Many species in the families of Rosaceae, Solanaceae, and Scrophulariaceae exhibit gametophytic self-incompatibility, a phenomenon controlled by two polymorphic genes at the S-locus, style-S (S-RNase) and pollen- S (SFB). Sequences of both genes show high levels of diversity, characteristic of genes involved in recognition of self-incompatibility systems in plants. In this study, S24- RNase and SFB24 alleles were cloned from Prunus armeniaca cv. Chuanzhihong (Chinese apricot). Sequence comparisons of deduced amino acid sequences revealed that the P. armeniaca S24-haplotype has different SFB alleles, but shares a single S-RNase allele with P. armeniaca S4-haplotype. Moreover, P. armeniaca S24-RNase haplotype has a single and three different alleles with S1-RNase of P. tenella (dwarf almond) and S1-RNase of P. mira (smooth pit peach), respectively. The functionalities of SFB24 and SFB4 have been evaluated by pollen tube growth and controlled field tests of P. tenella and P. mira. Genetic analysis of the two intercrosses showed that progenies segregated 1:1 into two S-genotype classes, which is consistent with the expected ratio for semi-compatibility. These findings imply that the allelic function of the S24-haplotype is identical to that of the S4-haplotype in a self-incompatibility reaction. Thus, these two Prunus S-haplotypes are in fact two neutral variants of the same S-haplotype. The evolution of the S-allele is also discussed in terms of both functions and differences between S24- and S4-haplotypes in Prunus.