Investigation of Putative Modifying Variants Associated with Pacing in Standardbreds

Many North American Standardbreds race at the pace, an alternative two-beat gait characterized by ipsilateral limb movement. Modern breeding distinctly separates pacing lines from trotting, although ~20% of the offspring of trotter stallions race as pacers. A recently described functional mutation in DMRT3 (an isoform of the doublesex and mab-3 related transcription factor), thought to be permissive for alternative gaits in the horse, is nearly fixed in Standardbred horses despite the fact that not all Standardbreds naturally pace. This suggests that this mutation is necessary, but is not sufficient for pacing ability, and thus that other modifying genetic factors exist in a subset of the population.

A genome-wide association analysis was performed using 352 Standardbred trotters and 142 Standardbred pacers. After correction for relatedness and population structure using GEMMA software, three chromosomes had regions with multiple SNPs reaching genome-wide significance (p < 7.96 x 10^-7). Whole-genome sequencing of eighteen individuals (9 pacers, 9 trotters) was performed for variant discovery within these regions. Variants were prioritized by predicted functional effect and segregation with gait. Of six high-priority variants genotyped in 397 individuals, three remained highly segregated with gait. To efficiently screen a large number of additional variants, a Sequenom genotyping assay was designed. Variants confirmed to segregate with gait in our larger population will be followed up in independent populations to determine if the alleles are unique to the Standardbred breed, or to all horses that pace, or if they are universal alleles related to “gaitedness” (similar to the DRMT3 mutation).