Oligonucleotide capture sequencing of the SARS-CoV-2 genome and subgenomic fragments from COVID-19 individuals.

TitleOligonucleotide capture sequencing of the SARS-CoV-2 genome and subgenomic fragments from COVID-19 individuals.
Publication TypeJournal Article
Year of Publication2021
AuthorsDoddapaneni, H, Cregeen, SJavornik, Sucgang, R, Meng, Q, Qin, X, Avadhanula, V, Chao, H, Menon, V, Nicholson, E, Henke, D, Piedra, F-A, Rajan, A, Momin, Z, Kottapalli, K, Hoffman, KL, Sedlazeck, FJ, Metcalf, GA, Piedra, PA, Muzny, DM, Petrosino, JF, Gibbs, RA
JournalPLoS One
Date Published2021
KeywordsCOVID-19, DNA, Complementary, Gene Frequency, Genetic Variation, Genome, Viral, Humans, Open Reading Frames, Real-Time Polymerase Chain Reaction, RNA, Viral, SARS-CoV-2, Sequence Analysis, DNA, Viral Load

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.

Alternate JournalPLoS One
PubMed ID34432798
PubMed Central IDPMC8386831
Grant ListU19 AI144297 / AI / NIAID NIH HHS / United States
1U19 AI144297 / NH / NIH HHS / United States

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