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Richard K. Kiyomoto and Mark H. Brand

Experiments were conducted on tissue proliferation (TP) development and in vitro and ex vitro growth of tissues from plants with (TP+) and without TP (TP-). In 1993 the increase in TP in one-, two-, and three-yr-old `Holden' and `Besse Howells' was 3%, 52%. and 32% and 10%, 26% and 21%, respectively. No differential mortality was observed. Shoot tip cultures initated from TP+ and TP- `Montego' showed 10-12 mo were required for miniaturiziation and multiplication in TP- shoot tips and 4 mo in TP+ shoot tips. TP- cultures require 10 uM 2-iP for normal shoot proliferation; whereas TP+ cultures had to be transferred to hormone-free medium after 6 mo to maintain normal shoot morphology. Cutting propagation from TP- and TP+ plants older than 5 yr, showed persistence of morphological aberrations associated with TP+ plants.

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K.H. Al-Juboory, D.J. Williams, and R.M. Skirvin

Shoots of greenhouse-grown Algerian ivy (Hedera canariensis L.) were surface disinfected and explanted on modified Murashige and Skoog (MS) medium supplemented with BA (10 μm) and NAA (2.5 μm). One month later the shoots were transferred to MS proliferation medium supplemented with TDZ (0.1 or 0.5 μm) and NAA (40 μm). An average of three microshoots developed on each stem treated with TDZ. Pruned shoots grown on MS medium supplemented with GA3 (20 μm) and BA (20 μm) branched better than unpruned shoots (3.7 vs. 1 per explant, respectively). Rooted shoots grown ex vitro grew and developed a shape suitable for commercial sale in 3 months. Chemical names used: N -(phenyl-methyl)-l H -purine-6-amine (BA); gibberellic acid (GA3); 1-naphthaleneacetic acid (NM); N -phenyl-W-1,2,3-thiadiazo-5-yl urea (Thidiazuron, TDZ).

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R. Lowe and D. Donnelly

Minituber production was investigated using ex vitro `Norland' plantlets in a rockwool-based hydroponic system. Productivity was evaluated for 12- and 16-h photoperiod pre-treatment, planting density (two, four, and six plantlets/slab), vertical or horizontal orientation, pinching, and hilling. Total yield differences did not result from photoperiod pre-treatments, but 12-h pre-treatment increased the number of minitubers in the desirable 10- to 40-g size range. Increased planting density reduced yield per plant but caused small increases in yield per slab. Planting orientation, pinching, or hilling had no effect on overall fresh weight yield, number, or size distribution. Short photoperiod pre-treatment, and planting densities of four to six plantlets/slab, oriented vertically, are recommended.

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Issam A. Hassaballa, M.G. Moughieth, N.A. Hagagy, and N.S. Zayed

Shoot tip and single-node cutting explants of `Hamawy' and `El-Amar' apricot cultivars were initiated from forced shoots of field-grown, virus-free trees. Explants were cultured on Murashige & Skoog (MS) Nitsch & Nitsch and Anderson media. Different modifications of MS medium were also evaluated. Antioxidant pretreatment reduced phenolic compounds and decreased necrosis. Modified MS was the best medium for plantlets regeneration, with positive effectiveness of adenine sulfate addition to the modified MS. Shoot multiplication was best on 2.0 mg·L–1 BAP and 1.0 mg·L–1 thidiazuron (TDZ). Also, half-strength MS medium was superior for shoot elongation Surface coverage, 16 hours light/8 hours dark cycle, and 2.0 mg·L–1 IBA induced good rooting. Rooted plantlets were successfully acclimated ex vitro.

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Xiaoling Yu and Barbara M. Reed

A micropropagation system was developed for hazelnut cultivars. Grafted greenhouse-grown plants produced many more viable explants than upper branches of mature field-grown trees. Shoots from grafted greenhouse-grown plants collected March through July and suckers of mature field-grown trees collected in July produced the most growing explants (46% to 80%). Three- to five-fold multiplication was obtained after 4 weeks of culture on NCGR-COR medium supplemented with 6.7 μm BA and 0.04 μm IBA. Roots were produced on 64% to 100% of shoots grown on half-strength NCGR-COR mineral salts and 4.9 μm IBA for 4 weeks. Ex vitro rooting by a brief dip in 1 or 5 mm IBA was equally successful. Transplant survival was 78% to 100%. Chemical names used: N 6-benzyladenine (BA); indole-3-butyric acid (IBA).

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Charlotte R. Chan and Robert D. Marquard

Traditional seed propagation (warm/cold stratification) was compared to embryo culture of Chionanthus virginicus L. to determine if germination could be promoted and time necessary to produce a sizable plant could be reduced. Embryos of C. virginicus were extracted from immature fruit collected 9, 16, and 23 Aug. 1995 and grown in vitro on Anderson's rhododendron medium. They germinated in 4 weeks and were transferred ex vitro to flats. Mature fruit from the same source were grown simultaneously using warm/cold stratification. The two groups were evaluated periodically over a 2-year period for percent germination, plant size, and seedling success. The embryo-cultured plants had a lower survival rate (16% vs. 44%) and were more labor intensive. After 2 years, embryo-cultured plants were 13.4-fold the mass and 4.7-fold taller than traditionally grown plants. Ten-month-old cultured plants were comparable in size to 2-year-old plants grown traditionally.

