Genotyping for male sterility (MS) and MS gene mapping with RAPD markers in Citrus, especially with precocious flowering seedlings from a cross of 'HY16' × grapefruit

Seedlings obtained from crosses with Citrus species and cultivars showed precocious flowering in spring after seed germination in autumn. Male-sterile anthers were observed in the flowers of seedlings from seven crosses. On the basis of segregation analysis for male sterility in these progenies, male-sterile genotype was estimated to be (S)r₁r₁R₂r₂r₃r₃ for male sterile 'HY16', (S)r₁r₁r₂r₂R₃r₃ for male sterile 'Kiyomi' and (F)r₁r₁R₂R₂R₃r₃ or (F)R₁r₁R₂R₂r₃r₃ for male fertile 'Foster Pink' grapefruit. Here, male sterility is a recessive character; r₂ and r₃ are complementary genes with upper stream expression of r₁, and r₁ is an epistatic gene to r₂ and r₃ genes. Male sterile seedlings appear when r₁ is recessive homozygous (r₁r₁) and either r₂ or r₃ is recessive homozygous (r₂r₂ or r₃r₃). A population of 101 individuals from the cross of 'HY16' × 'Foster Pink', in which male sterile and male fertile seedlings segregated in a 1:1 ratio, was chosen for mapping the R gene with RAPD markers. Reproducible polymorphisms of amplified DNA fragments were obtained; 260 RAPD primers were tested by means of bulked segregant analysis. Five RAPD markers amplified in most of male-fertile individuals and 'Foster Pink'. On the other hand, two RAPD markers were found in most of male-sterile individuals and 'Foster Pink'. The linkage analysis showed that the five markers flanked with R₁ or R₃ gene with the closest distance of 8.4 cM in grapefruit and the two markers flanked with r₁ or r₃ gene with the closest distance of 15.5 cM in grapefruit.