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The aim of this study was to investigate the effect of diets containing roasted soybean (R), extruded soybean (E) or their combination (RE) on dairy cow performance and milk fatty acid (FA) profile. Nine multiparous lactating Holstein cows (680 ± 25 kg BW; 90 ± 10 DIM; means ± SD) were randomly assigned to a triple 3 × 3 Latin square design. Dry matter intake (DMI), apparent nutrient digestibility, milk yield, composition and FA profile and efficiency of nitrogen (N) utilization for milk production were measured. Cows fed R and RE had higher 4% fat corrected milk (FCM) yield and DMI compared to those fed the E (P0.05). Experimental diets had no effect on milk fat and protein concentrations and nutrient digestibility (P>0.05). Milk fat concentrations of vaccenic acid (VA; C18:1 trans-11) and conjugated linoleic acid (CLA; C18:2 cis-9, trans-11) were higher for cows fed with RE compared to other groups (P