• Journal of The Korean Society of Grassland and Forage Science
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• v.31 no.3
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• pp.217-222
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• 2011

Success in molecular breeding for better adapted varieties to environmental stresses depend upon the concerted efforts by various research including tissue culture, transformation, genetics and breeding. In order to optimize tissue culture conditions of Siberian wildrye grass, the effects of plant growth regulators on callus induction and plant regeneration were investigated with mature seeds. The highest callus induction frequency was observed when the mature seeds were cultured on MS medium supplemented with 5 mg/L 2,4-D. The highest plant regeneration frequency was observed when callus was transferred to N6 medium supplemented with 1 mg/L 2,4-D and 3 mg/L BA. Regenerated plants were grown normally when shoots were transplanted to the soil. A short tissue culture period and regeneration system would be beneficial for molecular breeding of Siberian wildrye grass by the production of transgenic plant.

• FLOWER RESEARCH JOURNAL
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• v.16 no.1
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• pp.12-16
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• 2008

This study was carried out to produce haploid plants to verify a systematic breeding program and genetic analysis. Effect of developmental stage of pollen grains and pre-treatment temperature on production of haploid plants was investigated. Microscopic investigation of the explants (Lilium Asiatic hybrid 'reamland' revealed that the length of flower bud at 23.0-24.9, 25.0-26.9, and 27.0-28.9 mm long coincided with tetrad, uninucleate, and binucleate, respectively. When the efficiency of the anther culture from microgametogenetic stages was tested, late uninucleate to early binucleate stage, having the length of 23.0 to 28.0 mm long flower bud, was the best. The frequencies of the callus induction and plant regeneration from the stage mentioned above were 17.8 and 6.7%, respectively. When calli were cultured on the MS medium containing picloram and zeatin at $25^{\circ}C$, shoots were obtained. Roots of regenerated plantlets were confirmed as haploid through an microscopic observation.

• Journal of Plant Biotechnology
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• v.50
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• pp.56-62
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• 2023

Transgenic lilies have been obtained using Agrobacterium tumefaciens (AGL1) with the plant scale explants, followed by DL-phosphinothricin (PPT) selection. In this study, scales of lily plants cv. "red flame" were transformed with the pCAMBIA3301 vector containing the gus gene as a reporter and the blpR gene as a selectable marker, as well as a gene of interest showing herbicide tolerance, both driven by the CaMV 35S promoter. Using a 20-minute infection time and a 5-day cultivation period, factors that optimized and demonstrated a high transformation efficiency were achieved. With these conditions, approximately 22-27% efficiency was observed for Agrobacterium-mediated transformation in lilies. After transformation with Agrobacterium, scales of lilies were transferred to MS medium without selective agents for 2 weeks. They were then placed on selection MS medium containing 5 mg/L PPT for a month of further selection and then cultured for another 4-8 weeks with a 4-week subculture regime on the same selection medium. PPT-resistant scales with shoots were successfully rooted and regenerated into plantlets after transferring into hormone-free MS medium. Also, most survived putatively transformed plantlets indicated the presence of the blpR gene by PCR analysis and showed a blue color indicating expression of the gus gene. In conclusion, when 100 scales of lily cv. "red flame" are transformed with Agrobacterium, approximately 22-27 transgenic plantlets can be produced following an optimized protocol. Therefore, this protocol can contribute to the lily breeding program in the future.

• Journal of Plant Biotechnology
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• v.39 no.3
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• pp.140-145
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• 2012

The flowering locus T (FT) gene, of which expression will be controlled at high temperature by heat shock promoter (it printed as to HSproFT), was introduced into spray-type chrysanthemum (Dendranthema grandiflorum (Ramat.) Kitamura) 2 cultivars ('Pink PangPang' and 'Pink Pride' by co-cultivation with Agrobacterium tumefaciens strain C58C1 harboring pCAMBIA2300 containing the HSproFT gene. After leaf segments of the 2 cultivars were infected with the A. tumafaciens with C58C1 as explants, shoots were regenerated from the explants cultured on the $1^{st}$ selection medium (MS basal salts + 1.0 mg/L BA, 0.5 mg/L IAA + 10 mg/L kanamycin + 0.7% plant agar, pH 5.8). The shoots were transferred into the $2^{nd}$ selection medium (MS basal salts + 1.0 mg/L BA, 0.5 mg/L IAA + 20 mg/L kanamycin + 0.7% plant agar, pH 5.8). One hundred seventeen plantlets from 'Pink PangPang' and 5 ones from 'Pink Pride' were confirmed as transformants by PCR analysis. Twenty six of the transformants and non-transformants were acclimatized and established well in a green house. Eights of 26 transgenic lines showed flower bud 1.7~10 days earlier than nontransgenic plants, and 24 of them flowered 1~6 days earlier than non- transgenic plants. The shape and color of flower of all HSproFT-transgenic lines were not different with those of non- transgenic plants.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2002.11a
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• pp.29-30
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• 2002

