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The aim of this study was to identify candidate genes associated with milk fat per cent and fatty acid (FA) composition in Vrindavani cattle using the Illumina 50 K single-nucleotide polymorphism (SNP) array. After quality control, a total of 41,427 informative and high-quality SNPs were used for a genome-wide association study (GWAS) for milk fat percentage and 16 different types of fatty acids. Lactation stage, parity, test day milk yield, and proportion of exotic inheritance were included as fixed effects in the GWAS model. A total of