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Masayuki Oda and Tatsuto Nakajima

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Takahiro Tezuka, Hisa Yokoyama, Hideyuki Tanaka, Shuji Shiozaki and Masayuki Oda

Mature seeds of Ilex species usually contain immature embryos and are extremely difficult to germinate. Ilex latifolia and I. rotunda, two species that are grown as ornamentals, also produce seeds that are difficult to germinate. In the present study, we investigated some factors affecting seed germination in those species. Although seeds of I. latifolia and I. rotunda could imbibe water, they did not germinate. When embryos were cultured in vitro, germination was observed in I. latifolia but not in I. rotunda. Interestingly, a transient decrease in germination frequency occurred in I. latifolia embryos isolated from seeds collected in September or October. Among five types of I. latifolia seeds that differed in the presence of the endocarp, testa, and endosperm, germinability of isolated embryos was highest. Good germination was also observed in quarter-seeds with or without endocarp followed by half-seeds. Treatment of seeds with sodium hydroxide (NaOH) had no effect on seed germination in I. latifolia. Cold stratification at 5 °C increased the germination frequency of I. latifolia embryos. In conclusion, the data suggested a mechanical barrier by the endocarp and inhibitors contained in the endosperm, testa, and/or endocarp inhibited seed germination in I. latifolia. Although no seeds or embryos of I. rotunda germinated, cold stratification in combination with other treatments deserves further investigation. Chemical name: sodium hydroxide (NaOH).

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Masayuki Oda, Kunihiko Okada, Hidekazu Sasaki, Shigeki Akazawa and Masahiro Sei

Eggplant (Solanum melongena L.) at the two-leaf stage were grafted on scarlet eggplant (Solanum integrifolium Poir.) by hand or a newly developed robot. The scion and rootstock were fixed with an elastic tube for hand grafting or with an adhesive and a hardener for robotic grafting. After acclimatization, the grafted plants were planted at the three- or 11-leaf stage in a glasshouse. Plants grafted by the robot showed a higher percentage of survival, and attained the three- and 11-leaf stages 8 days earlier on average than those grafted by hand. Stems were longer, and shoot fresh mass and fruit yield of plants were higher for the three-leaf-than for the 11-leaf-stage planting, irrespective of the grafting method. Such vigorous growth and high yield by robotic grafting were absent for the 11-leaf-stage planting but obvious for the three-leaf-stage planting.

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Masafumi Johkan, Genjirou Mori, Kazuhiko Mitsukuri, Keiichirou Mishiba, Toshinobu Morikawa and Masayuki Oda

The aim of this study was to establish an alternative method to produce clones of tomato plants by modification of the complete decapitation method, which regenerates multiple shoots from the cut surfaces of the main and lateral stems of plants grown in vivo. Shading the stems of tomato plants drastically increased the number of regenerated shoots from 2.4 in controls with unshaded stems to 36.2 in shaded stems. In shaded stems, the concentrations of chlorophyll and phenolic compounds were stable for 3 weeks after cutting, whereas these amounts increased in unshaded stems. Inhibiting the production of phenolic compounds in the shaded stem tissues was associated with an acceleration of shoot formation in vivo.

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Takahiro Tezuka, Masashi Harada, Masahumi Johkan, Satoshi Yamasaki, Hideyuki Tanaka and Masayuki Oda

Adventitious shoots can be regenerated from the cut surface of the primary shoot and lateral branches in decapitated plants in vivo. This inherent regenerative ability of plants is useful for mass propagation. In the present study, we conducted histological observations of shoot regeneration and applied auxin and cytokinin to decapitated seedlings in four tomato cultivars. The cultivars produced different numbers of adventitious shoots after decapitation; ‘Petit’ produced the largest number of adventitious shoots (78.5 ± 10.2) and ‘Momotaro’ produced the fewest (12.1 ± 3.3). Histological observation of ‘Petit’ revealed that adventitious shoots regenerated from calli formed at the cut surface of stems. Adventitious shoot formation was inhibited by the presence of lateral branches. Shoot regeneration was prevented by application of 1-naphthaleneacetic acid to ‘Petit’. Application of 6-benzyladenine promoted shoot regeneration in ‘Momotaro’. These results suggest auxin synthesized de novo from the lateral branches inhibited shoot regeneration after decapitation and endogenous cytokinin might stimulate shoot regeneration. Chemical names: 1-naphthaleneacetic acid (NAA); 6-benzyladenine (BA)

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Kazuhiko Mitsukuri, Takaya Arita, Masahumi Johkan, Satoshi Yamasaki, Kei-ichiro Mishiba and Masayuki Oda

Habenaria radiata is a terrestrial orchid with beautiful bird-shaped petals. The wild H. radiata population has been severely affected by environmental disruption and overexploitation. In micropropagation of H. radiata, although aseptic germination has been studied, tissue culture methods have not yet been established. Shoot apexes and leaf explants from vegetative plants and flower stalks, stolons, and floret explants from reproductive plants were chosen for this study. Explants were cultured on half-strength inorganic salts and full-strength vitamins of Murashige and Skoog (1/2 MS) medium containing 30 g·L−1 sucrose, 8 g·L−1 agar (pH 5.6) supplemented with 4.44 μM N6-benzyladenine, and 0.54 μM α-naphthaleneacetic acid. After 8 weeks of culture, the highest survival rate was obtained with floret explants excised from plants at the reproductive phase. In floret culture, the number of adventitious bud formation per explant was 5.4 per upper floret and 4.0 per lower floret. Dark preconditioning, which inhibited browning and contamination, of explants before shoot apex culture increased survival rates of explants (53%) and bud formation (83%). Consequently, a tissue culture method using florets and shoot apexes as explant material was established for H. radiata.

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Masahumi Johkan, Tomoko Chiba, Kazuhiko Mitsukuri, Satoshi Yamasaki, Hideyuki Tanaka, Kei-ichiro Mishiba, Toshinobu Morikawa, Masayuki Oda, Chihiro Yamamoto and Hiroshi Ohkawa

There is concern that high temperatures resulting from global warming could reduce fruit set of tomato (Solanum lycopersicum). However, fruit set of parthenocarpic tomato genotypes, which often bears seedless fruit, is not reduced when grown under a high temperature. The cause of seedless fruit development was studied with the aim of increasing the seed number in parthenocarpic tomato. Ovule number at anthesis in parthenocarpic and non-parthenocarpic fruit did not differ, but the proportion of undeveloped ovules increased with time after anthesis in parthenocarpic tomato, whereas most ovules in non-parthenocarpic tomato developed normally. Pollen grains germinated on the stigma and extruded pollen tubes in parthenocarpic and non-parthenocarpic tomatoes, but in parthenocarpic tomato, pollen tube elongation was markedly inhibited in the style base. Elongation of pollen tubes on agar containing indoleacetic acid (IAA) was depressed in parthenocarpic and non-parthenocarpic tomato plants. p-Chlorophenoxyisobutyric acid (PCIB), which inhibits auxin action, did not affect the fruit set and fresh weight in either type of tomato, although seed number per fruit in parthenocarpic tomato was significantly increased from 13 ± 2 to 74 ± 6 seeds by PCIB treatment. These results indicated that a high IAA concentration in the ovary of parthenocarpic tomato inhibited pollen tube elongation, and that poor fertilization resulted in failure of ovule development. Moreover, floral organs in parthenocarpic tomato were normally developed as in non-parthenocarpic tomato, and seed development could be induced in parthenocarpic tomato by PCIB treatment.