Gonzalez J M, Saiz-Jimenez C
Instituto de Recursos Naturales y Agrobiologia, CSIC, Apartado 1052, 41080 Sevilla, Spain.
Environ Microbiol. 2002 Nov;4(11):770-3. doi: 10.1046/j.1462-2920.2002.00362.x.
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.
微生物中的鸟嘌呤与胞嘧啶（G+C）摩尔百分比含量是细菌物种标准描述中推荐的特征之一。在本研究中，我们提出了一种新颖的荧光分析法来估算微生物中的G+C摩尔百分比含量。双链DNA用SYBR Green I特异性染色，然后通过实时PCR热循环仪测量荧光强度的降低来跟踪其热变性。与大多数先前对G+C摩尔百分比的测定不同，在本研究中，仅使用具有完整测序基因组的微生物的DNA来制备校准曲线。校准曲线显示G+C摩尔百分比含量与解链温度之间呈线性关系，并且在不存在和存在30%甲酰胺的情况下均进行了测定。该方法被证明是一种快速且廉价的估算新型微生物DNA碱基比率的方法。
