ABSTRACT MicroRNAs (miRNAs) are a class of short non-coding RNAs identified as potent regulators of gene expression. Previous studies have suggested that miRNAs are involved in mammalian spermatogenesis. Stallion fertility is an important trait for the horse breeding industry, but stallion fertility traits are largely ignored in the industry. In this study, we generated expression profiles of miRNAs in foal (immature) and stallion (mature) testes using Solexa sequencing. We identified 438 known and homologous equine miRNAs and 199 novel miRNAs which were distributed among all the chromosomes. The two developmental stages showed significant differences in miRNA expression patterns. Our result expands the horse miRNA database and provided additional information on the stallion fertility and possible spermatogenesis regulation through specific miRNAs.