The goal of this research was to develop the effectiveness of in vitro culture method for A. formosanus and study the environment in vitro conditions affecting on growth. The first series of experiments were examined to investigate the response of three different basal media, MS (Murashige and Skoog, 1962), Knudson (KC; Knudson, 1946) and modified hyponex on growth and multiplication during in vitro culture. Multiple shoot proliferation was induced in shoot tip explants on Hyponex (H3) media supplemented with BA (1 mg1$\^$-1/) or TDZ (1-2 mg1$\^$-1/). Addition of activated charcoal (1%) to the TDZ containing medium promoted rapid shoot tip proliferation (11.1 shoots per explant) but the same medium had an opposite effect resulting in poor proliferation in the nodal explants. However, the regenerated shoots had slow growth rate and failed to elongate. This problem was overcome by transferring the shoot clumps to a hormone free H3 media supplemented with 2% sucrose and 0.5% activated charcoal. Using bioreactor culture for scaling up was also shown the best way for multiple shoot induction and growth of this plant. The second series of experiments was studied to investigate the effect of physical environment factors on growth of in vitro plantlets. The Anoectochilus formosanus plantlets were cultured under different air exchange rate (0.1, 0.9, 1.2h$\^$-1/), without sucrose or supplement 20g.1$\^$-1/ (photoautotrophic or photomixotrophic, respectively), and different photosynthesis photon flux (40, 80, 120 ,${\mu}$mol.m$^2$.s$\^$-1/- PPF). Under non-enrichment CO$_2$ treatment, slow growth was observed in photoautotrophical condition as compared with photomixotrophical condition on shoot height, fresh weigh and dry weight parameters; High air exchange (1.2.h-l) was found to be inadequate for plant growth in photomixotrophical condition. On the contrary, under CO$_2$, enrichment treatment, the plant growth parameters were sharply (visibly) improved on photoautotrophic treatments, especially on the treatment with air exchange rate of 0.9.h-1. The growth of plant in photoautotrophic condition was not inferior compared with photomixotrophic, and the best growth of plantlet was observed in treatment with low air exchange rate (0.9.h-1). Raising the PPF level from 80 to 120${\mu}$mol.m$\^$-2/.s$\^$-1/ decreased the plant height, particularly at 120${\mu}$mol.m$\^$-2/.s$\^$-1/ in photoautotrophic condition, fresh weight and dry weight declined noticeably. At the PPF of 120${\mu}$mol.m$\^$-2/,s$\^$-1/, chlorophyll contents lowed compared to those grown under low PPF but time courses of net photosynthesis rate was decreased noticeably. Light quality mainly affected morphological variables, changes of light quality also positively affected biomass production via changes in leaf area, stem elongation, chlorophyll content. Plant biomass was reduced when A. formosanus were grown under red LEDs in the absence of blue wavelengths compare to plants grown under supplemental blue light or under fluorescent light. Stem elongation was observed under red and blue light in the present experiment. Smaller leaf area has found under blue light than with other lighting treatments. Chlorophyll degradation was more pronounced in red and blue light compared with white light or red plus blue light which consequent affected the photosynthetic capacity of the plant. The third series of experiment were studied to investigate the effect of physical environment factors on growth of ex vitro plants including photosynthesis photon flux (PPF), light quality, growing substrates, electrical conductivity (EC) and humidity conditions. In the present experiments, response of plant on PPF and light quality was similar in vitro plants under photosynthesis photon flux 40${\mu}$mol.m,$\^$-2/.s$\^$-1/ and white light or blue plus red lights were the best growth. Substrates testing results were indicated cocopeat or peat moss were good substrates for A. formosanus growth under the greenhouse conditions. In case of A. formosanus plants, EC is generally maintained in the range 0.7 to 1.5 dS.m-1 was shown best results in growth of this plant. Keeping high humidity over 70% under low radiation enhanced growth rate and mass production.

