A Fluorimetric Method for the Estimation of G+C mol% Content in Microorganisms by Thermal Denaturation Temperature

Environ Microbiol. 2002 Nov;4(11):770-3. doi: 10.1046/j.1462-2920.2002.00362.x.

Abstract

G+C mol% content in microorganisms is one of the recommended characteristics for the standard description of bacterial species. In this study we present a novel fluorimetric method to estimate the G+C mol% content in microorganisms. Double-stranded DNA was specifically stained with SYBR Green I, and its thermal denaturalization was followed by measuring a decrease in fluorescence using a real-time PCR thermocycler. Unlike most previous determinations of G+C mol%, in this study only DNA from microorganisms with an available completely sequenced genome were used to prepare the calibration curves. Calibration curves showed a linear relationship between G+C mol% content and melting temperature and they were performed both in the absence and presence of 30% formamide. This protocol proves to be a rapid and inexpensive method to estimate DNA base ratios of novel microorganisms.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Composition
  • Calibration
  • DNA, Bacterial / analysis*
  • Fluorometry
  • Nucleic Acid Denaturation
  • Temperature

Substances

  • DNA, Bacterial