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  • Author or Editor: Toyoki Kozai x
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Broccoli (Brassica oleracea L. Botrytis Group `Ryokurei') plantlets, cultured photoautotrophically (without sugar in the medium) in vitro for 3 weeks at 23C and 160 μmol·m–2·s–1 photosynthetic photon flux (PPF), were stored for 6 weeks at 5, 10, or 15C under 0 (darkness) or 2 μmol·m–2·s–1 PPF (continuous lighting) supplied by fluorescent lamps (white light). Dry weight of the plantlets stored for 6 weeks at 5 or 10C in light was not significantly different from that of the plantlets before storage. Dry weight of the plantlets decreased as temperature increased and was maintained at higher levels in light than in darkness. Chlorophyll concentrations of the plantlets were higher at the lower temperatures. Chlorophyll fluorescence kinetics indicated higher activities of chlorophyll of the plantlets stored in light than in darkness. Lighting at as low as 2 μmol·m–2·s–1 PPF was important to preserve photosynthetic and regrowth abilities and dry weight of the plantlets during low-temperature storage.

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A storage method of transplants in vitro was developed using light compensation points in conjunction with low temperatures. Broccoli (cv. Ryokurei) plantlets, aseptically germinated and cultured for three weeks in vitro, were used as model transplants. Culture conditions were: 23C air temperature, 160 μmol m-2s-1 PPF, and 3.6 air exchanges per hour of the vessel. Prior to storage, light compensation points were determined at 3, 5, 10, and 15C for the plantlets cultured with or without 20 g liter-1 sugar in the medium. Plantlets were stored for six weeks at 5, 10, and 15C under either 0 or 2 μmol m-2s-1 continuous PPF. The light compensation points varied with air temperature and with medium sugar level. Plantlet dry weight during storage was best maintained by keeping CO2 exchange rate of the plantlets close to zero throughout the storage period. High transplant qualities were successfully preserved at light compensation points: 2 μmol m-2s-1 PPF at 5-10C without sugar, and at 5C with sugar in the medium. This method may be applicable for storage of other crop transplants, plug seedlings and cuttings as well.

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Growth and net photosynthetic rate of potato (Solanum tuberosum L.) `Benimaru' plantlet in vitro were studied under a conventional photomixotrophic condition [with 20 g sucrose/liter in the medium and under 70 μmol·m-2·s-1 photosynthetic photon flux (PPF)] with minimal ventilation (MV) and under photoautotrophic conditions (without sugar in the medium and under 190 μmol·m-2·s-l PPF) with enhanced natural ventilation using an air diffusive filter (DV) or with forced ventilation (FV). Fresh weight of the plantlets cultured in the FV and DV treatments was 2.4 times that of the plantlets cultured in the MV treatment. Net photosynthetic rate and dry weight per plantlet were the highest in FV followed by DV. For photoautotrophic micropropagation, FV was superior to DV.

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Commercial transplant production in Japan has been increasing rapidly since 1985. Transplant production began with plug seedlings for bedding plants, followed by carnation and Chrysanthemum plug transplants vegetatively-propagated using cuttings. Next, production more recently includes plug seedlings of lettuce and cabbage, and micropropagated tubers of potato plants and grafted transplants of tomato, eggplant, cucumber, and watermelon plants. The reasons for the rapid increase in commercial production of transplants will be reviewed. The current “cutting edge” practices include hardening before shipping or planting. The pros and cons of current transplant production systems in Japan will be discussed. Recent research advances in production of micropropagated, grafted and seedling transplants are reviewed with special reference to environmental control for hardening or acclimatization. Research on robotic or automated systems for micropropagation, grafting, and transplanting currently developed in Japan are described.

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A photoautotrophic or sugar-free medium micropropagation system (PAM) using five large culture vessels (volume = 120 L each) with a forced ventilation unit for supplying CO2-enriched air was developed and applied to commercial production of calla lily (Zantedeschia elliottiana) and china fir (Cunninghamia lanceolata) plantlets. The culture period of calla lily plantlets in the PAM was reduced by 50%, compared with that in a conventional, photomixotrophic micropropagation system (PMM) using small vessels each containing a sugar-containing medium. Percent survival ex vitro of calla lily plantlets from the PAM was 95%, while that from the PMM was 60%. The production cost of calla lily in the PAM was reduced by about 40%, compared with that in the PMM, and the initial investment per plantlet for the PAM was ≈10% lower than that for the PMM. The sales price of ex vitro acclimatized calla lily plantlet was increased by 25% due to its higher quality, compared with plantlets produced in the PMM.

