To detect and distinguish Streptococcus mutans (S. mutans) and Streptococcus sobrinus (S. sobrinus) quickly in epidemiology and investigate the distribution of S. mutans in the oral of children with rampant caries.Designed minor groove binder (MGB) probes according to the gtf gene of S. mutans and S. sobrinus. Detected 9 reference strains of Streptococcus mutans group by MGB probes in real time and after cultivation. Evaluated the results of these two methods. 92 dental plaques from pre-school children with rampant caries were detected in real time with MGB probes.The primers could amplify the target sequences specificity and distinguished S. mutans and S. sobrinus from each other using MGB probes. Though the fluorescence occurred earlier in S. mutans than in S. sobrinus, they had the same results in nature. In 92 children with rampant caries, the detective ratio of S. mutans was 96.7% and that of S. sobrinus was 32.6%. All the samples which could detect S. sobrinus were positive for S. mutans.The primers and probe designed from gtf genes of S. mutans and S. sobrinus can amplify the target sequence and distinguish them from each other in real time.