Eight-year-old peach trees (Prunus persica (L.) Batsch cv. ‘Hakuho’ grafted on ‘Ohatsumomo’ peach rootstock), planted in 1500 L beds in a plastic greenhouse to avoid rain and groundwater effects, were used to evaluate the relationships among sap flow (SF), water-deficit stress parameters, and atmospheric conditions. Under a mild water-deficit stress condition, sap-flow velocity (SFV) decreased in the afternoon but not in the morning. Two indices were derived from the daily SFV profile: i) the slope of the regression line of SFV in the daytime (index C) and ii) the decreasing rate of SFV from the morning to each hour in the afternoon (index D). These indices showed higher correlation coefficients to the tree water status expressed by predawn leaf water potential (Ψ_PD) (r² = 0.41) and maximum daily shrinkage of the trunk (MDS) (r² = 0.58**) than to daily total SF (index A) and SFV at each hour in the daytime (index B). In a nonstressed condition, SFV was more highly correlated to photosynthetic photon flux density (PPFD) (r² = 0.84**) than to temperature, relative humidity, and vapor pressure deficit (VPD). The correlation coefficients (r²) of indices with MDS and Ψ_PD increased to 0.77** and 0.92**, respectively, when the SFVs of low light intensity (< 800 μmol·m⁻²·s⁻¹ of PPFD) were eliminated. The index which most precisely detected water-deficit stress was the decreasing rate of SFV from the morning (07:00–11:00 h) to any hour from 12:00 to 15:00 h with the elimination of SFV values when PPFDs were low.

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Assessment of the tree vigor of grafted rootstocks in citrus requires intense labor, a long period, and large fields because tree vigor is judged through cultivation tests with adult trees. We have developed a method to predict tree vigor at an early stage based on the stem hydraulic conductance of 5-month-old seedlings. We used 15 cultivars (strains), including 11 crossed strains, 3 cultivars of commonly used trifoliate orange (common type, ‘Pomeroy’, and ‘USDA’), and ‘Swingle citrumelo’. The growth characteristics of 7-year-old satsuma mandarin trees grafted onto those rootstocks were compared with those of 5-month-old rootstock seedlings. The results revealed a relatively high correlation (r² = 0.633) between the trunk circumference of 7-year-old trees and the stem hydraulic conductance of seedlings, and furthermore, control cultivars existed approximately on that regression line. Therefore, it seems possible to predict tree vigor from the stem hydraulic conductance of seedlings in a very short period without using a large field and intense labor.

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Fruit trees have a long juvenile period after germination before flowering and fruit set. In spite of its horticultural importance, the physiological and molecular mechanisms of juvenility in fruit tree development are unknown; therefore, the relationship between the expression of the AGAMOUS-LIKE6 (AGL6) homolog and juvenility was investigated in apple (Malus domestica Borkh.). The results of sequence alignment, the phylogenetic tree, and Southern blot analysis suggested that MdMADS11 is an AGL6 ortholog in apple. An increase in the abundance of the mRNA of MdMADS11, which promotes flowering, corresponded to changes in leaf area and serration with the transition from the juvenile to adult phase in apple seedlings. In contrast, the abundance of mRNA of MdJOINTLESS, which may suppress flowering, decreased with the phase transition. Expression analysis of adult apple trees suggested that MdMADS11 also plays a role in seasonal flower initiation and floral organ formation in the adult phase. The results provide new insight into the molecular mechanism of the phase transition between juvenile and adult fruit trees.

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Gametophytic self-incompatibility (SI), an important character for breeding seedless cultivars of Citrus, is known in pummelo, mandarin, and many hybrid cultivars with pummelo in their pedigrees. Only a little is known about the allelic variation in the self-incompatibility gene (S), S allele frequencies, and the genotypes of self-incompatible and semi-self-compatible cultivars. In this study, pollination of ‘Banpeiyu’ S₁ seedlings with ‘Banpeiyu’ and pollination between the S₁ seedlings were performed to determine homozygous S₁ seedlings for the S gene. Seventy-eight Citrus accessions, including 55 pummelo accessions, were pollinated with each of two homozygous S₁ seedlings (S₁S₁ and S₂S₂). Pollen tube arrest in the style base of their pollinated pistils indicated that 23 accessions, including ‘Banpeiyu’, have an S₁ allele each and 16 accessions, including ‘Banpeiyu’, have an S₂ allele each. Frequency of accessions with S₁ allele was 29.9% (23 of 77 accessions examined) and S₁ allele frequency was 16.4% (23 of 140 alleles excluding S_f allele). Frequency of accessions with S₂ allele was 21.3% (16 of 75 accessions examined) and S₂ allele frequency was 11.6% (16 of 138 alleles excluding S_f allele). Pummelo accessions collected from Kagoshima Prefecture had S₁ alleles with two and half times higher frequency (56.3%) than that in all accessions examined. Of the 79 accessions, six accessions (‘Banpeiyu’, ‘Iriki Buntan’, ‘Kaopang’, Nagashima Buntan No. 6, Nagashima Buntan No. 7, and ‘Soyu’) were S₁S₂ genotypes. The Citrus cultivars, whose S genotypes have been fully determined in this study, were ‘Banpeiyu’ (S₁S₂), ‘Iriki Buntan’ (S₁S₂), ‘Kaopang’ (S₁S₂), ‘Soyu’ (S₁S₂), ‘Kinukawa’ (S_fS₂), and ‘Kawano Natsudaidai’ (S_fS₂).

