Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data

M.D. Parker; B.B. Lindsey; S. Leary; S. Gaudieri; A. Chopra; M. Wyles; A. Angyal; L.R. Green; P. Parsons; R.M. Tucker; R. Brown; D. Groves; K. Johnson; L. Carrilero; J. Heffer; D.G. Partridge; C. Evans; M. Raza; A.J. Keeley; N. Smith; A.D. Filipe; J.G. Shepherd; C. Davis; S. Bennett; V.B. Sreenu; A. Kohl; E. Aranday-Cortes; L. Tong; J. Nichols; E.C. Thomson; D. Wang; S. Mallal; T.I. de Silva

doi:10.1101/gr.268110.120

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Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data

Journal article

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Peer reviewed

Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data

M.D. Parker, B.B. Lindsey, S. Leary, S. Gaudieri, A. Chopra, M. Wyles, A. Angyal, L.R. Green, P. Parsons, R.M. Tucker, …

Genome Research, Vol.31(4), pp.645-658

2021

DOI: https://doi.org/10.1101/gr.268110.120

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Abstract

We have developed periscope, a tool for the detection and quantification of subgenomic RNA (sgRNA) in SARS-CoV-2 genomic sequence data. The translation of the SARS-CoV-2 RNA genome for most open reading frames (ORFs) occurs via RNA intermediates termed “subgenomic RNAs.” sgRNAs are produced through discontinuous transcription, which relies on homology between transcription regulatory sequences (TRS-B) upstream of the ORF start codons and that of the TRS-L, which is located in the 5′ UTR. TRS-L is immediately preceded by a leader sequence. This leader sequence is therefore found at the 5′ end of all sgRNA. We applied periscope to 1155 SARS-CoV-2 genomes from Sheffield, United Kingdom, and validated our findings using orthogonal data sets and in vitro cell systems. By using a simple local alignment to detect reads that contain the leader sequence, we were able to identify and quantify reads arising from canonical and noncanonical sgRNA. We were able to detect all canonical sgRNAs at the expected abundances, with the exception of ORF10. A number of recurrent noncanonical sgRNAs are detected. We show that the results are reproducible using technical replicates and determine the optimum number of reads for sgRNA analysis. In VeroE6 ACE2+/− cell lines, periscope can detect the changes in the kinetics of sgRNA in orthogonal sequencing data sets. Finally, variants found in genomic RNA are transmitted to sgRNAs with high fidelity in most cases. This tool can be applied to all sequenced COVID-19 samples worldwide to provide comprehensive analysis of SARS-CoV-2 sgRNA.

Details

Title: Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
Authors/Creators: M.D. Parker (Author/Creator) - University of Sheffield
B.B. Lindsey (Author/Creator) - University of Sheffield
S. Leary (Author/Creator) - Murdoch University
S. Gaudieri (Author/Creator) - Murdoch University
A. Chopra (Author/Creator) - Murdoch University
M. Wyles (Author/Creator) - University of Sheffield
A. Angyal (Author/Creator) - Interac
L.R. Green (Author/Creator) - Interac
P. Parsons (Author/Creator) - University of Sheffield
R.M. Tucker (Author/Creator) - University of Sheffield
R. Brown (Author/Creator) - Interac
D. Groves (Author/Creator) - Interac
K. Johnson (Author/Creator) - Sheffield Teaching Hospitals NHS Foundation Trust
L. Carrilero (Author/Creator) - University of Sheffield
J. Heffer (Author/Creator) - University of Sheffield
D.G. Partridge (Author/Creator) - University of Sheffield
C. Evans (Author/Creator) - Sheffield Teaching Hospitals NHS Foundation Trust
M. Raza (Author/Creator) - Sheffield Teaching Hospitals NHS Foundation Trust
A.J. Keeley (Author/Creator) - University of Sheffield
N. Smith (Author/Creator) - Interac
A.D. Filipe (Author/Creator)
J.G. Shepherd (Author/Creator) - University of Glasgow
C. Davis (Author/Creator) - University of Glasgow
S. Bennett (Author/Creator) - University of Glasgow
V.B. Sreenu (Author/Creator) - University of Glasgow
A. Kohl (Author/Creator) - University of Glasgow
E. Aranday-Cortes (Author/Creator) - University of Glasgow
L. Tong (Author/Creator) - University of Glasgow
J. Nichols (Author/Creator) - University of Glasgow
E.C. Thomson (Author/Creator) - University of Glasgow
D. Wang (Author/Creator) - University of Sheffield
S. Mallal (Author/Creator) - Murdoch University
T.I. de Silva (Author/Creator) - University of Sheffield
Publication Details: Genome Research, Vol.31(4), pp.645-658
Publisher: Cold Spring Harbor Laboratory Press.
Identifiers: 991005542607407891
Copyright: © 2021 Parker et al.
Murdoch Affiliation: Institute for Immunology and Infectious Diseases
Language: English
Resource Type: Journal article

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Collaboration types: Domestic collaboration; International collaboration
Citation topics: 1 Clinical & Life Sciences; 1.104 Virology - General; 1.104.1353 Coronavirus Research
Web Of Science research areas: Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology; Genetics & Heredity
ESI research areas: Molecular Biology & Genetics