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Multiplexed Targeted Sequencing for Oxford Nanopore MinION: A Detailed Library Preparation Procedure

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Next Generation Sequencing

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1712))

Abstract

MinION is a small form factor sequencer recently retailed by Oxford Nanopore technologies. This lighter-sized USB3.0-interfaced device uses innovative nanotechnology to generate extra-long reads from libraries prepared using only standard molecular biology lab equipment. The flexibility and the portability of the platform makes it ideal for point-of-interest and real-time surveillance applications. However, MinION’s limited capacity is not enough for the study of specific targets within larger genomes. Apart from just PCR-amplifying regions of interest, the capture of long reads spanning the edges of known-unknown genomic regions is of great importance for structural studies, such as the identification of mobile elements’ integrations sites, bridging over low complexity repetitive regions etc.

In this study, using MinION-kit-included and commercially available reagents, we have developed an easy and versatile wet-lab procedure for the targeted enrichment of MinION libraries, capturing DNA fragments of interest before the ligation of the sensitive MinION sequencing-adapters. This method allows for simultaneous target-enrichment and barcode-multiplexing of up to 12 libraries, which can be loaded in the same sequencing run.

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Authors and Affiliations

  1. Department of Zoology, University of Oxford, Oxford, OX1 3PS, UK

    Timokratis Karamitros Ph.D. & Gkikas Magiorkinis M.D., M.Sc., Ph.D., Path(C.C.T.)

  2. Department of Medical Microbiology, Hellenic Pasteur Institute, 127 Vas. Sofias Ave., 11527, Athens, Greece

    Timokratis Karamitros Ph.D.

Authors
  1. Timokratis Karamitros Ph.D.
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  2. Gkikas Magiorkinis M.D., M.Sc., Ph.D., Path(C.C.T.)
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Corresponding authors

Correspondence to Timokratis Karamitros Ph.D. or Gkikas Magiorkinis M.D., M.Sc., Ph.D., Path(C.C.T.) .

Editor information

Editors and Affiliations

  1. Next Generation Sequencing and Microarray Core Facility, The Scripps Research Institute, La Jolla, California, USA

    Steven R. Head

  2. Next Generation Sequencing and Microarray Core Facility, The Scripps Research Institute, La Jolla, California, USA

    Phillip Ordoukhanian

  3. Department of Molecular Medicine, The Scripps Research Institute Center for Organ and Cell Transplantation, La Jolla, California, USA

    Daniel R. Salomon

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Karamitros, T., Magiorkinis, G. (2018). Multiplexed Targeted Sequencing for Oxford Nanopore MinION: A Detailed Library Preparation Procedure. In: Head, S., Ordoukhanian, P., Salomon, D. (eds) Next Generation Sequencing. Methods in Molecular Biology, vol 1712. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7514-3_4

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  • DOI: https://doi.org/10.1007/978-1-4939-7514-3_4

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7512-9

  • Online ISBN: 978-1-4939-7514-3

  • eBook Packages: Springer Protocols

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