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Akihiko Sato and Masahiko Yamada

The textural properties of grape berry flesh were evaluated with a puncture test using a total of 87 cultivars, consisting of 62 vinifera cultivars (Vitis vinifera L.) and 25 labruscana cultivars (Vitis labruscana Bailey) in order to search cultivars with crisp flesh texture for table grape breeding. Crisp texture is the most desirable texture for tableuse, and was determined as easy breakdown on mastication and firm flesh, which was measured as small deformation at the first major peak (DFP) and large maximum force (MF), respectively, in force-deformation curve of the test. For the vinifera cultivars, the mean values of DFP and MF were significantly smaller and larger in table-use cultivars than wine-use and dual-purpose cultivars, respectively. The result indicated that the vinifera wine-use and the dual-purpose cultivars had soft and non-crisp flesh. Based on the puncture tests, the cultivars with a crisp texture (2.5 mm ≥ for DFP and 0.9 N ≤ for MF) were limited to 11 vinifera table-use cultivars such as `Baladi', `Cardinal', and `Muscat of Alexandria'; and one vinifera dual-purpose cultivar (`Terbash'). The DFP was generally high in labruscana cultivars, which include table-use and dual-purpose cultivars, resulting in lacking cultivars with crisp texture among labruscana cultivars. These results indicate that the cultivars with a crisp texture were restricted to a small source within vinifera cultivars.

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Masahiko Yamada, Akihiko Sato and Yasuo Ukai

Environmental variance components were estimated for calyx-end fruit cracking in pollination-constant and nonastringent cultivars and selections of Japanese persimmon (Diospyros kaki Thunb.). The cracking value of a tree in a cultivar or selection (genotype) (X) was evaluated as the number of fruit that cracked divided by the total number (25) of fruit evaluated from each tree. Because the mean value of X was correlated with the variance of X, analyses of variance were performed using its square root value. The variance associated with genotyp× year interaction was the largest of environmental variance components. The variances associated among years and among trees within genotypes were very small. The mean percentage of cracked fruit in evaluation for 10 years was 3% for `Fuyu', 11% for `Matsumotowase-Fuyu', and 12% for `Izu'. On the basis of the environmental variance components obtained, it is proposed that all offspring genotypes exhibiting a phenotypic cracking incidence of less than 20% and 11% should be selected in single-year and three-year evaluations, respectively, when those genotypes are evaluated using 25 fruits from a single tree, in order to successfully select all genotypes with an genotypic incidence of less than 3%.

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Akihiko Sato, Masahiko Yamada and Hiroshi Iwanami

Berry texture of grapes (Vitis labruscana Bailey, V. vinifera L., and their hybrids) can be characterized by two factors: 1) difficulty of breakdown in mastication and 2) firmness, which can be mechanically measured as deformation at first major peak (DFP) and maximum force (MF) of the force-deformation curve in flesh puncture tests. Crisp texture (easily breakable and firm flesh texture), one of the most important factors for the quality of table grapes, corresponds to a combination of small DFP (≤2.5 mm) and large MF (≥0.9 N). Obtaining offspring with crispy flesh is a primary objective in grape interspecific hybrid breeding at the National Institute of Fruit Tree Science, Japan. In this study, the expected proportion (EP) of offspring with crispy flesh as a genotypic value was estimated using a population consisting of 23 full-sib families each with eight offspring. An analysis of variance in the offspring, which estimated between-family and within-family variances, and the regression analysis of the family mean (Fm) of eight offspring in each full-sib family on mid-parental value (MP) were conducted for DFP and MF. The results revealed that the total genetic variation in offspring was mostly explained by the variance due to the regression and the within-family variance for both DFP and MF. No significant heterogeneity of within-family variance was detected by Bartlett's test for either DFP or MF. Therefore, a simple model was used to calculate EP: Fm is solely determined by the regression of Fm on MP, and all families have an equal within-family genetic variance due to segregation. Since merely a weak correlation relationship existed between DFP and MF, its influence was omitted in calculating EP. The EP of offspring having crisp texture was estimated to be 11% for an MP value of DFP of 2.5 mm (DFP for `Italia'), 6% for a DFP of 3.5 mm (DFP for `Athens'), and 3% for a DFP of 4.5 mm (DFP for `Bath'); the MP value of MF was assumed to be 0.7 N (MF for `Steuben' and `Italia') in this calculation.

