Development of a system for S locus haplotyping based on the polymorphic SLL2 gene tightly linked to the locus determining self-incompatibility in radish (Raphanus sativus L.)

Abstract

To develop a reliable system for identifying multiple S haplotypes controlling self-incompatibility (SI) in radish (Raphanus sativus L.), the genomic organization of the S locus region was identified from radish draft genome sequences. An initial attempt to find the S receptor kinase (SRK) gene, the female determinant of SI, failed due to presence of 15 homologous genes. Using synteny between the radish and Chinese cabbage genomes, the putative S locus region was identified in the radish R7 linkage group. One scaffold anchored to this R7 region contained the S-locus glycoprotein (SLG) gene, which is one of the S locus genes. Using the high homology between the SLG and S domain of SRK, the full-length radish SRK gene containing the largest 6861-bp intron1 was assembled by connecting two scaffolds harboring the S receptor and kinase domains, respectively. A scaffold containing the full-length S-locus cysteine-rich protein (SCR)/S locus protein 11 (SP11) gene, the male-determinant of SI, was identified using information reported previously. Finally, 53,785, 42,804, and 10,165 bp sequences containing the S locus genes and their flanking sequences were obtained. Unlike the various orientations of the SRK or SCR/SP11 genes, the position of SLL2 located at the border region of the S locus was conserved among haplotypes. Sequencing of the SLL2 gene from 31 inbred lines showing differential SI responses revealed 26 polymorphic alleles. Four additional SLL2 alleles were identified from analysis of diverse breeding lines. Based on the polymorphic SLL2 sequences, a new S haplotyping system was developed for efficient marker-assisted selection of the S haplotypes in radish.