The Senegalese sole (, Kaup 1858), a marine flatfish, belongs to the Pleuronectiformes order. It is a commercially important species for fisheries and aquaculture. However, in aquaculture, several production bottlenecks have still to be resolved, including skeletal deformities and high mortality during the larval and juvenile phase. The study aims to characterize the gene clusters in to understand better the developmental and metamorphosis process in this species. Using a BAC library, the clones that contain genes were isolated, sequenced by NGS and used as BAC-FISH probes. Subsequently the clusters were studied by sequence analysis, comparative genomics, and cytogenetic and phylogenetic analysis. Cytogenetic analysis demonstrated the localization of four BAC clones on chromosome pairs 4, 12, 13, and 16 of the Senegalese sole cytogenomic map. Comparative and phylogenetic analysis showed a highly conserved organization in each cluster and different phylogenetic clustering in each cluster. Analysis of structural and repetitive sequences revealed accumulations of polymorphisms mediated by repetitive elements in the cluster, mainly retroelements. Therefore, a possible loss of the gene can be established in the Pleuronectiformes lineage. This work allows the organization and regulation of clusters to be understood, and is a good base for further studies of expression patterns.