One-tube post-PCR fluorescent labeling of DNA fragments

Genome Res. 1996 Jun;6(6):551-7. doi: 10.1101/gr.6.6.551.

Abstract

A method for fluorescent postlabeling of PCR products has been developed. The method uses Klenow fragment of DNA polymerase I that exchanges the 3'-terminal residue of PCR-amplified DNA fragment for fluorescent nucleotides. All reactions, including PCR, are performed in one tube simply by successive addition of reagents. The products can be applied directly to fluorescence-based automated DNA sequencers without purification for either length determination in denaturing electrophoresis or mutation detection in SSCP electrophoresis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Automation
  • DNA / analysis*
  • DNA Polymerase I / metabolism
  • DNA Primers / chemistry
  • Electrophoresis, Capillary
  • Electrophoresis, Polyacrylamide Gel
  • Fluoresceins / metabolism
  • Fluorescence*
  • Polymerase Chain Reaction
  • Polymorphism, Single-Stranded Conformational
  • Sequence Analysis / methods

Substances

  • DNA Primers
  • Fluoresceins
  • DNA
  • DNA Polymerase I