• Journal of Plant Biotechnology
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• v.31 no.4
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• pp.255-259
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• 2004

Agrobacterium tumefaciens-mediated cotyledonary node transformation was used to produce transgenic soybean. Cotyledonary node explants of three cultivars and one genotype were co-cultivated with strains Agrobacterium (LBA4404, GV3101, EHA101, C58) containing the binary vectors (pCAMBIA3301 and pPTN289) carrying with CaMV 35S promoter-GUS gene as reporter gene and NOS promoter-bar gene conferring resistance to glufosinate (herbicide Basta) as selectable marker. There was a significant difference in the transformation frequency depend on bacteria strain. The EHA101 strain of the bacterial strains employed gave the maximum efficiency (3.6%). One hundred-six lines transformed showed the resistance in glufosinate. Histochemical GUS assay showed that at least 11 plants transformed with the GUS gene were positive response. The soybean transformants were obtained from the Thorne (5 plants), 1049 (5 plants) and Bakun (1 plant), respectively. Southern blot analysis and leaf painting assay revealed that the GUS and bar gene segregated and expressed in their progeny.

• Journal of Plant Biology
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• v.39 no.2
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• pp.127-135
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• 1996

An efficient system of somatic embryogenesis was established for the red pepper plant (Capsicum annuum L. cv. Nokkwang) usign immature zygotic embryos. The size of the immature zygotic embryos and the concentrations of 2, 4-D and sucrose were found to be critical. Somatic embryos were induced via callus or directly from explants and regenerated into plantlets successfully. When zygotic embryos 1~2 mm long were cultured on the modified Murashige-Skoong (MS) medium supplemented with 2 mg/L 2, 4-D for 3 weeks in the dark, somatic embryos were induced directly from the apical region of zygotic embryos with the highest frequency being approximately 90%. To mature the somatic embryos, ABA and an ethylene inhibitor AgNO3 were used. The highest frequency of shoot regeneration (25% in each) resulted at 2$\mu$M ABA or 20$\mu$M AgNO3 treatment at rates 3.7 and 1.6 times control, respectively. Shoots developed mainly from the cotyledonary node on CoCl2-containing medium, and from the upper side of cotyledon on medium containing AgNO3 while the embryos on the control medium produced shoots from both the cotyledonary node and the upper region of cotyledons both at frequencies of 50%. Indirect somatic embryogenesis via callus was induced at an efficiency of approximately 10% with zygotic embryos 3~4 mm long cultured on MS medium containing 5~10 mg/L, 2, 4-D for 5~7 weeks under a continuous light condition. The plants regenerated from the somatic embryos were morphologically normal. Using scanning electron microscopy, the direct and indirect somatic embryogeneses were observed to follow the globular, heart and torpedo stages, similar to zygotic embryogenesis. Also, suspensors appeared in the early globular and ovoid-shaped late globular embryos during indirect somatic embryogenesis.

• Journal of Plant Biotechnology
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• v.43 no.3
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• pp.341-346
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• 2016

The objectives of this study were to 1) evaluate the efficiency of the protocol of Agrobacterium-mediated transformation of cucumber to introduce phytoene synthase-2a carotene desaturase (PAC genes); 2) demonstrate the integration of PAC genes into the genome of putative transgenic cucumber based on growth on selection medium, PCR and Southern analysis; 3) evaluate the expression of PAC genes in transgenic cucumber based on the analysis of RT-PCR and Northern blot hybridization. Out of 5,945 cotyledonary-node explants inoculated with Agrobacterium, 65 (1.1%) explants produced 238 shoots. Integration of PAC genes into the genome of the cucumber was demonstrated based on the analysis of gDNA-PCR, 21 out of the 238 plants regenerated; while 6 plants proved positive for Southern blot hybridization. Transgene expression was demonstrated based on analysis of RT-PCR, 6 plants proved positive out of the 6 plants analyzed; while 4 plants out of 6 proved positive during Northern blot hybridization. This study successfully demonstrated the production of transgenic cucumber, integration, and expression of the PAC gene in cucumber.

