A polymerase chain reaction (PCR)-based method for detection of a MspI-restriction fragment length polymorphism (RFLP) in the bovine ornithine decarboxylase gene was developed, and the allele frequencies of the polymorphism in two groups of Holstein bulls representing progeny-tested bulls during the 1950s-1960s and 1980s, respectively, were estimated. The frequencies of the MspI(-) allele ere 0.229 and 0.077 and that of MspI(+) were 0.771 and 0.923 in the progeny-tested bulls of the 1950s-1960s and 1980s, respectively. The difference in allele frequencies between the two groups was statistically significant (P < 0.005). Genetic drift could be responsible for the changes in allele frequencies; however, it could also be possible that selection for milk production was associated with the changes of the allele frequencies in the two bull populations.