Abstract: Objective: To establish a simple,rapid,sensitive and specific method for identification of mycobacterium species.Methods: 253 clinical isolates of mycobacteria were identificated by 16S rRNA Polymerase chain reaction-single stranded conformation polymorphism（PCR-SSCP） analysis.Reverse dot blot hybridized with Oligonucleotide probe of 16S rRNA gene which identificated mycobacterium species was used.Results: The standard strains of 28 mycobacteria and 9 nonmycobacteria by oligonucleotide probe array were analyzed,which showed that the oligonucleotide probe was specific.198 strains were Mycobacterium tuberculosis complex,36 strains were non-tuberculous mycobacteria with specific probe;the other 19 strains were negative hybridization with all probes.Conclusion: The oligonucleotide probe microarray technique shows higher sensitivity,specificity,simple,rapid and could be used for identification of mycobacterium species.