High-coverage SARS-CoV-2 genome sequences acquired by target capture sequencing

J Med Virol. 2020 Oct;92(10):2221-2226. doi: 10.1002/jmv.26116. Epub 2020 Jun 19.

Abstract

In this study, we designed a set of SARS-CoV-2 enrichment probes to increase the capacity for sequence-based virus detection and obtain the comprehensive genome sequence at the same time. This universal SARS-CoV-2 enrichment probe set contains 502 120 nt single-stranded DNA biotin-labeled probes designed based on all available SARS-CoV-2 viral sequences and it can be used to enrich for SARS-CoV-2 sequences without prior knowledge of type or subtype. Following the CDC health and safety guidelines, marked enrichment was demonstrated in a virus strain sample from cell culture, three nasopharyngeal swab samples (cycle threshold [Ct ] values: 32.36, 36.72, and 38.44) from patients diagnosed with COVID-19 (positive control) and four throat swab samples from patients without COVID-19 (negative controls), respectively. Moreover, based on these high-quality sequences, we discuss the heterozygosity and viral expression during coronavirus replication and its phylogenetic relationship with other selected high-quality samples from the Genome Variation Map. Therefore, this universal SARS-CoV-2 enrichment probe system can capture and enrich SARS-CoV-2 viral sequences selectively and effectively in different samples, especially clinical swab samples with a relatively low concentration of viral particles.

Keywords: SARS coronavirus; gene expression; genetic networks; genetic variability; mutation.

MeSH terms

  • Biotin / chemistry
  • COVID-19 / diagnosis*
  • COVID-19 / pathology
  • COVID-19 / virology
  • DNA Probes / chemical synthesis
  • DNA Probes / metabolism*
  • DNA, Single-Stranded / genetics*
  • DNA, Single-Stranded / metabolism
  • Genome, Viral*
  • Genotype
  • Humans
  • Mutation
  • Nasopharynx / virology
  • Phylogeny
  • Reverse Transcriptase Polymerase Chain Reaction / standards
  • SARS-CoV-2 / classification
  • SARS-CoV-2 / genetics*
  • SARS-CoV-2 / isolation & purification
  • SARS-CoV-2 / pathogenicity
  • Sensitivity and Specificity
  • Whole Genome Sequencing / methods*

Substances

  • DNA Probes
  • DNA, Single-Stranded
  • Biotin