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Fusarium wilt of melon (Cucumis melo L.), caused by Fusarium oxysporum f. sp. melonis, is regarded as a severe disease worldwide. Many resources that are resistant to races 0, 1, and 2 have been reported; nevertheless, no resource of high resistance to either race 1,2w or 1,2y has been reported, although partial resistance has been found. Wild Cucumis species have been reported as genetic resources which have resistance to some diseases and pests of melon. This study identified novel sources of high resistance to race 1,2y in wild Cucumis species. In all, we tested 76 accessions belonging to 11 wild species of the genus Cucumis. After artificial inoculation, the disease severity in each plant was evaluated using a 0–3 disease severity scale (0 = no symptoms, 1 = beginning of yellowing symptom on leaves, 2 = leaves strongly affected, 3 = plant death). The disease index (DI) was calculated as DI = summation of (disease severity × number of plants in that disease severity) × 100/(3 × total number of plants). Accessions showing high resistance (DI = 0) were 34 among six species: C. africanus, C. anguria, C. metuliferus, C. prophetarum, C. subsericeus, and C. zeyheri. By contrast, most accessions in C. dipsaceus, C. meeusei, C. pustulatus, and C. sagittatus exhibited high susceptibility (DI = 80–100). Among these four species, no accession showed high resistance. This is the first report of genetic resources having high resistance to race 1,2y. To introduce this high resistance of race 1,2y to melon, we should investigate methods to overcome reproductive barriers to interspecific crosses between melon and wild Cucumis species.

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To differentiate the fruit texture characteristics of five major cucumber types (Beit Alpha, greenhouse, Japanese, pickling, and slicer) that are grown worldwide and to identify beneficial traits for cucumber breeding, we compared their physical properties [flesh firmness, placental firmness, skin toughness, crispness index, ratio of placental firmness to flesh firmness (relative placental firmness), and ratio of skin toughness to flesh firmness (relative skin toughness)]. We also compared the time-dependent changes in the physical properties of each type. Although differences existed in the standard size among the various types, physical properties clearly differed among the types and can be summarized as follows: the Japanese type has firmer flesh and placenta, lower relative skin toughness, and the highest crispness; Beit Alpha and greenhouse types have softer flesh, placenta, and skin, and the lowest crispness; the pickling type has firmer (tougher) flesh, placenta, and skin; and the slicer type has softer flesh and placenta, and the toughest skin. The physical properties of sequentially harvested fruits in each cultivar varied widely, indicating that fruit texture is strongly affected by cultivar (or type) and fruit size (i.e., timing of harvest). It is suggested that much attention should be paid to cucumber texture to breed favorable cultivars.

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We evaluated 41 Capsicum accessions collected in Myanmar for resistance to bacterial wilt and Phytophthora blight. Eighteen accessions were highly resistant to bacterial wilt. They consisted of 11 C. annuum, 6 C. frutescens, and 1 C. chinense, and had a wide range of fruit sizes and shapes. Seventeen were moderately resistant. On the other hand, only 1 accession was highly resistant to Phytophthora blight, and all other accessions were susceptible. These results indicate the existence of a range of materials in Myanmar that are highly resistant to bacterial wilt, but few accessions that are resistant to Phytophthora blight. The accession that was highly resistant to Phytophthora blight was also classified as moderately resistant to bacterial wilt, and will therefore be a useful genetic resource in breeding to produce combined resistance to both diseases.

