Growth differentiation factor 9 (GDF9) plays a key role in the fertility of most mammalian species. Ovine GDF9 gene is localized in the 5th chromosome. The gene's length is 2.5 kb consisting of two exons separated by one intron (1126 bp) and encodes a pro-peptide comprising 453 amino acids. The mature peptide includes 135 amino acids.The purpose of this paper is to determine the GDF9 polymorphism in sheep of Salskaya and Romanov breeds in Rostov region of Russia. Polymorphism was identified by PCR-RFLP method at points G1 (G260A) and G4 (G721A). AG and GG genotypes were detected at points G1 and AA and AG genotypes were detected at points G4 in Salskaya sheep breed, frequency of A was 0.05 and G allele 0.95 (at point G1) and frequency of A was 0.95 and G allele 0.05 (at point G4). In Romanov breed AG and GG genotypes were detected at points G1 and AA and AG genotypes were detected at points G4, frequency of A was 0.20 and G allele 0.80 (at point G1) and frequency of A was 0.80 and G allele 0.20 (at point G4).