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Michio Kanechi, Masakatsu Ochi, Michiko Abe, Noboru Inagaki, and Susumu Maekawa

The effects of natural ventilation and CO2 enrichment during the rooting stage on the growth and the rates of photosynthesis and transpiration of in vitro cauliflower (Brassica oleracea L.) plantlets were investigated. In vitro plantlets were established in airtight or ventilated vessels with or without CO2 supplied (≈1200 μg·L-1) through gas permeable films attached to the vessel's cap for 15 days before transplanting ex vitro. Leaves generated in vitro in ventilated vessels had a higher photosynthetic rate than those produced in airtight vessels, which lead to greater leaf expansion and shoot and root dry matter accumulation during in vitro culture and acclimatization. Enhanced photosynthesis in leaves of ventilated plantlets was positively correlated with chlorophyll content. Increasing photosynthetically active radiation from 70 to 200 μmol·m-2·s-1 enhanced the growth of in vitro plantlets under ventilated conditions but it depressed photosynthesis of the leaves grown photomixotrophically with sugar and CO2 enrichment which might be due to the feedback inhibition caused by marked accumulations of sucrose and starch. Higher CO2 levels during in vitro culture enhanced photosynthesis under photoautotrophic conditions, but inhibited it under photomixotrophic conditions. Fifteen days after transplanting ex vitro, high photosynthetic ability and stomatal resistance to transpiratory water loss of ventilated plantlets in vitro had important contributions to rooting and acclimatization. Our findings show that the ventilated culture is effective for accelerating photoautotrophic growth of plantlets by increasing photosynthesis, suggesting that, especially for plantlets growing in vitro without sugar, CO2 enrichment may be necessary to enhance photosynthetic ability.

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Jeffrey P. Schnurr and Zongming Cheng

A selection of Betula platyphylla, from an open pollinated population, was made for upright growth habit, cold hardiness, and a dark green canopy. A micropropagation system was developed to overcome the difficulty with conventional propagation techniques. Shoot-tip cultures were best established in 3/4 strength MS medium supplemented with 0.1 μM thiadiazuron. After 5 weeks in culture, shoots were transferred to woody plant medium (WPM) with 4.4 μM BA. The highest proliferation rate occurred at 24 C on WPM, solidified with agar, and supplemented with 2.2 μM BA. Shoots rooted in vitro and ex vitro and have been established in the field. A regeneration system has also been developed using leaves from aseptic cultures. The optimum conditions for shoot regeneration include a 2-week dark treatment before exposure to a 16-h day/8-h night cycle. Large, healthy leaf explants cultured on WPM with 20 μM BA regenerated shoots at the highest frequency. Regenerated shoots, when transferred to the micropropagation system, proliferate successfully. Currently, a transformation system for this selection is being developed.

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Wenhao Dai, Cielo Castillo, and Victoria Magnusson

In vitro shoot cultures for two birch species, Asian white birch (Betula platyphylla) and paper birch (Betula papyrifera), were initiated from shoot tips of mature trees and maintained in MS (Murashige and Skoog) medium containing 3% sucrose and 5–10 μM (micromolar) benzyladenine (BA). The effect of such factors as genotype, basal medium, and plant growth regulator (PGR) on proliferation was investigated. Shoots were proliferated in both MS and woody plant medium (WPM) supplemented with different concentrations of thidiazuron (TDZ), BA, and kinetin (Kin). Two birch species responded differently to these factors. In general, more shoots were proliferated in WPM than in MS medium. The maximum proliferation rate of Asian white birch was achieved by being cultured in WPM containing 4–8 μM TDZ, while paper birch gave rise to the maximum proliferation rate in WPM supplemented with 20 μM BA. Interactions between genotype and medium or cytokinin were found. Shoots produced on media with TDZ had thick stems and small, dark green leaves. Microshoots can be rooted both in vitro and ex vitro with or without IBA treatment. Plants were regenerated from leaf tissues of Asian white birch. Adventitious shoots regenerated when in vitro leaves were cultured on WPM supplemented with 10–20 μM BA with 2-week dark treatment. The effect of genotype, PGR, and culture condition on in vitro regeneration of birch species is being tested.

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Jung Eek Son, Yil Jang, and Jung Hyuk Seo

Supporting materials for rooting have a considerable influence on the growth and quality of in vitro plantlets. Various supporting materials (rockwool, perlite, vermiculite, and polyurethane) and nutrient supply cycles (12, 24, 36, and 48 hours) were examined to find the optimum conditions for photoautotrophic micropropagation of potato plantlets in the nutrient-circulated micropropagation (NCM) system. In the NCM system, nutrient solution was circulated between the culture vessel and the nutrient reservoir. A plug cell tray with 70 plantlets was placed inside. The number of air exchanges was 10 hours under forced ventilation. Nodal leafy cuttings of plantlets were cultured at CO2 concentrations (mol·mol-1)/PPF s (mol·m-2·s-1) of 350/80, 700/120, and 1500/250 on day 5-11, 12-18, and 19-28, respectively, for all treatments. All growth factors of in vitro plantlets grown for 28 days using rockwool, perlite and vermiculite were greater than those grown using polyurethane. Dry weight of plantlets grown using rockwool was eight times greater than those grown using polyurethane. The same results were obtained in the growth and survival percentages 14 days after transplanting to ex vitro conditions. Optimum nutrient supply cycles were 12, 24, and 48 hours when perlite, rockwool, and vermiculite were used as supporting materials, respectively. It was considered that the range of optimum nutrient supply cycle was affected by water retention characteristics of supporting materials. This study proved that the supporting material and the nutrient supply cycle were very important environmental factors in photoautotrophic mass propagation.