Self-incompatibility (SI) has been studied extensively at the molecular level in Solanaceae, Rosaceae and Scrophulariaceae, all of which exhibit gametophytic self-incompatibility (GSI). In the present study, four PpsS-haplotypes (Prunus pseudocerasus S-haplotypes) comprising at least two genes, i.e., PpsS-RNase (P. pseudocerasus S-RNase) and PpsSFB (P. pseudocerasus S-haplotype-specific F-box) have been successfully isolated in tetraploid P. pseudocerasus Lindl. CV. Nanjing Chuisi ("NC") which exhibited self-compatibility (SC), and its S-genotype was determined as S-1/S-3'/S-5/S-7. These PpsS-RNases, which were expressed exclusively in style, shared the typical structural features with S-RNases from other Prunus species exhibiting GSI. All PpsSFBs showed similar structure characteristics of SFBs from other Prunus species, and matched with the necessary conditions for pollen S-determinant. No mutations leading to dysfunction of S-haplotype were found in their full-length c-DNA sequences, except for PpsS-3'-haplotype which was not amplified by PCR. These four S-haplotypes complied with tetrasomic inheritance. Diploid pollen grains with S-genotypes S-7/S-1, S-7/S-5 and S-1/S-5 can grow the full length of the style after self-pollination, while pollen grains with S-3'/S-7, S-3'/S-1 and S-3'/S-5 cannot. These results suggest that PpsS-haplotypes-1, -5 and -7 are functional, and that competitive interaction between two of them confer self-compatibility on cultivar "NC". Furthermore, in terms of recognition specificity, diploid pollen grains carrying PpsS-3'-haplotype are equal to monoploid pollen grains carrying the other functional S-haplotype.