DNA sequence diversity of the astringency-controlling locus in Diospyros kaki

Astringency due to proanthocyanidins (PAs) in the mature fruit is a very unique characteristic of persimmons. Pollination-constant and non-astringent (PCNA) type cultivars stop accumulating PAs in the fruit early in fruit development, allowing the fruit to be edible without artificial de-astringency treatment. The PCNA trait is conferred by a recessive allele at a single locus, ASTRINGENCY (AST), and a SCAR marker, which is tightly linked to the PCNA trait, is widely used for marker-assisted selection for the PCNA trait in modern persimmon breeding. In this study, 110 cultivars and strains of Diospyros kaki and a single strain of D. oleifera were evaluated for genetic diversity in an approximately 4.5 kb region around the SCAR marker using amplicon-based Pacbio sequencing. In the PCR marker region, considerable sequence variation was observed in the presumed ast haplotypes, but to a lesser extent than in the presumed AST haplotypes. Haplotype network analysis showed that haplotypes with indel-3, a genetic marker currently used for the marker-assisted selection of PCNA trait, formed a single cluster. These results collectively suggested that ast was generated by a single mutation event from the original AST. In addition, several polymorphisms were detected in the region corresponding to the primer binding sites of the SCAR marker for PCNA selection. The results presented in this study indicate that the current PCNA selection could be improved by designing primers in less diverse regions based on the analysis of the sequence diversity in the AST region.