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We studied the genomic sequence of the anthocyanidin synthase (ANS) gene in acyanic flower lines of lisianthus and detected a 94-bp deletion of the sequence in comparison with that of cyanic flower lines. Cross-pollination was made between acyanic and cyanic flower lines to yield F₁ progeny, followed by its self-pollination. The segregated individuals between cyanic and acyanic flowers among the F₂ population were of 24 and 8, respectively (3 : 1), which suggested that one gene of the F₁ hybrid was unequivocally involved in anthocyanin production. The homozygous ANS genes that have a set of 94-bp deletion co-segregated a recessive phenotype of white color in the F₂ population. Genomic PCR of the ANS genes of the 14 independent lisianthus lines of acyanic flowers, such as white, yellow, pale yellow, and pale green flowers, revealed that all possessed a homozygous 94-bp deletion. The data support that the acyanic flower colors are associated with a 94-bp deletion in the ANS gene as a result of the loss of anthocyanin accumulation.

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The intron structures of two variants of 1-aminocyclopropane-1-carboxylate synthase (ACS) genes (DcACS1a and DcACS1b) in carnation (Dianthus caryophyllus) and genes homologous to them (ACS1 homologous genes) in other 10 Dianthus species (16 strains in total) were studied by comparing the sizes of the PCR amplificates and nucleotide sequence of the introns. All 16 sequenced homologous ACS1 genes, including DcACS1 genes themselves, had five exons and four introns. The exons had similar nucleotide sequences and consequently similar deduced amino-acid sequences. The sizes of three introns (intron-1, -2, -3) were variable among the homologous genes, whereas that of the fourth intron (intron-4) was almost identical. The variation in introns was probably caused by the insertion (and deletion) of nucleotide fragments of given lengths. Interestingly, the 3'-UTR of DcACS1a was different from that of DcACS1b, and the latter was similar to other 14 ACS1 homologous genes. Moreover, the length of Thr repeat in the C-terminal region was long in DcACS1a protein but short in DcACS1b protein, and the latter resembled ACS1 homologous proteins in other Dianthus species. The present findings suggest that (1) the variation in intron structure between two variants of carnation DcACS1 is reminiscent of the variation that occurred universally in Dianthus species, (2) DcACS1a is probably a gene intrinsic to carnation, and (3) DcACS1b was acquired from another, as yet unknown, Dianthus species, in the course of breeding modern carnation cultivars.

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Anthocyanin pigments and their related compounds were examined in the flowers of F₁ and F₂ plants generated by crosses between two acyanic lines, 54Y having a speckled mutation for pale yellow flowers and 78WWc-1 carrying the c-1 mutation conferring white flowers, in Japanese morning glory. The mutable speckled allele generates speckled flowers when a dominant genetic element, speckled-activator, is present in its genome. The Speckled and C-1 loci are tightly linked, and 78WWc-1, but not 54Y, bears the specked-activator. The flower color of F₁ progeny is red-purple, and F₂ population displays a ratio of 8 red-purple flowers: 4 white flowers: 3 speckled flowers with red-purple spots pale-yellow background: 1 pale-yellow flower. The anthocyanin components of red-purple flowers in F₁ and F₂ plants as well as red-purple spots of the speckled flower in the F₂ generation were identical. Wedding Bells Anthocyanin (WBA) was accumulated in red-purple flowers as the dominant anthocyanin along with its precursor, pelargonidin 3-sophoroside-5-glucoside, and eight of its acylated derivatives as minor anthocyanins. The major flavonoid pigment in the pale-yellow flowers of 54Y and the F₂ plants as well as the pale-yellow background of the speckled flower in the F₂ generation was identified as chalcononaringenin 2'-glucoside by chemical and spectroscopic methods. Furthermore, chlorogenic acid and small amounts of caffeic acid as well as aureusidin 4-glucoside were obtained from the pale-yellow flowers. Chlorogenic acid and caffeic acid were also detected in the white flowers of F₂ and 78WWc-1. These results suggest that anthocyanin biosynthesis in speckled and c1 mutants stopped at the steps mediated by chalcone isomerase (CHI) and chalcone synthase (CHS), respectively, and that anthocyanin biosynthesis was completely recovered in the red-purple tissue parts of the speckled flower. Moreover, we found that anthocyanin composition varied, especially in the red-purple flowers and red-purple tissue parts in the speckled flowers of F₂ plants, implying that WBA biosynthesis depends on unknown genetic backgrounds.

