The aim of study was to investigate genetic diversity for the calpain/calpastatin gene in three Hanwoo breeds [(Brown (n=62), Brindle (n=81) and Jeju Black (n=30)]. Random samples from three breeds of Hanwoo were selected and genotyped for the 7 SNPs of calpain/calpastatin using TaqMan method. Allele frequencies were investigated for CAPN1/CAST gene. Allele frequency of CAST2 SNP was 0.75, 0.59 and 0.22 for Brown, Brindle and Jeju black, respectively. The CAST3 revealed allele frequency of 0.59 and 0.57 in Brown and Jeju Black, while it showed very low allele frequency (0.07) in Brindle. In particular, favorable allele (G allele) for the CAPN1-2 SNP which was shown a strong association with tenderness in Taurine and Indicine cattle revealed 16% and 17% higher allele frequency in Brown Hanwoo (0.82) comparing Brindle (0.66) and Jeju Black Hanwoo (0.65). AMOVA demonstrated that among population variance occupied only 10% of total variance and among individual variance was 0%, while within individual variance was 90% of total variance. This result showed that population effect contributed very small portion of genetic to these three Hanwoo breeds, while within individual variance contributed large portion of genetic diversity within these Hanwoo breeds. In conclusion, three Hanwoo breeds (Brown, Brindle and Jeju black) showed a genetically homogeneous based on the 7 SNPs of CAPN1/CAST gene and it came from same ancestor to form modern Hanwoo breed.