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  • Author or Editor: Toyoki Kozai x
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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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Providing continuous light (24-h photoperiod) at a relatively low photosynthetic photon flux (PPF) is one possible way to reduce both initial and operational costs for lighting and cooling during transplant production with an artificial light. However, physiological disorders (i.e., chlorosis and necrosis) are often observed in several species under continuous light with a constant temperature. The objective of this study was to find an effective air-temperature regime under the continuous light to avoid such physiological disorders, and simultaneously enhance floral development, using tomato [Lycopersicon esculentum Mill.] as a model. The seedlings with fully expanded cotyledons were grown for 15 d at a PPF of 150 μmol·m–2·s–1, a relative humidity of 70%, and a CO2 concentration of about 380 μmol·mol–1 (atmospheric standard). Leaf chlorosis was observed when the air temperature was constant regardless of average air temperature (16, 22,or 28 °C). Neither leaf chlorosis nor necrosis was observed when the air temperatures were alternated [periods of high (28 °C) and low (16 °C) air temperatures of 16/8, 12/12, and 8/16 h·d–1]. Faster floral development was observed in the seedlings grown at lower average air temperatures. These results indicated that physiological disorders of tomato seedlings grown under continuous light could be avoided, and at the same time floral development could be enhanced, by lowering the average air temperature through modification of the periods of high and low air temperatures.

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We report the results of serial studies aimed at clarifying several factors affecting organogenesis in rhizome culture of temperate Cymbidium species and their hybrids. The growth patterns and regeneration ability of rhizomes derived from asymbiotic seed or shoot tip culture vary according to media composition, kinds and concentrations of plant growth regulators, culture conditions, and species and varieties. N6-benzyladenine was the best cytokinin for inducing shoot formation, for switching rhizome tissues into protocorm-like bodies, and for directly forming multiple shoots from branched rhizomes. Activated charcoal appeared to be necessary for producing healthy plantlets and for stimulating shoot growth at levels of 0.1% to 0.3% but concomitantly decreased rhizome growth. Sucrose at 5% was the most effective concentration for shoot induction from rhizomes. The above results support the conclusion that organogenic pathways between tropical, subtropical, and temperate Cymbidium species may be controlled by the genetic backgrounds of the species or cultivars.

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The two-spotted spider mite (Tetranychus urticae Koch.) has a worldwide distribution and is one of the most harmful agricultural pests for a variety of plant species. To reveal a visible light wavelength that reduces the population growth rate (as r m, unit: d–1) of the mite, we investigated the r m under wavelength peaks of 468, 515, and 658 nm with half bandwidths of 23, 44, and 22 nm using blue, green, and red light-emitting diodes, respectively. In all treatments, light intensity was set at 2.3 W·m–2 with a light period of 16 h·d–1, and air temperature was set at 25 °C. The survival percentage (l x) and the number of eggs deposited per female (m x) on each age (x, unit: d) were determined to calculate the r m by the following equation: ∑exp(–r m · xl x /100 · m x = 1. The r m and the total m x decreased with increasing wavelength. This result shows that the population growth rate can be reduced under red light (658 nm).

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Cymbidium (cv. Lisa rose) PLB (protocorm-like bodies) were cultured in liquid 1/2 MS medium with/without 20 mg g-1 sucrose under continuous lighting conditions. The vessels were shaken at 100 rpm under PPF (photosynthetic photon flux) of 20 and 140 μmol m-2 s-1 and CO2 concentrations outside the vessel (Cout) of 450 and 2000 μmol mol-1 conditions. Photoautotrophic growth was obtained at high PPF and high Cout. The chlorophyll content of the PLB in the medium without sucrose at high PPF and high Cout was almost 3 times that with sucrose at low PPF and low Cout. The number of newly developed PLB with sucrose at low PPF and low Cout was 1.6 times that without sucrose at high PPF and high Cout; the dry weight per unit PLB with sucrose at low PPF and low Cout was almost 3 times that without sucrose at high PPF and high Cout. Photoautotrophic growth of the PLB might be further promoted at higher CO2 concentration (> 1%).

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Potato (Solanum tuberosum L. cv. Benimaru) plantlets were cultured under four lighting cycles (photoperiod/dark period: 16 h/8 h, 4 h/2 h, 1 h/0.5 h, and 0.25 h/0.125 h) photoautotrophically (without sugar in the medium), and photomixotrophically (with sugar in the medium) in vitro for 28 days. Simulations of time courses of CO2 concentration in the vessel (Ci) and dry weight accumulation of the plantlets cultured photoautotrophically were conducted using a previously developed model (Niu and Kozai, 1997). While underestimation and overestimation of time courses of Ci in some treatments were observed, the simulated results of Ci and dry weight accumulation of the plantlets generally agreed with the measured ones. The difference of net photosynthetic rate response to Ci throughout the culture period was examined between the plantlets cultured photoautotrophically and photomixotrophically. Quantitative relationship between daily net photosynthetic rate (daily net production) and vessel ventilation rate per plantlet was simulated under various CO2 levels outside the vessel for given sizes of potato plantlets cultured photoautotrophically in vitro to aid appropriate CO2 enrichment and vessel design in commercial micropropagation.