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Vigorous shoots in chrysanthemums (Chrysanthemum morifolium) often develop from shoot tips (apical meristems with two leaf primordia) cultured in vitro following particle bombardment. The average fresh weight of regenerated shoots from a bombarded shoot tip of chrysanthemum ‘Jinba’ was 10 times more than that regenerated from unbombarded shoot tips. The average number of leaves per bombarded shoot tip was also more than that from an unbombarded shoot tip. The average number of leaves developing from a shoot tip increased with an increase in the amount of gold particles shot into the shoot tips. In addition, when the area destroyed in a shoot apical meristem (SAM) was varied by bombardment through nylon mesh with different pore sizes, the total number of leaves produced from each shoot tip increased with the size of the destroyed SAM area. Knowing the origin of these vigorous shoots, which may be from the bombarded meristem or from the lateral meristems, is important for the screening of transgenic plants. When the entire surface of a SAM was destroyed by bombardment, it was unable to rebuild itself; instead, lateral meristems were initiated at the base of the leaf primordia. Furthermore, the initiation of lateral meristems at the base of the leaf primordia was also observed in instances of restoration of the area of a partially destroyed SAM. This result indicates that vigorous lateral shoots initiate and develop from the bases of leaf primordia when SAMs are damaged to varying degrees. When leaf primordia-free shoot apical meristems (LP-free SAMs) were cultured after bombardment, vigorous shoots failed to develop from the wounded SAM; instead, the wounded LP-free SAMs regenerated a SAM by repairing the wounded areas and developed a non-vigorous single shoot. It was concluded the vigorous shoots do not participate in transgenic plant production because vigorous shoots arise from unbombarded lateral meristems. Finally, an effective and versatile method for transgenic plant production was established by combining micro-wound treatment on a SAM by bombardment and LP-free SAM culture to suppress the growth of vigorous lateral shoots after wounding.

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Lilium brownii var. colchesteri has unique and ornamental floral characteristics in graceful harmony with flower and anther colors, flower shape, elegant fragrance, and flower color change from yellowish cream to white during anthesis. There are, however, few accessions conserved in Japan up to the present, and they often show abnormally shaped flowers and mosaic leaves seemingly due to virus infection. Virus-free bulblets were established by combining meristematic tip culture and chemotherapy. At initial diagnosis with RT-PCR, Cucumber mosaic virus (CMV), Lily mottle virus (LMoV) and Lily symptomless virus (LSV) were detected from leaf tissues of the mother plants. All regenerated bulblets obtained by meristematic tip culture of the mother plants were still infected by at least one of the viruses. The meristematic tip of the bulblets infected with either LSV or LMoV was selected for subsequent culture with 2,4-dioxohexahydro-1,3,5-triazine (DHT), an antiviral chemical. LSV was eliminated successfully in mericlones from bulblets with LSV, whereas LMoV was not from those with LMoV. The virus-free bulblets were transferred to new medium without DHT, and multiplied by in vitro scaling. They were then acclimated in a phytotron glass room at 20°C. The plants were confirmed to be virus-free after 18-months’ acclimation. It was concluded that the combination of meristem tip culture and chemotherapy is practical for producing virus-free plants of L. brownii var. colchesteri.

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Japanese cultivated gentians (Gentiana triflora, G. scabra and their hybrids) are economically important floricultural plants and their use is growing annually worldwide. We isolated 48 simple sequence repeat (SSR) markers using a SSR-enriched genomic library constructed from a gentian double haploid (DH) line. We demonstrated the utility of these SSR markers as a molecular tool for identification of vegetative-propagated gentian cultivars that are difficult to distinguish from their morphology. Five SSR loci showed high genetic polymorphism among 12 Japanese gentian cultivars, ranging from five to eight alleles per locus, with expected heterozygosity between 0.656 and 0.826, and enabled simple and precise identification of cultivars. It was confirmed that the markers were suitable to estimate genetic similarity among cultivars and were also applicable to ten related Gentiana species. This is the first report of the development of a comprehensive set of SSR markers in Japanese cultivated gentians to facilitate genomic analyses in Gentiana, including species of economic importance.

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The vase life of cut flowers is one of the most important breeding targets in ornamentals; therefore, we started a research breeding program in 1992 to improve the vase life of carnation (Dianthus caryophyllus L.) flowers by means of conventional breeding techniques. Crossings and selections of six generations were effective to improve vase life. The mean vase life was 7.4 days for the initial parental generation derived from crossing six parental cultivars; in contrast, after six cycles of crossing and selection, it was improved to 15.9 days, resulting in a net increase of 8.5 days. Moreover, the significant increase of 0.9 days in mean vase life was observed from the fifth to sixth generations, indicating that the breeding progress of flower vase life was continuing in the sixth generation. We selected two ethylene-resistant lines (203-42S and 204-41S) in the fifth generation. The response time to 10 μL·L⁻¹ ethylene was 23.0 h for line 203-42S and 34.2 h for 204-41S, whereas that of a control cultivar, ‘White Sim’ was 7.6 h. The mean vase life of the 18 selected sixth-generation lines ranged from 15.5 to 32.7 days (between 258% and 536% the value of ‘White Sim’). In particular, line 532-6 with an ultra-long vase life showed the longest vase life among all cultivars and lines; the mean vase life of line 532-6 was 32.7 days in 2007 and 27.8 days in 2008 (536% and 463% the value of ‘White Sim’, respectively) at 23°C and 70% RH under a 12-h photoperiod, without chemical treatment. Closer observation of petals during senescence showed that line 532-6 was characterized by a lack of brownish discoloration of petals, which was a senescence symptom of other selected lines with low ethylene production, when the flower lost its ornamental value.

