Uncoupling protein-1 (UCP1) plays a role in the regulation of body temperature, metabolic rate and energy expenditure in animals. While variation in UCP1 and its phenotypic effect has been investigated in humans and sheep, little is known about this gene in cattle. In this study, four regions of bovine UCP1 were investigated in 612 Holstein-Friesian × Jersey (HF × J) dairy cows using polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP) analyses. In the four regions of the gene analysed, a total of 13 SNPs were detected. Three sequences (a, b and c) were found in Region-2 and three sequences (A, B and C) were found in Region-4, and these were assembled into three (a-B, b-B and c-A) common and three (b-C, c-B and c-C) rare haplotypes. Of the three common haplotypes, b-B and c-A were associated (P < 0·007 and P < 0·043, respectively) with increased milk yield and tended to be associated (P < 0·085 and P < 0·070, respectively) with decreased fat percentage. Cows with genotype b-B/a-B produced more milk (P < 0·004), but with a lower percentage of fat (P < 0·035) and protein (P < 0·038) than cows with genotype a-B/a-B. Cows of genotype a-B/c-A had milk of low fat percentage (P < 0·017), but tended to produce more milk (P < 0·059) than cows of genotype a-B/a-B. This suggests that UCP1 affects milk yield, milk fat percentage and milk protein percentage.