Isolation of European pear S-haplotypes using S-homozygotes derived from bud-selfing

Gametophytic self-incompatibility is a mechanism that prevents self-fertilization through a specific pollen-pistil recognition, genetically controlled by the single S locus. In the Pyrinae the S locus has a quite complex structure including a single female (pistil-expressed) gene, the S-RNase, and a number of male (pollen-expressed) F-box genes that act coordinately in a non-self-recognition system. While each Shaplotype segregates as a single unit, gene duplication and genetic exchange seem to play an essential role in the evolution of pollen S-specificities; as a result, extant Shaplotypes differ for the number, position and arrangement of F-box genes. So far, the genomic characterization of some S-haplotypes has been achieved using genomic libraries; the use of S-homozygotes can be an alternative source of information to characterize the pool of F-box genes belonging to a single haplotype. To overcome the self-incompatibility barrier, selfing can be forced by manually pollinating pistils of immature flowers ("bud-selfing"): As the concentration of S-RNases in the early stages of the pistil development is not yet fully effective, a portion of self-pollen tubes can escape the inhibition. The progeny obtained show a mendelian (1:2:1) segregation for the two parent S haplotypes, allowing the isolation of S-homozygotes in half the seedlings; since a low vigor is expected as a consequence of the increase in homozygosity, these genotypes are maintained and multiplied in vitro. This material will be useful for the identification and characterization of F-box genes in different Shaplotypes, as a first step for investigating their roles in the determination of S specificities as well as their evolutionary dynamics.