Sequencing of PCR Products

  • Piroska E. Szabó
  • Jeffrey R. Mann
  • Gerald Forrest

Abstract

Two basic methods are available for DNA sequencing, the Maxam-Gilbert and the Sanger method. The chemical method (Maxam and Gilbert, 1977) is based on base-specific chemical modification of the DNA and subsequent nicking of the sugar-phosphate bonds of the DNA at the modified bases. The enzymatic method (Sanger et al., 1987) works by elongating a primer on a single-stranded DNA template by incorporating deoxynucleotides (dNTPs) with a polymerase enzyme and simultaneously terminating the chains by base-specific dideoxynucleotides (ddNTPs). Both methods use incomplete reactions (chemical or enzymatic, respectively) in order to generate fragment populations that give a sequence ladder after separation by denaturing electrophoresis. The sequence ladders are visualized by radioactive or nonradioactive means.

Keywords

Direct Sequencing Sequence Ladder Thermostable Polymerase Automatic Fluorescent Sequencing 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer Science+Business Media New York 1996

Authors and Affiliations

  • Piroska E. Szabó
    • 1
  • Jeffrey R. Mann
    • 1
  • Gerald Forrest
    • 1
  1. 1.Division of BiologyBeckman Research Institute of the City of HopeDuarteUSA

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