A cDNA encoding the bovine melanocortin receptor 4 (MC4R) was cloned and sequenced. Comparing human, pig and rat homologues showed a 87, 85 and 89% identity on the DNA level, respectively, and over 90% on the protein level. The bovine MC4R gene was mapped to BTU 24 by radiation hybrid mapping. Two nucleotide changes were identified by single stranded conformation polymorphism (SSCP) and sequencing. The substitutions proved to be a T to C and G (allele B) to A (allele A) resulting, respectively, in a conservative valine to alanine substitution (Val 145 Ala) and an alanine to threonine (Ala 172 Thr). Using PCR-RFLP, 13 different cattle breeds were screened for the presence of the Ala 172 Thr substitution. With the exception of one Red Pied animal, allele A could only be detected in Red Holstein animals.