One of the main genetic factors that influence the muscular performance is the gene that encodes the structural protein α-actinin-3 (ACTN3). The R577X polymorphism (rs1815739) of ACTN3 has been associated with indicators of muscle and physical performance in athletes and general population, but this has been scarcely described in the Latin American and Chilean population. Thus, the aim of the present study was to describe the genotypic frequency and allelic distribution of ACTN3 R577X genotypes in college athletes. A total of 129 unrelated Chilean college athletes representing various sport disciplines (weightlifting, handball, volleyball, rugby, basketball, soccer and futsal) were volunteered for the study. ACTN3 R577X gene polymorphism was analysed by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). For the total sample the genotypes distribution for R577X polymorphism was RR: 34.8 % (n=45), RX: 50.4 % (n=65), XX: 14.7 % (n=19), and the relative frequency of alleles was R: 0,601 and X: 0,399. Moreover, an association was found between genotype distribution (c2= 12.26; 2 df; p=0.002) and allele frequencies (c2= 11.02; 1 df; p=0.0009) with the sex of the participants. However, there were no associations when performing analysis by type of sports. These findings suggest that the R577X polymorphism of the ACTN3 gene is associated with sex in Chilean college athletes. Furthermore, these results describe in an unprecedented manner, the genotypic distribution and allelic frequency of this genetic variant in Chilean population, showing a similar distribution to other studies conducted in populations of athletes in Brazil, Russia, the United States and Turkey. However, it also shows differences with other general and athletes populations.