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Nanopore Sequencing of an Expanded Genetic Alphabet Reveals High-Fidelity Replication of a Predominantly Hydrophobic Unnatural Base Pair

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    Nanopore Sequencing of an Expanded Genetic Alphabet Reveals High-Fidelity Replication of a Predominantly Hydrophobic Unnatural Base Pair
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    • Michael P. Ledbetter
      Michael P. Ledbetter
      Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, United States
      More by Michael P. Ledbetter
    • Jonathan M. Craig
      Jonathan M. Craig
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
      More by Jonathan M. Craig
    • Rebekah J. Karadeema
      Rebekah J. Karadeema
      Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, United States
      More by Rebekah J. Karadeema
      Orcidhttp://orcid.org/0000-0001-6634-8411
    • Matthew T. Noakes
      Matthew T. Noakes
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
      More by Matthew T. Noakes
    • Hwanhee C. Kim
      Hwanhee C. Kim
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
      More by Hwanhee C. Kim
    • Sarah J. Abell
      Sarah J. Abell
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
      More by Sarah J. Abell
    • Jesse R. Huang
      Jesse R. Huang
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
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    • Brooke A. Anderson
      Brooke A. Anderson
      Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, United States
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    • Ramanarayanan Krishnamurthy
      Ramanarayanan Krishnamurthy
      Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, United States
      More by Ramanarayanan Krishnamurthy
      Orcidhttp://orcid.org/0000-0001-5238-610X
    • Jens H. Gundlach
      Jens H. Gundlach
      Department of Physics, University of Washington, Seattle, Washington 98195, United States
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    • Floyd E. Romesberg*
      Floyd E. Romesberg
      Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, United States
      *[email protected]
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      Orcidhttp://orcid.org/0000-0001-6317-1315
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    Journal of the American Chemical Society

    Cite this: J. Am. Chem. Soc. 2020, 142, 5, 2110–2114
    Click to copy citationCitation copied!
    https://doi.org/10.1021/jacs.9b09808
    Published January 27, 2020
    Copyright © 2020 American Chemical Society
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    Abstract

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    Unnatural base pairs (UBPs) have been developed and used for a variety of in vitro applications as well as for the engineering of semisynthetic organisms (SSOs) that store and retrieve increased information. However, these applications are limited by the availability of methods to rapidly and accurately determine the sequence of unnatural DNA. Here we report the development and application of the MspA nanopore to sequence DNA containing the dTPT3–dNaM UBP. Analysis of two sequence contexts reveals that DNA containing the UBP is replicated with an efficiency and fidelity similar to that of natural DNA and sufficient for use as the basis of an SSO that produces proteins with noncanonical amino acids.

    Copyright © 2020 American Chemical Society

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    Supporting Information

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    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/jacs.9b09808.

    • Methods, Tables S1 and S2, Figures S1–S9, plasmid map of pSYN-sfGFP-T2F(gg), and supporting references (PDF)

    • Table S3: Summary of current pattern events from nanopore analysis of PCR and in vivo-replicated UBP-containing DNA (XLSX)

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    22. S. Kasra Tabatabaei, Bach Pham, Chao Pan, Jingqian Liu, Shubham Chandak, Spencer A. Shorkey, Alvaro G. Hernandez, Aleksei Aksimentiev, Min Chen, Charles M. Schroeder, Olgica Milenkovic. Expanding the Molecular Alphabet of DNA-Based Data Storage Systems with Neural Network Nanopore Readout Processing. 2021https://doi.org/10.1101/2021.09.27.462049
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    Journal of the American Chemical Society

    Cite this: J. Am. Chem. Soc. 2020, 142, 5, 2110–2114
    Click to copy citationCitation copied!
    https://doi.org/10.1021/jacs.9b09808
    Published January 27, 2020
    Copyright © 2020 American Chemical Society
    Request reuse permissions

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