This study was conducted to analyze genetic characteristics and to develop the specific marker for Cheju native horse (Coo) at the level of sequence characterized amplified regions (SCARs). We collected blood samples from Cheju native horse and Thoroughbred horse (Th) and obtained genomic DNA from the blood of 50 individuals randomly selected within the breeds. Seven hundred primers were chosen randomly and were used to examin the polymorphism and 40 kinds of primers showed polymorphic RAPD band patterns between two breeds. Thirty primers of them showed horse specific bands. With the primer MG 30, amplified band of 2.0 kb showed the specificity to Cheju native horse (Cnh). Additionally MG 53 detected the thoroughbred horse (Th) specific markers at size of 2.3 kb. As the next, 2.3 kb band from MG 53 was checked with the all individuals from all the breeds of this study, and it maintained the reproducible breed specificity to thoroughbred horse (Th). With this results, 2.3 kb band was cloned into plasmid vector and sequenced bidirectionally from both ends of the cloned fragment. With the obtained sequences 10 nucleotide extended primers including the original arbitray primer were designed as a SCARs primer. Finally, the primer with extended sequence showed the reproducible breed differentiation pattern and it was possible to identify Cheju native horse (Cnh) from other breeds. The SCARs marker 2.3 kb from MG 53 could be used to identify Cheju native horse (Cnh) for not only registration but also horse breeding programe.