Construction of high-resolution linkage map of the Ms locus, a restorer-of-fertility gene in onion (Allium cepa L.)

For the purpose of developing closely-linked molecular markers to the Ms locus, a restorer-of-fertility gene in onions (Allium cepa L.), bulked segregant analysis and randomly amplified polymorphic DNA (RAPD) analyses were utilized. Five RAPD markers polymorphic between male-fertile and male-sterile bulks were identified. These RAPD markers were converted into a simple PCR marker or cleaved amplified polymorphic sequence (CAPS) markers after sequencing the RAPD products and obtaining flanking sequences of the RAPD markers by genome walking. A linkage map was constructed with the Ms locus and flanking markers using a F-2 population. There was no recombinant between the Ms locus and two CAPS markers, jnurf05 and jnurf17. To increase resolution among these closely linked molecular markers and the Ms locus, a total of 1,346 F-2:3 and 2,927 F-2:4 plants were analyzed with two flanking markers for detection of recombinants. Segregation of male-fertility phenotypes in large-sized populations confirmed allelic segregation distortion in favor of the recessive Ms allele. Analysis of the recombinants with closely linked markers revealed only two recombinants between the Ms locus and the jnurf05 markers among 4,273 segregating plants, showing very tight linkage between the two loci. However, linkage disequilibrium between the two loci was not too strong among the breeding lines. Despite weak linkage disequilibrium, these tightly linked markers are useful in accurate marker-assisted selection of the Ms alleles and ultimate isolation of the Ms gene by map-based cloning approach.