A New One-Tube Reaction Assay for the Universal Determination of Sweet Cherry (Prunus avium L.) Self-(In)Compatible MGST- and S-Alleles Using Capillary Fragment Analysis

The sweet cherry plant (Prunus avium L.) is primarily self-incompatible, with so-called S-alleles responsible for the inability of flowers to be pollinated not only by their own pollen grains but also by pollen from other cherries having the same S-alleles. This characteristic has wide-ranging impacts on commercial growing, harvesting, and breeding. However, mutations in S-alleles as well as changes in the expression of M locus-encoded glutathione-S-transferase (MGST) can lead to complete or partial self-compatibility, simplifying orchard management and reducing possible crop losses. Knowledge of S-alleles is important for growers and breeders, but current determination methods are challenging, requiring several PCR runs. Here we present a system for the identification of multiple S-alleles and MGST promoter variants in one-tube PCR, with subsequent fragment analysis on a capillary genetic analyzer. The assay was shown to unequivocally determine three MGST alleles, 14 self-incompatible S-alleles, and all three known self-compatible S-alleles (S3′, S4′, S5′) in 55 combinations tested, and thus it is especially suitable for routine S-allele diagnostics and molecular marker-assisted breeding for self-compatible sweet cherries. In addition, we identified a previously unknown S-allele in the ’Techlovicka´ genotype (S54) and a new variant of the MGST promoter with an 8-bp deletion in the ´Kronio´ cultivar.