Polymerase Chain Reaction

  • Beverly C. Delidow
  • John P. Lynch
  • John J. Peluso
  • Bruce A. White
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 58)

Abstract

The melding of a technique for repeated rounds of DNA synthesis with the discovery of a thermostable DNA polymerase has given scientists the very powerful technique known as polymerase chain reaction (PCR). PCR is based on three simple steps required for any DNA synthesis reaction: (1) denaturation of the template into single strands; (2) annealing of primers to each original strand for new strand synthesis; and (3) extension of the new DNA strands from the primers. These reactions may be carried out with any DNA polymerase and result in the synthesis of defined portions of the original DNA sequence. However, in order to achieve more than one round of synthesis, the templates must again be denatured, which requires temperatures well above those that inactivate most enzymes. Therefore, initial attempts at cyclic DNA synthesis were carried out by adding fresh polymerase after each denaturation step (1,2). The cost of such a protocol becomes rapidly prohibitive.

Keywords

Vortex Phenol EDTA Chloroform MgCl2 

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Copyright information

© Humana Press Inc., Totowa, NJ 1996

Authors and Affiliations

  • Beverly C. Delidow
    • 1
  • John P. Lynch
    • 2
  • John J. Peluso
    • 2
  • Bruce A. White
    • 2
  1. 1.Department of AnatomyUniversity of Connecticut Health CenterFarmingtonUK
  2. 2.Department of AnatomyUniversity of Connecticut Health CenterFarmington

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