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Ayako Ikegami, Keizo Yonemori, Akira Sugiura, Akihiko Sato and Masahiko Yamada

Japanese persimmon (Diospyros kaki Thunb.) cultivars are classified into four types depending on the nature of astringency loss of the fruit. The pollination-constant, non-astringent (PCNA) persimmons lose their astringency on the tree as the fruits develop. This PCNA trait is qualitatively inherited and recessive to the other three types, pollination-constant, astringent (PCA), pollination-variant, nonastringent (PVNA), and pollination-variant, astringent (PVA). In fact, crosses among Japanese PCNA cultivars yield only PCNA type in F1 generation as shown in recent breeding programs at the National Institute of Fruit Tree Science. Despite these previous results, we demonstrated here that non-PCNA (PVNA, PVA, and PCA) type offspring were derived at relatively high rates in the F1 generation from a cross between `Luo Tian Tian Shi', a PCNA accession from China, and the Japanese PCNA cultivar, `Taishu', despite the fact that `Luo Tian Tian Shi' was confirmed to be a true PCNA type by measuring tannin cell size, a principal morphological characteristic to distinguish PCNA cultivars from non-PCNA ones. When segregations of tannin cell size and tannin content in three progenies of the breeding populations derived from Chinese PCNA `Luo Tian Tian Shi' × Japanese PCNA `Taishu', Japanese PCNA `Shinshu' × Japanese PCNA `Taishu', and Japanese PVNA (non-PCNA) `Kurokuma' × Japanese PCNA `Taishu' were investigated, all offspring between Japanese PCNA cultivars contained only small tannin cells and were PCNA types, and those between Japanese PVNA × PCNA cultivars contained only large tannin cells and were non-PCNA types. However, hybrids between `Luo Tian Tian Shi' and `Taishu' segregated into populations of small and large tannin cells, indicating that `Luo Tian Tian Shi' is likely heterozygous for astringency. Therefore, Chinese PCNA `Luo Tian Tian Shi' should be different from Japanese PCNA cultivars in genetic makeup.

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Atsushi Kono, Akihiko Sato, Yusuke Ban and Nobuhito Mitani

We evaluated the resistance of 133 grapevine cultivars or selections, including Vitis vinifera and American hybrids, on the basis of lesion number and length to identify sources of resistance to grapevine anthracnose. All germplasms tested in this study showed anthracnose symptoms to some extent, and the distribution of lesion number and diameter was continuous. Most table grape V. vinifera cultivars were highly susceptible, showing many large lesions. However, V. vinifera wine grapes were more resistant with smaller lesions. Some American hybrid grapes such as ‘Ontario’ showed very few and small lesions. There was a significant positive correlation between lesion number and size in American (r = 0.63, P = 0.0041) and Japanese hybrids (r = 0.56, P < 0.001), whereas there was no correlation between these characters in V. vinifera. Japanese tetraploid cultivars were neither highly susceptible nor resistant. High anthracnose susceptibility of most well-known table grape V. vinifera cultivars, including ‘Muscat of Alexandria’, ‘Italia’, ‘Rizamat’, ‘Kattakurgan’, and ‘Thompson Seedless’, indicates that resistance should be introgressed from other cultivars such as American hybrids or wine grapes when these susceptible table grapes or their descendants are used in breeding anthracnose-resistant table grapes.

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Atsushi Kono, Akihiko Sato, Bruce Reisch and Lance Cadle-Davidson

Grapevine downy mildew (DM), caused by the oomycete Plasmopara viticola (Berk. & Curt.) Berlese & de Toni, is a major disease, especially in humid viticultural areas. Development of resistant cultivars is an important objective for grapevine breeding. To establish a reliable and inexpensive quantitative method to aid in breeding for DM resistance, we improved the method for counting the number of sporangia on leaf discs, and developed a method for counting the number of sporangia in solution. To prevent the loss of DM sporangia from adhesion onto plastic ware, we found as little as 0.01% Tween 20 was effective. On the other hand, this detergent was shown to have a severe inhibitory effect upon DM infection of leaves. We developed a sporangia counting method using dried droplets of DM suspensions, and the method was highly correlated with counting by hemacytometer (R 2 > 0.96). The nonparametric Spearman’s rank correlations between visual rating and the number of the sporangia were as high as ρ = 0.82 to 0.91, suggesting that evaluation by the visual rating could provide a good estimate of the sporangia numbers on leaf discs. We established a high-throughput and inexpensive method with acceptable accuracy for DM resistance evaluation based on a leaf disc assay, and our results suggested that visual ratings of infected leaf discs provide a good estimate of sporangia numbers.

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Hiroshi Iwanami, Nobuyuki Hirakawa, Hiroyasu Yamane and Akihiko Sato

Crosses between seedless cultivars had been conducted to produce seedless table grape efficiently by combining with ovule and embryo culture in Vitis vinifera L. But very few plants grew normally in this method. Four plant growth regulators (Cycocel, B-Nine, Uniconazole-P, Ethrel) were applied to shoots 4 weeks before anthesis to develop the seeds of two seedless cultivars `Flame seedless' and `A1706'. Correlation was significant in each cultivar between the shoot length at anthesis and the number of seed traces per berry in all combined treatments. Analysis of covariance revealed that the number of seed traces per berry was significantly higher when the shoots were applied with Uniconazole-P (240 ppm) than B-nine (2000 ppm), Cycocel (500 ppm) and Ethrel (400 ppm) in `Flame seedless' and Uniconazole-P and B-nine than Ethrel in `A1706'. Ovules of these two seedless cultivars crossed with seedless cultivar `Perlette' after the application of four plant growth regulators were cultured on half-strength MS medium with 10 μm IAA and the percentage of developed embryos in ovules was higher when the shoots were applied with Uniconazole-P and B-nine than Cycocel, Ethrel in `Flame seedless' and B-nine than others in `A1706'. These results indicate that the use of certain plant growth regulators promotes the embryo development.

