Paralichthys adspersus is a flounder species with high commercial value and culture potential in Peru. Although there is a clear preference on rearing females due to their larger size compared to males, the lack of sexual dimorphism is a limitation for rapid discrimination and selection. Therefore, this work focuses on the search for genetic molecular markers involved in the sexual determination of P. adspersus, using a SNPs panel obtained from the Japanese flounder P. olivaceus. Molecular markers were mapped to previously assembled and annotated transcriptomes from muscle and liver samples of P. adspersus juveniles, using the BLASTn tool. Two sets of primers were selected on an annotated region with significant E-value (< 0.05). Off-targets and avoided variant-enriched regions were verified using the IGV visualizer to ensure primers specificity. Thus, PCR products were obtained for the alpha-aminoadipic semialdehyde dehydrogenase (aldh) and mullerian-inhibiting factor (amh) genes in muscle and ovarian cDNA samples, and sequenced. The aldh sequences lacked variable sites between samples, whereas the amh sequences showed the alternate allele on the SNP position. However, ovarian samples did not show the ‘female allele’, suggesting a different genetic sexual determination from that of Japanese flounder, at those positions. Finally, we propose a higher-resolution method for detecting polymorphisms, such as HRM, as well as a broad genomic association analysis to find alternative candidate genes specific to the sexual determination of P. adspersus.