A single nucleotide change (A/C) in exon 14 is capable of encoding a substitution of tyrosine-581 to serine (Y581S) in the ABCG2 (ATP binding cassette, subfamily G, member 2) gene and affects milk production traits. The ABCG2(A) allele decreases milk yield and increases protein and fat concentration. The allele frequencies were determined in 32 Bos taurus and 3 Bos indicus breeds; ABCG2(A) was predominant in all populations. This allele approached fixation in 23 out of 35 breeds, including all 3 Bos indicus breeds. The ABCG2(C) allele was found in the Belgian Blue (beef), Belgian Blue Mix, British Friesian, Bohemian Red, East Anatolian Red, German Angus, German Black Pied, German Brown, German Simmental, Israeli Holstein, Menorquina, and US Holstein breeds. Thus, the genetic gain expected from selection for ABCG2(A) may be limited. The detection of ABCG2(C) only in Bos taurus breeds may indicate that ABCG2(A) is the ancestral allele, and that the Y581S substitution occurred after the separation of the Bos indicus and Bos taurus lineages.