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High-accuracy long-read amplicon sequences using unique molecular identifiers with Nanopore or PacBio sequencing.
Karst, Søren M; Ziels, Ryan M; Kirkegaard, Rasmus H; Sørensen, Emil A; McDonald, Daniel; Zhu, Qiyun; Knight, Rob; Albertsen, Mads.
Afiliación
  • Karst SM; Center for Microbial Communities, Department of Chemistry and Bioscience, Aalborg University, Aalborg, Denmark.
  • Ziels RM; Department of Civil Engineering, The University of British Columbia, Vancouver, British Columbia, Canada.
  • Kirkegaard RH; Center for Microbial Communities, Department of Chemistry and Bioscience, Aalborg University, Aalborg, Denmark.
  • Sørensen EA; Center for Microbial Communities, Department of Chemistry and Bioscience, Aalborg University, Aalborg, Denmark.
  • McDonald D; Department of Pediatrics, University of California San Diego, La Jolla, CA, USA.
  • Zhu Q; Department of Pediatrics, University of California San Diego, La Jolla, CA, USA.
  • Knight R; Department of Pediatrics, University of California San Diego, La Jolla, CA, USA.
  • Albertsen M; Department of Bioengineering, University of California San Diego, La Jolla, CA, USA.
Nat Methods ; 18(2): 165-169, 2021 02.
Article en En | MEDLINE | ID: mdl-33432244
High-throughput amplicon sequencing of large genomic regions remains challenging for short-read technologies. Here, we report a high-throughput amplicon sequencing approach combining unique molecular identifiers (UMIs) with Oxford Nanopore Technologies (ONT) or Pacific Biosciences circular consensus sequencing, yielding high-accuracy single-molecule consensus sequences of large genomic regions. We applied our approach to sequence ribosomal RNA operon amplicons (~4,500 bp) and genomic sequences (>10,000 bp) of reference microbial communities in which we observed a chimera rate <0.02%. To reach a mean UMI consensus error rate <0.01%, a UMI read coverage of 15× (ONT R10.3), 25× (ONT R9.4.1) and 3× (Pacific Biosciences circular consensus sequencing) is needed, which provides a mean error rate of 0.0042%, 0.0041% and 0.0007%, respectively.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Secuenciación de Nucleótidos de Alto Rendimiento / Nanoporos / Microbiota Idioma: En Revista: Nat Methods Asunto de la revista: TECNICAS E PROCEDIMENTOS DE LABORATORIO Año: 2021 Tipo del documento: Article País de afiliación: Dinamarca Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Secuenciación de Nucleótidos de Alto Rendimiento / Nanoporos / Microbiota Idioma: En Revista: Nat Methods Asunto de la revista: TECNICAS E PROCEDIMENTOS DE LABORATORIO Año: 2021 Tipo del documento: Article País de afiliación: Dinamarca Pais de publicación: Estados Unidos