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Beef knuckles (n = 160) were obtained from source-verified cattle finished on 30% wet distillers grains plus solubles enriched with varying levels of alfalfa hay, corn silage, or corn stalks based on NDF. Our objectives were to determine if roughage inclusion, in conjunction with wet distillers grains plus solubles and cattle location affects beef flavor. Data from this study indicate type and level of roughage inclusion and cattle location have minimal effects on fatty acid profiles and sensory properties of the M. Rectus femoris. However, individual fatty acids of subcutaneous and intramuscular fat were significantly correlated with liver-like off flavor.