• Horticultural Science & Technology
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• v.34 no.3
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• pp.461-471
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• 2016

This study was conducted to establish an in vitro propagation system for sea-milkwort (Glaux maritima L.), which is an endangered coastal plant species with high horticultural value. Two phenotypes, 'Red type (RT)' and 'Pistachio type (PT)' based on the colors of stem and flower, were obtained from a personal horticulturist in 2009 and used for this study as plant materials. The stock plants showed typical morphologies in flower, capsule, and seed appearances as previously reported. Low temperature treatment at $4^{\circ}C$ for four or more weeks after in vitro sowing maximized seed germination percentage, indicating that imbibition of seed and subsequent low temperature treatment are crucial for its germination. The in vitro seedlings had phenotypic variation, falling into 'RT' and 'PT' classes like the stock plants. Although slight differences depending on genotype and medium were recognized, the fourth or fifth nodes detached from the in vitro seedlings revealed the best multiplication efficacy when estimated on the basis of total number of nodes of newly developed axillary shoots. In addition, the nodes from 'RT' and 'PT' regenerated the most shoots on medium supplemented with $0.5mg{\cdot}L^{-1}$ BA alone and $0.5mg{\cdot}L^{-1}$ BA plus $0.5mg{\cdot}L^{-1}$ IAA, respectively. The node culture-derived plantlets were well acclimatized in a culture room ex vitro and completed the pseudo-annual life cycle coincident with that in the natural salt march habitat with the current cultivation method of applying fresh water-irrigation under an inland environment. This work represents the first report of in vitro propagation of sea-milkwort. Thus, our study will contribute to exo-habitat conservation and natural habitat restoration of this endangered species in addition to development of a horticultural product.

• Journal of Korean Society of Forest Science
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• v.81 no.3
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• pp.273-279
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• 1992

Protoplasts isolated from leaf mesophyll tissues of Populus koreana ${\times}$ P. nigra var. italica were fused with those of P. euramericana cv. Guardi. Well expended healthy leaves of 5 to 7 week-old-plantlet grown in vitro were used as source materials. Leaves from P. koreana ${\times}$ P. nigra var. italica and P. euramericana cv. Guardi were digested in enzyme solution I (2.0% Cellulase, 1.2% Hemicellulase, 0.4% Macrozyme, 2.0% Driselase, 0.05% Pectolyase ; w/v) and enzyme solution II (1.0% Cellulase, 1.2% Hemicellulase, 0.4% Macrozyme, 2.0% Driselase, 0.05% Pectolyase ; w/v), respectively, The highest frequency of fusion among the protoplasts originated from the two source materials was approximately 21% using 40% PEG or 15% dextran. In addition, fusion frequency was enhanced by incorporating 30mM of $Ca^{2+}$ in eluting solution at pH 10.5. Dividing cells and/or mint-calli were obtained by culturing the fusion products in a liquid 8p-KM medium supplemented with 0.6M sucrose, $0.45{\mu}M$ 2, 4-D, and $0.5{\mu}M$ BA. Shoots were regenerated from the fusion product-derived calli after culture on MS medium containing $5.0{\mu}M$ zeatin. To verify the putative hybrid or cybrid, SDS-PAGE was carried out. From the 24 regenerants, just two plants showed intermediate protein band patterns compared with those of the original source plants.

• Journal of Plant Biotechnology
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• v.40 no.2
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• pp.79-87
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• 2013

The aim of this study was to establish plant regeneration from leaf explants of Sedum tosaense Makino, which is globally rare and endangered species. The leaf explants of S. tosaense were cultured on the MS medium supplemented with different concentration of BA and NAA for callus induction. Callus induction was showed the highest (100%) on MS medium containing $2.0mg{\cdot}L^{-1}$ BA and $1.0mg{\cdot}L^{-1}$ NAA. The highest number of shoots were regenerated when callus were cultured on MS medium containing $2.0mg{\cdot}L^{-1}$ BA and $1.0mg{\cdot}L^{-1}$ NAA for 5 weeks. The axillary bud were cultured on the MS media supplemented with combination of BA and NAA for in vitro propagation. The highest number of adventitious shoot (7.9 per explants) formed at $1.0mg{\cdot}L^{-1}$ NAA and $2.0mg{\cdot}L^{-1}$ BA. For rooting, MS medium supplemented with or without $2.0g{\cdot}L^{-1}$ activated charcoal was tested. The optimal results were observed using MS medium supplemented with $2.0g{\cdot}L^{-1}$ activated charcoal, on which 85.7 (No. of root), 4.6 cm (length of root). 1,200 ppm $CO_2$ and 350 ppm $CO_2$ were supplied for make certain the effects of $CO_2$ on pre-acclimatization by photoautotrophic culture. 1,200 ppm $CO_2$ treatment was established higher than 350 ppm $CO_2$ treatment. Soil acclimatization of in vitro plantlets was the best in mixture soil consisted of peat moss and perlite with 100% survival rate and they showed the maximum growth.