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

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...

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Autores principales: Parker, Matthew D., Lindsey, Benjamin B., Leary, Shay, Gaudieri, Silvana, Chopra, Abha, Wyles, Matthew, Angyal, Adrienn, Green, Luke R., Parsons, Paul, Tucker, Rachel M., Brown, Rebecca, Groves, Danielle, Johnson, Katie, Carrilero, Laura, Heffer, Joe, Partridge, David G., Evans, Cariad, Raza, Mohammad, Keeley, Alexander J., Smith, Nikki, Filipe, Ana Da Silva, Shepherd, James G., Davis, Chris, Bennett, Sahan, Sreenu, Vattipally B., Kohl, Alain, Aranday-Cortes, Elihu, Tong, Lily, Nichols, Jenna, Thomson, Emma C., Wang, Dennis, Mallal, Simon, de Silva, Thushan I.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Cold Spring Harbor Laboratory Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8015849/
https://www.ncbi.nlm.nih.gov/pubmed/33722935
http://dx.doi.org/10.1101/gr.268110.120
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author Parker, Matthew D.
Lindsey, Benjamin B.
Leary, Shay
Gaudieri, Silvana
Chopra, Abha
Wyles, Matthew
Angyal, Adrienn
Green, Luke R.
Parsons, Paul
Tucker, Rachel M.
Brown, Rebecca
Groves, Danielle
Johnson, Katie
Carrilero, Laura
Heffer, Joe
Partridge, David G.
Evans, Cariad
Raza, Mohammad
Keeley, Alexander J.
Smith, Nikki
Filipe, Ana Da Silva
Shepherd, James G.
Davis, Chris
Bennett, Sahan
Sreenu, Vattipally B.
Kohl, Alain
Aranday-Cortes, Elihu
Tong, Lily
Nichols, Jenna
Thomson, Emma C.
Wang, Dennis
Mallal, Simon
de Silva, Thushan I.
author_facet Parker, Matthew D.
Lindsey, Benjamin B.
Leary, Shay
Gaudieri, Silvana
Chopra, Abha
Wyles, Matthew
Angyal, Adrienn
Green, Luke R.
Parsons, Paul
Tucker, Rachel M.
Brown, Rebecca
Groves, Danielle
Johnson, Katie
Carrilero, Laura
Heffer, Joe
Partridge, David G.
Evans, Cariad
Raza, Mohammad
Keeley, Alexander J.
Smith, Nikki
Filipe, Ana Da Silva
Shepherd, James G.
Davis, Chris
Bennett, Sahan
Sreenu, Vattipally B.
Kohl, Alain
Aranday-Cortes, Elihu
Tong, Lily
Nichols, Jenna
Thomson, Emma C.
Wang, Dennis
Mallal, Simon
de Silva, Thushan I.
author_sort Parker, Matthew D.
collection PubMed
description 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.
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spelling pubmed-80158492021-04-21 Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data Parker, Matthew D. Lindsey, Benjamin B. Leary, Shay Gaudieri, Silvana Chopra, Abha Wyles, Matthew Angyal, Adrienn Green, Luke R. Parsons, Paul Tucker, Rachel M. Brown, Rebecca Groves, Danielle Johnson, Katie Carrilero, Laura Heffer, Joe Partridge, David G. Evans, Cariad Raza, Mohammad Keeley, Alexander J. Smith, Nikki Filipe, Ana Da Silva Shepherd, James G. Davis, Chris Bennett, Sahan Sreenu, Vattipally B. Kohl, Alain Aranday-Cortes, Elihu Tong, Lily Nichols, Jenna Thomson, Emma C. Wang, Dennis Mallal, Simon de Silva, Thushan I. Genome Res Method 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. Cold Spring Harbor Laboratory Press 2021-04 /pmc/articles/PMC8015849/ /pubmed/33722935 http://dx.doi.org/10.1101/gr.268110.120 Text en © 2021 Parker et al.; Published by Cold Spring Harbor Laboratory Press http://creativecommons.org/licenses/by/4.0/ This article, published in Genome Research, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.
spellingShingle Method
Parker, Matthew D.
Lindsey, Benjamin B.
Leary, Shay
Gaudieri, Silvana
Chopra, Abha
Wyles, Matthew
Angyal, Adrienn
Green, Luke R.
Parsons, Paul
Tucker, Rachel M.
Brown, Rebecca
Groves, Danielle
Johnson, Katie
Carrilero, Laura
Heffer, Joe
Partridge, David G.
Evans, Cariad
Raza, Mohammad
Keeley, Alexander J.
Smith, Nikki
Filipe, Ana Da Silva
Shepherd, James G.
Davis, Chris
Bennett, Sahan
Sreenu, Vattipally B.
Kohl, Alain
Aranday-Cortes, Elihu
Tong, Lily
Nichols, Jenna
Thomson, Emma C.
Wang, Dennis
Mallal, Simon
de Silva, Thushan I.
Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title_full Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title_fullStr Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title_full_unstemmed Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title_short Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data
title_sort subgenomic rna identification in sars-cov-2 genomic sequencing data
topic Method
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8015849/
https://www.ncbi.nlm.nih.gov/pubmed/33722935
http://dx.doi.org/10.1101/gr.268110.120
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