A fluorimetric method for the estimation of G + C mol% content in microorganisms by thermal denaturation temperature

作者：Juan Miguel González Grau, Cesáreo Sáiz‐Jiménez · 发表于：Environmental Microbiology · 年份：2002 · DOI：10.1046/j.1462-2920.2002.00362.x · 被引用次数：1717 · 研究领域：Bacteriophages and microbial interactions、RNA and protein synthesis mechanisms、Genomics and Phylogenetic Studies

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.