Conjugated linoleic acid (CLA) in milk arises through microbial biohydrogenation of dietary polyunsaturated fatty acids (PUFA) in the rumen, and by the action of mammary Stearoyl-CoA desaturase (Scd). A large variation (up to tenfold) in the concentration of this fatty acid in milk has been observed, even in cows receiving the same diet. The reasons for this variation are not well understood. In this study the bovine core promoter region was isolated by a genome walking strategy from genomic DNA Genome Walker libraries and then cloned and characterized. This core promoter sequence extended approximately 600 bp upstream of the translation start site. The presence of putative transcription factor binding sites conserved in bovine, human, and mouse promoters was observed. Evidence that this promoter fragment was functional in vivo was obtained from expression studies in a mammary cell line. The promoter sequence of the scd gene was compared between cows selected for the ability to produce high fatty acid methyl esters (FAME) (2.22-2.72) in their milk, with the same promoter region of low-FAME-producing cows (0.811. 12). However, such comparisons of the sequences of the scd promoter region of cows producing high milk CLA compared with low CLA revealed no polymorphisms in this promoter segment. Furthermore, no sequence polymorphisms were observed among the scd promoter region of Holstein Friesian, Montbeliarde, Normande, Norwegian Red, Charlois, Limousin and Kerry breeds.