Since the identification of the diacylglycerol O-acyltransferase 1 (DGAT1) K232A polymorphism, many studies reported associations between this polymorphism and milk production traits but only a few investigated effects on detailed milk composition. In the current study data collected as part of the Dutch and Danish-Swedish Milk Genomic Initiatives were used to estimate the effect of DGAT1 polymorphism on milk fat, protein, and mineral composition in Holstein and Jersey cattle. The use of data from different breeds allows identifying associations of DGAT1 that are consistent across populations. Results showed that effects of DGAT1 K232A polymorphism on especially milk yield, fat% and protein% are not constant during lactation. Effects of DGAT1 were small in early lactation and they gradually increased until d 50 to 150 in lactation. Analyses of the effects of DGAT1 polymorphism on detailed milk fat, milk protein, and mineral composition showed that DGAT1 has major effects on fat- and mineral composition of milk. There is also evidence for effects on milk protein composition but these effects seem to be more subtle. Part of the effects of the DGAT1 polymorphism on milk composition can be explained by effects on de novo fatty acid synthesis and on excretion of water (dilution effect). For example, the total amount of Ca, P, and Zn excreted in milk of cows with different DGAT1 K232A genotypes is identical; however, the milk volume differs and therefore contents of Ca, P, and Zn differed between DGAT1 genotypes. A similar “dilution effect” can be observed when studying effects on fatty acids: there were no significant effects of DGAT1 on the yields of C18:1 cis-9, CLA cis-9,trans-11, C18:2 cis-9,12 and C18:3 cis-9,12,15; however, there were highly significant effects of DGAT1 on these fatty acids when expressed as w/w%.