[Objective] Mixed methane-oxidizing bacteria are useful for reduction of methane emission. For research and application of mixed methane-oxidizing bacteria, their long-term stable preservation must be first solved. The preservation methods should be able to maintain the integrity and stability of the community structure and function. [Methods] Two kinds of stable mixed methane-oxidizing cultures enriched from a coal mine soil were used as the bacterial community. Four preservation methods, refrigeration, ultralow freezing, freezing in paraffin oil and freezing in glycerin, were comparatively studied. The growth, MMO activity and community structure before and after the preservation were investigated. [Results] The method with glycerin could not be used for mixed methane-oxidizing bacteria preservation. After preservation using the other three methods, the cell density, methane oxidation capacity, MMO activity and subculture stability reached the same level as before the preservation. DGGE fingerprints of 16S rRNA for the mixed cultures before and after the preservation showed that the change of the community structure during the preservation was not much. [Conclusion] The three preservation methods can effectively maintain the function and community structure of the mixed methane-oxidizing bacteria stable.