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

被引:13
作者
Doddapaneni, Harsha [1 ]
Cregeen, Sara Javornik [2 ,3 ]
Sucgang, Richard [2 ]
Meng, Qingchang [1 ]
Qin, Xiang [1 ]
Avadhanula, Vasanthi [3 ]
Chao, Hsu [1 ]
Menon, Vipin [1 ]
Nicholson, Erin [3 ,4 ]
Henke, David [3 ]
Piedra, Felipe-Andres [3 ]
Rajan, Anubama [3 ]
Momin, Zeineen [1 ]
Kottapalli, Kavya [1 ]
Hoffman, Kristi L. [2 ,3 ]
Sedlazeck, Fritz J. [1 ]
Metcalf, Ginger [1 ]
Piedra, Pedro A. [3 ,4 ]
Muzny, Donna M. [1 ]
Petrosino, Joseph F. [2 ,3 ]
Gibbs, Richard A. [1 ]
机构
[1] Baylor Coll Med, Human Genome Sequencing Ctr, Houston, TX 77030 USA
[2] Baylor Coll Med, Dept Mol Virol & Microbiol, Alkek Ctr Metagen & Microbiome Res, Houston, TX 77030 USA
[3] Baylor Coll Med, Dept Mol Virol & Microbiol, Houston, TX 77030 USA
[4] Baylor Coll Med, Pediat, Houston, TX 77030 USA
来源
PLOS ONE | 2021年 / 16卷 / 08期
关键词
RNA; ALIGNMENT;
D O I
10.1371/journal.pone.0244468
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
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.
引用
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页数:18
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