Conversion of RFLP markers for the selection of pollination-constant and non-astringent type persimmons (Diospyros kaki Thunb.) into PCR-based markers

Abstract

Persimmon (Diospyros kaki Thunb.) cultivars are classified into 4 types depending on the relationship between astringency of the mature fruit and the effect of seeds on the loss of astringency, and only pollination-constant and non-astringent (PCNA)-type persimmons stably lose fruit astringency as a part of fruit development. This is a recessive trait, regarded to be controlled by a single locus, namely, the AST locus, which has a polysomic nature. Thus far, we have identified 2 restriction fragment length polymorphism (RFLP) markers, namely, Al and A2, each of which is separately linked to a different AST allele, and proved that the RFLP markers were useful for selecting PCNA-type persimmons. This study was conducted to convert the RFLP markers into polymerase chain reaction (PCR)-based markers. We isolated and characterized genomic regions corresponding to each RFLP marker by inverse PCR. Two primer pairs, E4/E9r and E4/A2r, were designed to generate 2 sequence characterized amplified region (SCAR) markers, i.e., PCR-A1 and PCR-A2, respectively. The PCR-A1 and PCR-A2 markers cosegregated with the Al and A2 markers, respectively, in 'Nishimura-wase'-derived progenies. Although the primer pair E4/A2r did not produce the PCR-A2 marker in the FU-275, which is a progeny derived from 'Aizumishirazu', all non-PCNA-type offspring but no PCNA-type offspring showed the PCR-A1 marker using the primer pair E4/E9r. Thus, it was revealed that the SCAR markers were useful for selecting PCNA-type offspring in these progenies. On the other hand, disruption of the relationship between the markers and the AST locus was observed in the KU-325, derived from 'Kurokuma', indicating that the selection of PCNA-type offspring using PCR-based markers is not effective for progeny derived from 'Kurokuma'. Herein, we discuss the possibility of applying PCR-based markers to genetic studies. JSHS © 2009.