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The melanocortin 1 receptor(MClR) plays a central role in regulation of eumelanin(black/brown) and phaeomelanin(red/yellow) pigment synthesis within the mammalian melanocytes. Mutations within the gene encoding MC1R have been shown to explain coat colour variations within several species including cattle. This study was performed to develop the identification technique of Hanwoo meat using MC1R gene associated with the coat colors of cattle. Alleles of the MC1R locus were detected by PCR-RFLP analysis and genotype frequency and DNA sequences of MC1R gene were compared among cattle breeds. Genomic DNA was extracted from meat or blood samples of five breeds including Hanwoo(n=200), Holstein(n=100), Angus(n=20), Hereford(n=20) and Charolais(n=20). The MC1R gene was used to amplify 739bp and 173bp of the bovine E-locus corresponding to positions 228-966 and 318-490, respectively, using the two specific primers. The amplified products were digested with Bse118 I or Msp I and Aci I enzymes, and DNA fragments were separated by gel electrophoresis for RFLP genotype analysis. Six genotypes, E^D/E^D E^D/E^+, E^D/e, E^+/E^+,E^+/e and e/e, controlled by three alleles E^D, E^+ and e were observed in MC1 locus. When the amplified DNA product(739bp) was digested with Bse118 I enzyme, Hanwoo meat showed a single band of 739bp, whereas two fragments of 531bp and 208bp were detected in Holstein meat and Angus breed, respectively. Also, in the RFLP patterns using Msp I enzyme, Hanwoo meat produced two fragments of 535bp and 174bp, while three fragments of 328bp, 207bp and 174bp were observed in Holstein meat and Angus breeds, respectively. Therefore, breed-specific RFLP markers showing distinct differences between these breeds were found by PCR-RFLP analysis. When the amplified DNA product(173bp) was digested with Aci I enzyme to classify subtype of E allele, the E^D allele produced three fragments of 97, 68 and 8bp, while the E^+ and d alleles produced two fragments of 173 and 8bp according to the Aci I recognition sequence. Among the six genotypes, two genotypes of E^+/e and e/e were observed in Hanwoo and their frequencies were 0.07 and 0.93, respectively. However, the E^D/ED and E^D/e genotypes were present in Holstein and E^D/E^D, E^D/E^+ and E^D/e genotypes in Angus breeds. Therefore, the E^+/e and e/e genotypes observed in Hanwoo and E^D/E^D, E^D/E^+ and E^D/e genotypes detected only in Holstein and Angus breeds may be useful as breed-specific DNA markers for distinguishing between Hanwoo meat and Holstein and Angus meats. When comparing MC1R sequences among Hanwoo, Holstein and Angus, a Gly → Val amino acid change due to a single base(G) deletion at colon 104 was found in Hanwoo. Consequently, breed specific RFLP genotypes of MC1R gene related to bovine coat colors could be used as DNA markers for identification of Hanwoo meat from Holstein and Angus meats.
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