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This study presents a profile of glucosinolate (GSL) levels in the edible portions of 31 cultivars of cabbage cultivated in Japan, and compares potential bioactivity levels on the basis of their ability to induce quinone reductase (QR), a phase II detoxification enzyme. Cabbage (Brassica oleracea var. capitata) seeds were sown in early March 2005 (summer-harvest) and early August 2005 (winter-harvest) and grown under uniform culture conditions. Total GSL concentration varied from 1.07 to 12.14 (mean 5.73) μmol·gDW⁻¹ in the summer-harvest, and from 3.42 to 10.26 (mean 6.16) μmol·gDW⁻¹ in the winter-harvest. The most predominant GSL was 3-indolylmethyl glucosinolate, 2-propenyl glucosinolate or 2-hydroxy-3-butenyl glucosinolate, depending on the cultivar. The values of relative QR activity were positively correlated only with 4-methylsulfinylbutyl glucosinolate (glucoraphanin, GR) content in both the summer-harvest (r = 0.51) and winter-harvest (r = 0.77). GR concentration ranged from 0.04 to 1.70 (mean 0.43) μmol·gDW⁻¹ in the summer-harvest, and from 0.11 to 1.39 (mean 0.44) μmol·gDW⁻¹ in the winter-harvest. There was no significant correlation between GR content and total GSL in either the summer-harvest (r = 0.31) or winter-harvest (r = 0.24). Among the same cultivars, there was a significant positive correlation between GR content in the summer-harvest and that in the winter-harvest (r = 0.63). These results suggest that the inducing activity of QR increased with GR content independent of total GSL, and that there are some inherent differences between cabbage cultivars in the intensity of this activity.

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A technique to determine the geographic origin of wine was examined using 99 samples from Japan and 4 other countries. The contents of 21 elements (Al, B, Ba, Ca, Cd, Cr, Cs, Cu, Fe, K, Li, Mg, Mn, Mo, Na, Ni, P, Pb, Sr, V, and Zn) were determined by inductively coupled plasma optical emission spectrometry. A linear discriminate analysis (LDA) model was created to identify wine from Japan and other countries. With these parameters, 99% correct classification was achieved. We developed an LDA model calibrated with the analytical results of 5 elements (B, Ca, Cr, K, and Mg) chosen by a backward stepwise procedure. This LDA model identified the origin of wines as Japan with 91% certainty. This result suggests that discriminating by inorganic element composition may be useful to identify Japanese wine for import clearance.

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The effects of 1-methylcyclopropene (1-MCP) on ascorbate (AsA) metabolism were studied and the possible molecular mechanisms were discussed for two cultivars of cauliflower (Brassica oleracea, L. var. botrytis), ‘Violet Queen’ and ‘Snow Crown’. Ethylene production was suppressed in ‘Violet Queen’, while it was increased in ‘Snow Crown’ by 1-MCP treatment. Meanwhile, the changing patterns of AsA metabolism after harvest were different between the two cauliflower cultivars. In ‘Violet Queen’, AsA content decreased in the control after harvest, and the loss of AsA was delayed by 1-MCP treatment. In ‘Snow Crown’, AsA content remained almost constant and was not affected by 1-MCP treatment. In ‘Violet Queen’, the gene expression of BO-APX1, BO-APX2, and BO-sAPX was down-regulated, while the gene expression of BO-DHAR and BO-GLDH was up-regulated by 1-MCP treatment. The regulation of these genes contributed to the suppression of AsA reduction in ‘Violet Queen’ treated by 1-MCP. In ‘Snow Crown’, simultaneous down-regulation of BO-APX1, BO-APX2, and BO-sAPX, which were responsible for AsA breakdown, and BO-MDAR1, BO-MDAR2, BO-DHAR, and BO-GLDH, which were responsible for AsA regeneration and biosynthesis, might lead to maintain the AsA level constant with 1-MCP treatment.

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