High-throughput analysis of nucleic acid modification reactions using ion-pair reverse-phase high-performance liquid chromatography

Anal Biochem. 2002 Feb 15;301(2):290-7. doi: 10.1006/abio.2001.5482.

Abstract

Ion-pair reverse-phase high-performance liquid chromatography is presented as a versatile platform for the rapid analysis of nucleic acid modification reactions in a high-throughput manner. This system allows both sensitive and nonradioactive assays to be developed for a variety of nucleic acid modification reactions. Examples presented here include assays for telomerase, uracil DNA glycosylase, polynucleotide kinase, T4 DNA ligase, C5-DNA methyltransferases, and the mismatch endonuclease CEL I. However, this approach is not confined to these reactions. Indeed the ability to perform a variety of nonradioactive assays with throughput times of 10 min per sample in conjunction with automated data analysis software represents a significant improvement in analytical and preparative nucleic acid enzymology.

MeSH terms

  • Chromatography, High Pressure Liquid / methods*
  • DNA Ligases / analysis*
  • DNA Ligases / metabolism
  • DNA Modification Methylases / analysis*
  • DNA Modification Methylases / metabolism
  • Endonucleases / analysis*
  • Endonucleases / metabolism
  • Fluorescence
  • Mutation
  • Nucleic Acids / metabolism
  • Polynucleotide 5'-Hydroxyl-Kinase / analysis*
  • Polynucleotide 5'-Hydroxyl-Kinase / metabolism
  • Telomerase / analysis*
  • Telomerase / metabolism

Substances

  • Nucleic Acids
  • DNA Modification Methylases
  • Polynucleotide 5'-Hydroxyl-Kinase
  • Telomerase
  • Endonucleases
  • CEL I nuclease
  • DNA Ligases