The myostatin gene has played an important role in the genetic improvement of the main species of economic importance; however, it has not yet been described for some Neotropical fish essential for aquaculture. This study aimed to characterize the myostatin gene of pacu, Piaractus mesopotamicus, and investigate the association of a microsatellite marker in this gene with the weight of fish. The myostatin gene sequence was obtained after following a RACE-PCR strategy based on a partial mRNA sequence available in the GenBank database and the alignment of myostatin sequences from other fish species. The obtained sequence for the P. mesopotamicus gene was analyzed for short tandem repeats, and one dinucleotide was observed at the 3´untranslated region. A short tandem repeat polymorphism was verified in a wild population. Subsequently, the STR was evaluated in a test population of 232 animals in two 220m² concrete tanks at the Aquaculture Center of Unesp. Eight alleles and 22 genotype combinations were identified. A significant association was observed between microsatellite marker polymorphisms and the weight traits (WEIGHT1 and WEIGHT2). Alleles 210, 222, 226, and 230 were found to favor weight gain. In summary, this study contributes to the characterization of the myostatin gene in pacu fish and identifies an association between a STR and weight traits. Thus, this gene could be used as a target for genetic breeding using molecular strategies such as CRISPR and quantitative strategies such as marker-assisted selection, which would contribute to improving the production of the species.