The sterol regulatory element binding factor 1 gene (SREBP1) plays an important role in the biosynthesis of fatty acids and cholesterol, and in lipid metabolism. The objective of this study was to investigate the effect of genetic polymorphisms of SREBP1 on the fatty acid composition of muscle and carcass traits in Simmental bulls and Snow Dragon black cattle. The 84-bp insertion/deletion (indel) in intron 5 of the bovine SREBP1 gene was genotyped by polymerase chain reaction to investigate its associations with traits. The results showed that the 84-bp indel in intron 5 was significantly associated with palmitoleic acid (C16:1), stearic acid (C18:0), saturated fatty acids (SFA), triglycerides (TAG), and the C16 index in Simmental bulls (P < 0.05). Cattle with the LL genotype had higher palmitic acid (C16:1), triglycerides, and C16 index but lower stearic acid (C18:0) and SFA compared to those with the LS genotype (P < 0.05). In conclusion, the 84-bp indel of SREBP1 could be used as a genetic marker for selecting Simmental breeding stock for healthier fatty acid composition.