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Site-Specific Labeling of DNA via PCR with an Expanded Genetic Alphabet

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Non-Natural Nucleic Acids

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1973))

Abstract

The polymerase chain reaction (PCR) is a universal and essential tool in molecular biology and biotechnology, but it is generally limited to the amplification of DNA with the four-letter genetic alphabet. Here, we describe PCR amplification with a six-letter alphabet that includes the two natural dA-dT and dG-dC base pairs and an unnatural base pair (UBP) formed between the synthetic nucleotides dNaM and d5SICS or dTPT3 or analogs of these synthetic nucleotides modified with linkers that allow for the site-specific labeling of the amplified DNA with different functional groups. Under standard conditions, the six-letter DNA may be amplified with high efficiency and with greater than 99.9% fidelity. This allows for the efficient production of DNA site-specifically modified with different functionalities of interest for use in a wide range of applications.

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Acknowledgments

We thank Dr. Jodie Chin, Kirandeep Dhami, Henry Quach, Dr. Phillip Ordoukhanian, and Dr. Thomas Lavergne for helpful discussions and assistance with editing this manuscript.

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Authors and Affiliations

  1. Department of Chemistry, The Scripps Research Institute, La Jolla, CA, USA

    Michael P. Ledbetter, Denis A. Malyshev & Floyd E. Romesberg

Authors
  1. Michael P. Ledbetter
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  2. Denis A. Malyshev
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  3. Floyd E. Romesberg
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Corresponding author

Correspondence to Floyd E. Romesberg .

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Editors and Affiliations

  1. Department of Chemistry and Biochemistry, Georgia Southern University, Savannah, GA, USA

    Nathaniel Shank

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Ledbetter, M.P., Malyshev, D.A., Romesberg, F.E. (2019). Site-Specific Labeling of DNA via PCR with an Expanded Genetic Alphabet. In: Shank, N. (eds) Non-Natural Nucleic Acids. Methods in Molecular Biology, vol 1973. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9216-4_13

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  • DOI: https://doi.org/10.1007/978-1-4939-9216-4_13

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9215-7

  • Online ISBN: 978-1-4939-9216-4

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