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Ayako Ikegami, Keizo Yonemori, Akira Kitajima, Akihiko Sato and Masahiko Yamada

Expression patterns of the genes involved in condensed tannin (CT) biosynthesis during fruit development was investigated in a Chinese pollination-constant, nonastringent (PCNA) persimmon (Diospyros kaki Thunb.) `Luo Tian Tian Shi'. The transcript levels of phenylalanine ammonia-lyase (PAL) and dihydroflavonol reductase (DFR) in `Luo Tian Tian Shi' were detected at high levels throughout the fruit growth. Chalcone synthase (CHS) and flavonol 3-hydroxylase (F3H) also continued to be transcribed during fruit growth, although their levels decreased earlier than PAL and DFR. In contrast, expression levels of these genes declined into undetectable levels at an early stage of fruit development in Japanese PCNA persimmon. In addition, anthocyanidin reductase (ANR), which encodes a key enzyme of the proanthocyanidin biosynthesis, was transcribed at high levels in `Luo Tian Tian Shi' during fruit growth, but not in Japanese PCNA persimmon. By contrast, the expression of D. kaki serine carboxypeptidase-like protein 1 (DkSCPL1) that was obtained from suppression subtractive hybridization (SSH) analysis between artificially astringency-removed fruit and astringent fruit in a different experiment, declined earlier than the other flavonoid biosynthesis genes in `Luo Tian Tian Shi', coincident with the termination of the tannin cell development. In the F1 progeny of the cross between `Luo Tian Tian Shi' and Japanese PCNA `Taishu', similar expression patterns were obtained among segregated PCNA and astringent offspring. These results indicate that Chinese PCNA is different from Japanese PCNA in expression of the genes involved in CT biosynthesis. In conclusion, we clarified that expression of the genes (PAL to ANR, but not SCPL) involved in flavonoid biosynthesis was continuous in the Chinese PCNA cultivar, despite the termination of tannin cell development.

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Shinya Kanzaki, Keizo Yonemori, Akira Sugiura, Akihiko Sato and Masahiko Yamada

Japanese persimmon (Diospyros kaki Thunb.) cultivars are classified into four types depending upon the nature of astringency loss of the fruit. Among them, the pollination-constant and nonastringent (PCNA) type is the most desirable for fresh fruit consumption due to the trait of stable loss of astringency on the tree with fruit development. Lack of tannin accumulation is the main cause of natural astringency loss in PCNA-type fruit, and is qualitatively inherited. The PCNA trait is recessive to the non-PCNA trait. In this study, we investigated amplified fragment length polymorphism (AFLP) markers for the trait of natural astringency loss of PCNA-type fruit using bulked segregant analysis (BSA) for efficient selection of PCNA type plants in a breeding population. A total of 128 primer combinations were tested and one AFLP marker was found to be linked to the dominant allele controlling the trait for astringency. This marker, EACC/MCTA-400, was absent in all of the PCNA-type plants tested, whereas it was present in about half of the non-PCNA-type plants tested. However, RFLP analysis using this marker enabled the detection of the other dominant allele, and all PCNA-type plants could be distinguished from the non-PCNA-type plants. Application of this marker system will be useful for the selection of PCNA-type plants in persimmon breeding.

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Ayako Ikegami, Sai Eguchi, Keizo Yonemori, Masahiko Yamada, Akihiko Sato, Nobuhito Mitani and Akira Kitajima

Pollination-constant and nonastringent (PCNA) is one of the most desirable traits in persimmons as this type of cultivar loses its astringency while still on the tree before harvest. Among Japanese PCNA cultivars, the trait is qualitatively inherited and recessive to pollination-constant, astringent (PCA), pollination-variant, nonastringent (PVNA), and pollination-variant, astringent (PVA) types. However, in a previous trial, both astringent and nonastringent types segregated in the F1 population that resulted from a cross between a Chinese PCNA `Luo Tian Tian Shi' and a Japanese PCNA cultivar. Because of the unusual segregation, in this study, we crossed another Japanese PCNA `Okugosho' with `Luo Tian Tian Shi' to confirm the segregation of astringent types by measuring the tannin cell size and tannin concentration at harvest. Previously, we found that astringent types have larger tannin cells than PCNA-type. The F1 hybrid progenies from the cross segregated into both PCNA and astringent-type individuals in approximately 1:1 ratio. Likewise, the F1 population from the astringent-type `Yotsumizo' and `Iwasedo' × `Luo Tian Tian Shi' were ascertained to contain both PCNA and astringent types, which indicates that the PCNA trait of `Luo Tian Tian Shi' was dominant. Thus, this Chinese cultivar has the potential to become an important parental material for future breeding of PCNA persimmons.