Accurate detection and quantification of SARS-CoV-2 genomic and subgenomic mRNAs by ddPCR and meta-transcriptomics analysis

Commun Biol. 2021 Oct 22;4(1):1215. doi: 10.1038/s42003-021-02748-0.

Abstract

SARS-CoV-2 replication requires the synthesis of a set of structural proteins expressed through discontinuous transcription of ten subgenomic mRNAs (sgmRNAs). Here, we have fine-tuned droplet digital PCR (ddPCR) assays to accurately detect and quantify SARS-CoV-2 genomic ORF1ab and sgmRNAs for the nucleocapsid (N) and spike (S) proteins. We analyzed 166 RNA samples from anonymized SARS-CoV-2 positive subjects and we observed a recurrent and characteristic pattern of sgmRNAs expression in relation to the total viral RNA content. Additionally, expression profiles of sgmRNAs, as determined by meta-transcriptomics sequencing of a subset of 110 RNA samples, were highly correlated with those obtained by ddPCR. By providing a comprehensive and dynamic snapshot of the levels of SARS-CoV-2 sgmRNAs in infected individuals, our results may contribute a better understanding of the dynamics of transcription and expression of the genome of SARS-CoV-2 and facilitate the development of more accurate molecular diagnostic tools for the stratification of COVID-19 patients.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • COVID-19 / genetics*
  • COVID-19 / metabolism*
  • COVID-19 Nucleic Acid Testing / methods*
  • Computational Biology
  • Coronavirus Nucleocapsid Proteins*
  • Humans
  • Limit of Detection
  • Open Reading Frames
  • Phosphoproteins
  • Polymerase Chain Reaction / methods*
  • RNA, Messenger / metabolism
  • RNA, Viral / metabolism*
  • Real-Time Polymerase Chain Reaction
  • Reproducibility of Results
  • SARS-CoV-2*
  • Spike Glycoprotein, Coronavirus*
  • Transcriptome*

Substances

  • Coronavirus Nucleocapsid Proteins
  • Phosphoproteins
  • RNA, Messenger
  • RNA, Viral
  • Spike Glycoprotein, Coronavirus
  • nucleocapsid phosphoprotein, SARS-CoV-2
  • spike protein, SARS-CoV-2