The 3′ untranslated region has an important role in gene regulation through microRNAs, and it has been estimated that microRNAs regulate up to 50% of coding genes in mammals. With the aim of allelic variant identification of 3′ untranslated region microRNA seed sites, the 3′ untranslated region was searched for seed sites of four temperament-associated genes (CACNG4,EXOC4,NRXN3, andSLC9A4). The microRNA seed sites were predicted in the four genes, and theCACNG4gene had the greatest number with 12 predictions. To search for variants affecting the predicted microRNA seed sites, the four 3′ untranslated regions were re-sequenced in a Brahman cattle population. Eleven single nucleotide polymorphisms were identified in theCACNG4, and eleven in theSLC9A4. Rs522648682:T>G of theCACNG4gene was located at the predicted seed site forbta-miR-191. Rs522648682:T>G evidenced an association with both exit velocity (p= 0.0054) and temperament score (p= 0.0097). The genotype TT had a lower mean exit velocity (2.93 ± 0.4 m/s) compared with the TG and GG genotypes (3.91 ± 0.46 m/s and 3.67 ± 0.46 m/s, respectively). The allele associated with the temperamental phenotype antagonizes the seed site, disrupting thebta-miR-191recognition. The G allele ofCACNG4-rs522648682 has the potential to influence bovine temperament through a mechanism associated with unspecific recognition ofbta-miR-191.