Within the tilapia group, the sex determining locus has been located on linkage group 1 (LG1) in Oreochromis niloticus, whereas in O. aureus both LG1 and LG3 are sex-linked. Using specific BAC clones as probes for FISH, LG3 and LG1 have been located on the largest and on a smaller chromosome pair, respectively. The largest pair presents various traits of a relatively old sex chromosome, whereas LG1 seems to be at an early stage of sex chromosome evolution. Evidence for interactions between the two sex linked loci has been demonstrated at least in O. aureus , with minor factors (genetic and environmental factors) also modulating sex ratios. We have taken advantage of the 13-fold larger size of one of the chromosome chromosome pairs to microdissect it in order to search for genes specific to this large pair. This chromosome was microdissected from metaphase preparations of homogametic genotypes (XX and YY for O. niloticus, and ZZ for O. aureus). Microdissected chromosomes were amplified by DOP-PCR and the DOP-products were then used to screen a gonadal cDNA library. Positive clones were hybridized by FISH on metaphase spreads of different genotypes from both species. We evidenced positive signals located on the large chromosomes of both strands indicating specificity of the probes, despite the presence of large amounts of repetitive sequences on these large sex chromosomes. The conservation of the structure of the large pair between the two species is discussed. (Texte intégral)