions with low and high viral inputs. We found large differences in mappability and genome coverage, and variations in sensitivity, reproducibility, and precision of single-nucleotide variant calling across different protocols. For certain amplicon-based protocols, an appropriate primer trimming step is critical for accurate single-nucleotide variant calling. We ranked the performance of protocols based on six different metrics. Our findings offer guidance in choosing appropriate WGS protocols to characterize SARS-CoV-2 and its evolution.Graphical abstractDisplay OmittedHighlights•Benchmarking comparison on seven different SARS-CoV-2 genome sequencing protocols•Ranked protocol performances based on genome coverage and SNV detection•Raised concern of primer-masking on SNV calling from Qiagen Amplicon protocol•Proposed bioinformatics remedy on making correct SNV call from masked genome regionsBiological sciences; Molecular biology; Microbiology; Virology; Omics
