Genotyping of Citrus Accessions with S₉ and/or S₁₀ Alleles for Self-incompatibility and Their Allelic Distribution

Gametophytic self-incompatibility, one of the key characteristics for breeding seedless Citrus cultivars, occurs in pummelo (Citrus maxima), mandarin (Citrus reticulata), and their hybrid cultivars. Allelic variation in Citrus was reported for the self-incompatibility gene (S); however, S allele frequencies and S genotypes of full-and semi-self-incompatible cultivars have been reported for a small number of alleles. To extend our knowledge of S alleles, we tested 146 Citrus accessions, including 82 pummelo accessions, for S₉ and S₁₀ alleles. Each accession was pollinated with homozygous S₁ seedlings of ‘Hirado Buntan’ pummelo (S₉S₉ and S₁₀S₁₀). The pollen tube growth arrest in the lower styles of their pollinated pistils indicated that four accessions, including ‘Hirado Buntan [Citrus maxima (Burm.) Merr.]’, have the S₉ allele and five accessions, including ‘Hirado Buntan’, have the S₁₀ allele. The percentage of accessions with the S₉ allele was 3.2% (4 of 126 accessions examined), and the S₉ allele frequency was 1.8% (4 of 217 alleles, excluding the S_f allele). The percentage of accessions with the S₁₀ allele was 3.9% (5 of 127 accessions examined), and the S₁₀ allele frequency was 2.3% (5 of 217 alleles, excluding the S_f allele). Japanese mandarin (another sources of S alleles) and its relatives had neither the S₉ nor the S₁₀ allele. Pummelo accessions had S₉ and S₁₀ alleles at higher rates of 2.9% (2 of 70 accessions examined) and 7.0% (5 of 71 accessions examined), respectively. ‘Kabusu’ sour orange (a pummelo-mandarin hybrid; Citrus aurantium) and ‘Kikudaidai’ (a sour orange relative; Citrus canaliculata) had S₉ alleles. These results suggested that the two alleles originated from pummelo (the main sources of S alleles). The S genotypes with S₉ and/or S₁₀ alleles were fully determined in ‘Hirado Buntan’ (S₉S₁₀), ‘Kabusu’ sour orange (S_fS₉), the ‘Kikudaidai’ (S₉S₁₁) sour orange hybrid, and two local pummelo plants. The results of our study suggest that in comparison with the other S alleles reported, the pummelo plants with low frequencies of S₉ and/or S₁₀ alleles contributed to very low rates of evolution and development of Citrus species and cultivars during the long history of citrus cultivation, except for those generating sour oranges (Citrus auratinum), which are used as root stocks, and for flesh and rind processing and ornamental plants. Finally, we examined the degree of self-incompatibility between S₉ and S₁₀ alleles in the lower part of styles of S₁ seedlings of ‘Hirado Buntan’ and Citrus accessions with S₉ and/or S₁₀ alleles. The result indicated no difference in the self-incompatibility reaction between the two alleles.