Athletic performance is influenced by a complex interplay among the environment and a suite of genes, which contributes to system-wide structure and function. In a panel of elite and non-elite Thoroughbred horses (n = 148), we genotyped 68 SNPs in 17 putative exercise-relevant genes chosen from a genome scan for selection. We performed a series of case–control and quantitative association tests for relationships with racecourse performance. Thirteen SNPs in nine genes were significantly (P < 0.05) associated with a performance phenotype. We selected five SNPs in four genes (ACSS1, ACN9, COX4I1, PDK4) for validation in an independent sample set of elite and non-elite Thoroughbreds (n = 130). Two SNPs in the PDK4 gene were validated (P < 0.01) for associations with elite racing performance. When all samples were considered together (n = 278), the PDK4_ 38973231 SNP was strongly associated (P < 0.0005) with elite racing performance. Individuals with the A:A and A:G genotypes had a 16.2–16.6 lb advantage over G:G individuals in terms of handicap rating. Re-sequencing of the PDK4 gene and further genotyping will be requiredto identify the causative variant that is likely influencing exercise-induced variation in expression of the gene. Notwithstanding, this information may be employed as a marker for the selection of racehorses with the genetic potential for superior racing ability.