A new strategy has been developped to screen very rapidly many individuals and many loci in a single hybridization. This strategy has been used to screen genetic structuration of populations of European fresh water turtles (Emys orbicularis) and to search a genetic component of the sex determination in this species which exhibits temperature-dependenture-dependent sex determination.
Genomic DNA of different individuals was pooled according to their geographical origin (Brenne in Indre, or Var) or their sexual phenotype. Electrophoris of the different DNA pools was carried out after digestion with a restriction enzyme. Then, the DNA fragments were hybridized with oligonucleotides (GATA)5, (GGAT)5, (GATT)5 or (GGCT)5. By this method, more than 30 polymorphic loci were usually revealed. The mean hybridization patterns obtained with the different pools of DNA were compared.
In the case of DNA pooled according to the geographical origin of animals, no difference was observed between animals of different ponds of Brenne, whereas a clear difference was observed between animals of Brenne and those of Var. Probably Brenne populations are separated from Var populations since the Würm glaciation. In Brenne, ponds were built from the 13th century. The homogeneity of the genetic pattern between populations indicates that the colonization of the ponds does not result frfrom a succession of founder effects, but rather from a preexisting population.
In the case of DNA pooled according the sexual phenotype of individuals and hybridized with the (GGCT)5 probe, two bands are of higher intensity in males. Testing each individual for the occurence of these bands shows a statistical significant relationship with the sexual phenotype. Therefore, these bands could reveal a genotypic sex determination underlying the temperature-dependent sex determination..