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The light and dark periods can be easily controlled by the use of artificial lighting. To understand the effects of alternation of light and dark periods on plant growth and development, we studied the growth and development of tomato (`Momotaro') seedlings under nonperiodic alternation of light and dark periods. Tomato seedlings grown under two nonperiodic alternation treatments of NF (NF-1 and NF-2) were compared with seedlings grown under a periodic alternation treatment (P treatment) with 12-hour light and dark periods. In all treatments, photosynthetic photon flux (PPF) during the light period was maintained at 280 μmol·m-2·s-1; the sum of each light period and the following dark period was 24 hours; and each of the integrated light and dark periods was 132 hours during 11 days of the experiment. In NF-1, the initial light and dark periods were 7 and 17 hours, respectively, and the light period was extended 1 hour per day, while in NF-2, they were initially 17 and 7 hours, respectively, and the light period was shortened 1 hour per day. At the end of the experiment, dry weight per seedling was greater and flower-bud initiation of the first flower truss was earlier in NF-1 than in NF-2 and P, even though the integrated PPF during the experiment was the same in all treatments. These results demonstrate that growth and development of tomato seedlings can be enhanced without any increase in electric energy consumption for lighting by gradually extending the light period or shortening the dark period.

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Lettuce (Lactuca sativa L. cv. Summer-green) plug transplants were grown for 3 weeks under 16 combinations of four levels (100, 150, 200, and 300 μmol·m-2·s-1) of photosynthetic photon flux (PPF), two photoperiods (16 and 24 h), and two levels of CO2 (400 and 800 μmol·mol-1) in growth chambers maintained at an air temperature of 20 ±2 °C. As PPF increased, dry mass (DM), percent DM, and leaf number increased, while ratio of shoot to root dry mass (S/R), ratio of leaf length to leaf width (LL/LW), specific leaf area, and hypocotyl length decreased. At the same PPF, DM was increased by 25% to 100% and 10% to 100% with extended photoperiod and elevated CO2 concentration, respectively. Dry mass, percent DM, and leaf number increased linearly with daily light integral (DLI, the product of PPF and photoperiod), while S/R, specific leaf area, LL/LW and hypocotyl length decreased as DLI increased under each CO2 concentration. Hypocotyl length was influenced by PPF and photoperiod, but not by CO2 concentration. Leaf morphology, which can be reflected by LL/LW, was substantially influenced by PPF at 100 to 200 μmol·m-2·s-1, but not at 200 to 300 μmol·m-2·s-1. At the same DLI, the longer photoperiod promoted growth under the low CO2 concentration, but not under the high CO2 concentration. Longer photoperiod and/or higher CO2 concentration compensated for a low PPF.

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Growth and net photosynthetic rates of shoots of a triploid melon clone, `(L-14 × B) × L-14', were observed over 21 days following transfer from a multiplication MS medium containing 3% sucrose and 10 μM BA to a shoot development medium containing 1 μM BA at varying levels of sucrose in the medium (0%, 1%, and 3%), and light (50, 100, and 150 PPF) and CO2 (500, 1000, and 1500 ppm) in the headspace. Largest numbers of shoot buds were observed in media with 3% sucrose. Increased light and CO2 had a positive interactive effect. Fresh and dry weights were greatest at highest levels of sucrose, light, and CO2. Although there was less growth in the absence of sucrose, fresh or dry weight of shoot buds grown without sucrose in the media still doubled over the 21 days of culture. Net photosynthetic rates of buds were negative 4 days after initiation of culture and approximately zero after 20 days of treatment. When transferring buds to fresh, sugar-free media, net photosynthetic rates became highly positive. Buds that had been cultured in the absence of sucrose and at highest light levels had the highest net photosynthesis rates upon transfer to fresh, sugar-free media.

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Two triploid clones of melon from the same tetraploid parent were grown in vitro with and without sugar, rooted without sugar in media both in a laboratory controlled environment chamber (in vitro) and a greenhouse acclimatization unit (ex vitro), and compared for subsequent nursery growth in the greenhouse unit. The clone `(L-14 c B) × L-14' produced more shoots in both photomixotrophic (with sucrose) or photoautotrophic (sugar-free) conditions. Both genotypes were equally likely to root in sugar-free media, and `(L-14 × B) × L-14' rooted as well from either photoautotrophic and photomixotrophic shoots but `(L-14 × B) × Mainstream' rooted less frequently from photoautotrophic shoots. Seventy-six percent (76%) of the shoots were able to root photoautotrophically in vitro, whereas 47% of the ex vitro shoots were rooted. About 85% of plantlets from all treatments survived after transfer to the nursery. After growth in the greenhouse nursery, the sizes of plants (fresh and dry weight, leaf area) were the same for either clone, from either photoautotrophic or photomixotrophic shoots. Also, after growth in the nursery, plantlets that had been rooted in vitro were larger than those rooted ex vitro. Photoautotrophic rooting demonstrates a concept for integrating micropropagation and plug-type vegetable transplant production.

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