Oligonucleotide capture sequencing of the SARS-CoV-2 genome and subgenomic fragments from COVID-19 individuals.
Journal
PloS one
ISSN: 1932-6203
Titre abrégé: PLoS One
Pays: United States
ID NLM: 101285081
Informations de publication
Date de publication:
2021
2021
Historique:
received:
07
12
2020
accepted:
09
08
2021
entrez:
25
8
2021
pubmed:
26
8
2021
medline:
7
9
2021
Statut:
epublish
Résumé
The newly emerged and rapidly spreading SARS-CoV-2 causes coronavirus disease 2019 (COVID-19). To facilitate a deeper understanding of the viral biology we developed a capture sequencing methodology to generate SARS-CoV-2 genomic and transcriptome sequences from infected patients. We utilized an oligonucleotide probe-set representing the full-length genome to obtain both genomic and transcriptome (subgenomic open reading frames [ORFs]) sequences from 45 SARS-CoV-2 clinical samples with varying viral titers. For samples with higher viral loads (cycle threshold value under 33, based on the CDC qPCR assay) complete genomes were generated. Analysis of junction reads revealed regions of differential transcriptional activity among samples. Mixed allelic frequencies along the 20kb ORF1ab gene in one sample, suggested the presence of a defective viral RNA species subpopulation maintained in mixture with functional RNA in one sample. The associated workflow is straightforward, and hybridization-based capture offers an effective and scalable approach for sequencing SARS-CoV-2 from patient samples.
Identifiants
pubmed: 34432798
doi: 10.1371/journal.pone.0244468
pii: PONE-D-20-38498
pmc: PMC8386831
doi:
Substances chimiques
DNA, Complementary
0
RNA, Viral
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
e0244468Subventions
Organisme : NIAID NIH HHS
ID : U19 AI144297
Pays : United States
Organisme : NIH HHS
ID : 1U19 AI144297
Pays : United States
Commentaires et corrections
Type : UpdateOf
Type : UpdateOf
Déclaration de conflit d'intérêts
The authors have declared that no competing interests exist.
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