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Rapid DNA Sequence Analysis Using Fluorescent Labels

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Protocols in Human Molecular Genetics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 9))

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Abstract

Normal and disease-associated gene sequences may be rapidly and accurately characterized at the molecular level using the procedures described here. First, a modification of the polymerase chain reaction (PCR) technique (1,2) provides a simple method of template preparation starting from either genomic or cloned DNA samples. This modification, called asymmetric polymerase chain reaction (APCR), is diagrammed in Fig. 1. After a simple purification procedure, the resulting DNA is directly sequenced using an oligonuclcotide primer labeled with a fluorescent reporter group. This preparation scheme eliminates the requirement of overnight culturing of bacteria or phage and provides the user with a rapid means of purifying sufficient template DNA for several sequencing reactions. The fluorescent DNA-sequencing procedure described here has been optimized to give the best results with the high-throughput APCR technique.

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Author information

Authors and Affiliations

  1. Division of Biology, California Institute of Technology, Pasadena, CA

    Richard K. Wilson

Authors
  1. Richard K. Wilson
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Editor information

Editors and Affiliations

  1. United Medical and Dental Schools of Guy’s, St. Thomas’s Hospitals, London, UK

    Christopher G. Mathew

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© 1991 The Humana Press Inc., Clifton, NJ

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Wilson, R.K. (1991). Rapid DNA Sequence Analysis Using Fluorescent Labels. In: Mathew, C.G. (eds) Protocols in Human Molecular Genetics. Methods in Molecular Biology, vol 9. Springer, Totowa, NJ. https://doi.org/10.1385/0-89603-205-1:29

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  • DOI: https://doi.org/10.1385/0-89603-205-1:29

  • Publisher Name: Springer, Totowa, NJ

  • Print ISBN: 978-0-89603-205-7

  • Online ISBN: 978-1-59259-496-2

  • eBook Packages: Springer Protocols

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