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.
Kee Yoeup Paek and Toyoki Kozai
Kee-Yoeup Paek and Eun-Joo Hahn
Adventitious buds regenerated from homogenized leaf tissue of Saintpaulia ionantha Wendl. `Crimson Frost' were micropropagated to determine types and frequencies of the variants obtained. Plants grown for one year in a greenhouse showed 67% variation and 33% normality. A higher rate of variation was observed in leaf color rather than in leaf shape. The variations in leaf color and leaf shape were 67% and 19%, respectively. In regard to flower type, greater numbers of semidouble and double types were obtained as compared to single types. Both flower types showed a much higher rate of normal (mixed) color (81%) as compared to pink, red or white (19%). An sodium dodecylsulfate polyacrylamide gel electrophoresis profile of protein extracted from leaves of the stock plants and the variants indicated no difference between them and did not reflect the variation in phenotype.
Cheol Hee Lee, Kee Yoeup Paek, J. Brian Power, and Edward C. Cocking
This study was designed to assess the general limitations of somatic hybridization as one of the key technologies for genetic manipulation in plants. The limits of somatic hybridization against different taxonomic backgrounds, intraspecific to interfamilial, were also assessed. Protoplast culture studies provided essential information relating to the species cultural and morphogenetic capacity. several #elect Ion strategies for the recovery of somatic hybrid colonies/plants were developed and assessed using various combinations of protoplast sources and species in the genera Petunia, Nicotiana, Salpiglossis and Chrysanthemum. Morphological, cytological and biochemical analyses were performed to confirm the hybridity of plants or cell lines recovered following protoplasm fusion (using 4-5 methods) and selection.
The somatic hybrid callus/plants were obtained at intraspecific to interfamilial levels by complementation to chlorophyll proficiency, together with media selection or complementation of nitrate reductase deficient mutants as follows; P. Hybrida var. Monsanto (+) P. hybrida cv. Blue Lace (intraspecific), P. hybrida var. Monsanto (+) P. inflata and P. parviflora (interspecific), P. parviflora (+) N. tabacum (intergeneric), S. sinuata (+) P. hybrida var. Monsanto, P. parodii and N. tabacum (intertribal), and C. morifolium (+) S. sinuata.
From this study, it appeared that there were no taxonomic limits to the production and proliferation of somatic hybrid cell lines. However, obtaining morphologically normal hybrid plants met with limited success as the taxonomic relationships became more distant. The regeneration capacity of somatic hybrids seemed to be controlled by both parental species. Somatic incompatibility mechanism was also shown to operate on chromosome elimination. Such chromosome elimination may well be advantageous in plant improvement.
Xuan-Chun Piao, Debasis Chakrabarty, Eun-Joo Hahn, and Kee-Yoeup Paek
In vitro nodal cuttings of potato (Solanum tuberosum L.) `Atlantic' and `Russet Burbank' from bioreactor culture were hydroponically cultured for 28 days using a deep flow technique (DFT) system. The response of plant growth and photosynthesis to different levels of solution electrical conductivity (EC; 0.08, 0.15, 0.22 and 0.36 S·cm-1) and pH (3, 4, 5, 6 and 7) were studied. The best growth, characters of shoot length, total shoot and root fresh and dry weight, were obtained in nutrient solution of pH 6.0 and EC 0.15 S·cm-1 for `Atlantic', while pH 7.0 and EC 0.15 S·cm-1 were found to be best for `Russet Burbank'. Plantlet growth was reduced by low solution pH (3.0) and high EC level (0.36 S·cm-1). Photosynthetic rate, stomatal conductance, and transpiration rate were also found to be affected by EC levels. Down regulation of photosynthesis, as indicated by chlorophyll fluorescence results, were observed when potato plantlets were cultured under nutrient solution of higher EC level. Plantlet growth and photosynthetic rate increased as photosynthetic photon flux (PPF) levels increased from 50 to 250 μmol·m-2·s-1. Particularly, increasing PPF level had a more distinctive effect on plantlet growth than CO2 enrichment condition. It was apparent from this study that nutrient solution of pH 6.0 and 0.15 S·cm-1 EC in combination with high PPF level (250 μmol·m-2·s-1) were suitable for hydroponic culture of potato plantlets as it would maximize net photosynthetic rate, and achieve the highest growth rates.
Chun-Woo Nam, Dong-Lim Yoo, Su-Jeong Kim, Jong-Taek Suh, Myoung-Rae Cho, and Kee-Yoeup Paek
This experiment was carried out to determine correct applications of GA3 for the production of cut flowers of Zantedeschia albomaculata cv. Black Magic in highlands. Tubers were treated with GA3 in concentrations of 0, 100, and 200 mg·L-1. ABA contents in the tubers and roots were twice as high (20∼25 pmol/mL) in the control as in GA3-treated tubers and roots. ABA content in roots increased with increasing GA3 concentration. Growth of the calla was investigated according to the tuber hardness (3.3, 6.0 kg·cm-2), GA3 concentrations (0, 100, 200 mg·L-1), and GA3 treatment duration (24 h, 12 h, and 30 min before planting). Tubers with higher tuber hardness were strong against soft rot, regardless of the GA3 concentrations and treatment durations. Tubers with lower tuber hardness showed over 90% soft rot occurrence when treated with 200 mg·L-1 GA3 for 24 h before planting. However, soft rot did not occur when treatment with 200 mg·L-1 GA3 was used for 12 h before planting.
Nguyen Phuc Huy, Vu Quoc Luan, Le Kim Cuong, Nguyen Ba Nam, Hoang Thanh Tung, Vu Thi Hien, Dung Tien Le, Kee Yoeup Paek, and Duong Tan Nhut
Paphiopedilum spp. is one of the most commercially popular orchids because of its variety of shapes, sizes, and colors. However, it is at risk for extinction because of its exploitation. Regeneration of orchid plants using internode segments is extremely difficult. In this study, young P. callosum plants (1.5 cm) were exposed to eight dark–light cycles (14 days of dark and 1 day of light) for stem elongation to increase the number of nodes to obtain internode tissues. After 75 days of culture, the highest callogenesis (31.25%) was achieved when internode tissue was cultured on liquid Schenk and Hildebrandt (SH) medium containing 30 g·L−1 sucrose, 1.0 mg·L−1 Thidiazuron (TDZ), 1.0 mg·L−1 2,4-Dichlorophenoxyacetic acid (2,4-D), and cotton wool as the support matrix. The optimal media for induction of protocorm-like bodies (PLBs) were the same compositions as previously mentioned and were supplemented with 9 g·L−1 Bacto agar as the gelling agent. PLB clumps (5–6 PLBs/clump) produced the best shoots on medium containing 0.5 mg·L−1 α-Naphthaleneacetic acid (NAA) and 0.3 mg·L−1 TDZ. Among the organic substances tested, 200 g·L−1 potato homogenate (PH) added to Hyponex N016 medium supplemented with 1.0 mg·L−1 NAA, 30 g·L−1 sucrose, 170 mg·L−1 NaH2PO4, 1.0 g·L−1 peptone, and 9 g·L−1 Bacto agar resulted in the best rooting. The rooted plantlets with four to five leaves were acclimatized and had a 100% survival rate. The method presented in this research provides a strategy for the development of highly effective propagation of Paphiopedilum species using ex vitro explants for both conservation and horticultural purposes.