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We recently showed that spinach (Spinacia oleracea L.) transplants produced under a short photoperiod and low air temperature were characterized by a delay of bolting and short flower-stalk length at harvest (Chun et al., 2000a). The present study was conducted to determine whether these changes are caused by the short photoperiod itself or by the lower integrated photosynthetic photon flux (IPPF). Shoot and root dry weights of transplants increased significantly with increasing IPPF, but were not affected by a change in the photoperiod. However, the floral development indices of transplants were significantly greater under a 16-than under a 10- or 13-hours/day photoperiod, but were not affected by a change in IPPF. The percentage of bolted plants 3 days after transplanting (DAT) increased significantly with increasing photoperiod (from 0% at 10 hours/day to more than 85% at 16 hours/day). Flower-stalk length increased with increasing photoperiod (e.g., at 14 DAT, from 15 mm at the shorter photoperiods to 80 mm at 16 hours/day), but was not affected by a change in IPPF. These results show that the delay of bolting that occurs when the photoperiod is reduced during transplant production is due to the delay of floral development and not to retarded vegetative growth as a result of reduced IPPF.

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For vegetative propagation of sweetpotato, single or multi-node leafy cuttings are used as propagules. A quantitative understanding of leaf development is important for predicting the number of propagules produced after a given production period under various environmental conditions. For plant production in a relatively closed structure, controlling CO2 concentration is necessary, but effects of CO2 concentration on cutting production rates of sweetpotato are not well-investigated. Single-node cuttings each with a fully expanded leaf (the initial leaf blade length was 66 mm) were grown under one of three levels of CO2 concentration (400, 800, and 1200 μmol·mol-1), 250 μmol·m-2·s-1 PPF, 16 h/day photoperiod, and 29 °C air temperature. The plant dry weight increased faster in the higher CO2 concentrations. Changes in the number of harvestable cuttings during the production period was defined by changes in the number of leaves reaching a leaf blade length (LBL) longer than a given standard length (LS). The number of harvestable cuttings increased almost linearly with time after the LBL of the first leaf reached the LS, regardless of CO2 concentration. The effect of CO2 concentration on cutting production rate (number of harvestable cuttings per day) was varied with different LS. For example, at LS = 20, 30, and 40 mm, the cutting production rate increased slightly at higher CO2 concentrations, while at LS = 60 mm, it decreased significantly at higher CO2 concentrations. This indicates that, under the present experimental conditions, increasing CO2 concentration increased the number of small leaves that might not be usable as cuttings (propagules). Environmental control is necessary in vegetative propagation to increase the number of propagules and the biomass usable as propagules, thereby minimizing energy and resources needed for the propagule/transplant production process.

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For vegetative propagation of sweetpotato, single or multi-node leafy cuttings are used as propagules. A quantitative understanding of leaf development and the effects of environment conditions on leaf emergence and expansion rates is important for predicting the number of propagules produced after a given production period. Single-node cuttings each with a fully expanded leaf were grown under two levels of photosynthetic photon flux (PPF, 160 and 250 μmol·m–2·s–1) and photoperiod (10 and 16 h/day). The time courses of the number of leaves larger than the standard leaf area (As) were obtained by analyzing the time courses of leaf blade length recorded every day on each leaf. The number of leaves larger than a given As increased almost linearly after the first leaf reached to the As. PPF and photoperiod affected both the duration until the appearance of the first leaf with As and the leaf development rates (leaves per day). The effects of PPF were more pronounced than photoperiod for the development rate of the leaves regardless of As. Results obtained in these experiments were incorporated into our previously developed model, and the number of propagules produced under different environment conditions was predicted. Such techniques need to be used effectively for planning and environment control of vegetative propagation.

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A system was designed for measuring the CO2 exchange rates [net photosynthetic rate (Pn) and dark respiration rate] of in vitro plantlets in situ (in the vessel with natural ventilation). The system, excluding gas cylinders, was placed in a growth chamber so that the desired photosynthetic photon flux (PPF) and temperature could be maintained during the measurement. The CO2 concentration inside the culture vessel (Ci) was indirectly controlled by controlling the CO2 concentration outside the vessel (Co). The Pn of the plantlets was estimated based on the measured Ci and Co at steady state using a gas chromatograph according to the method described by Fujiwara et al. (1987). The performance of the system was demonstrated by measuring the in situ Pn of sweetpotato [Ipomoea batatas (L.) Lam., cv. Beniazuma] and tomato (Lycopersicon esculentum Mill., cv. Hana Queen) plantlets in vitro under a range of CO2 concentrations and PPF. The photosynthetic parameters of the Pn model (Niu and Kozai, 1997) for the plantlets were then estimated based on the measured Pn. The preliminary measurements demonstrated the potential application of the system.

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