• Korean Journal of Plant Tissue Culture
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• v.25 no.1
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• pp.69-74
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• 1998

The somatic embryos of Daucus carota L. cv. Hongshim developed in MS basal medium far 4 weeks had varied number of cotyledons. Palisade and spongy parenchyma of cotyledon were not clearly discriminated in all seedlings developed from the embryos. No independent existence of collateral vascular bundle was observed in all seedlings with various types of cotyledon ; instead, vascular bundles were either interconnected or partially connected with one another. Most of the cotyledonary bases on hypocotyl showed short cylinder structure which encircle plumule. The vascular tissues of cotyledonary bases and nodes of seedlings with jar-shaped or 1 cotyledon were connected in ring forms, showing the pattern of ectophloic shiphonostele, and similar ring form structure was also found in the vascular arrangement of 5 cotyledon seedlings. The vascular bundles of seedlings with 2, 3 and 4 cotyledons in many cases had independently arranged within the cotyledonary bases and nodes, showing the pattern of eustele. In hypocotyl, tetrarch or hexarch xylems prevailed in seedlings with jar-shaped cotyledon or 1 and 5 cotyledon; tetrarch xylems prevailed in 2 cotyledon seedlings; and triarch xylems prevailed in 3 cotyledon seedlings. In most of seedlings, cortex vascular bundles were dispensed in the region from cotyledonary node to hypocotyl, but double vascular bundles were also observed occasionally. In roots, diarch xylems were observed in most of seedlings with 2 cotyledons, triarch xylems in half of seedlings with 3 cotyledons, and diarch xylems in most of the remaining seedlings with the occasional occurrences of tetrarch xylems.

• Korean Journal of Plant Resources
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• v.26 no.4
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• pp.511-515
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• 2013

Hypocotyls explants of melon seedling were cultured on Murashige and Skoog's (MS) medium supplemented with 1 mg/L 2,4-dichlorophenoxy acetic acid (2,4-D) and 0.5 mg/L benzyl aminopurine (BA) for 6 weeks to produce somatic embryos. In somatic embryos produced through intervening bright yellow friable (BYF) from the explants, somatic embryos with two-cotyledon (26%) and horn-type cotyledon (74%) were observed. The procambial strand of cotyledons was originated from circular procambial tissues of lower hypocotyls. The circular procambial independently divided into two procambial strand at the edge of cotyledonary-node, and then connected to each cotyledon to form somatic embryos with two-cotyledon. When cotyledon was horn-type, the circular procambial strand in lower hypocotyls would continuously remain connected to the cotyledon. However, somatic embryos with two or horn type cotyledon formed an abnormal shoot apex without the tunica-corpus structure or dome shape in the inter-cotyledonary area. These results demonstrated that the variation of cotyledon in somatic embryos was closely related to procambial tissue differentiation and shoot apical formation.

• Plant Biotechnology Reports
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• v.3 no.3
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• pp.205-211
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• 2009

An efficient in vitro regeneration protocol for moth bean [Vigna aconitifolia (Jacq.) Marechal] via somatic embryogenesis has been developed. Embryogenic callus cultures were established from the cotyledonary node as explant on semi-solid Murashige and Skoog (MS) medium supplemented with $0.75mg\;1^{-1}$ 2,4-dichlorophenoxyacetic acid (2,4-D) and $1.5mg\;1^{-1}$ 6-benzylaminopurine (BA) and with various additives ($50mg\;1^{-1}$ ascorbic acid and $25mg\;1^{-1}$ each of adenine sulphate, citric acid and $_L-arginine$). Numerous somatic embryos differentiated on MS basal nutrient medium supplemented with $0.25mg\;1^{-1}$ 2,4-D and $0.5mg\;1^{-1}$ of kinetin (Kin). Sustained cell division resulted in the formation of cell aggregates, which progressed to the globular- and heart-shaped somatic embryos and then, if they differentiated properly, to the torpedo shape and cotyledonary stages. The transfer of embryos onto fresh MS basal medium containing $0.2mg\;1^{-1}$ BA and $2.0mg\;1^{-1}$ gibberellic acid enabled the embryos to achieve complete maturation and germination. More than 80% of somatic embryos were converted into true-to-type fertile plants. In vitro-regenerated plantlets with well-developed roots were successfully hardened in a greenhouse and established in soil.

• Journal of Plant Biology
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• v.31 no.4
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• pp.289-298
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• 1988

The pattern of elongation in the developing hypocotyl of Glycine max shows that the elongation generally proceeds from base to the cotyledonary node in acropetal diredtion, although earlier elongation takes place through the entire hypocotyl. Because the differentiation of the vascular cambium in the hypocotyl advances also acropetally, it can be seen that the acropetal wave of hypocotyl elongation is associated with the acropetal differentiation of the cambium in the hypocotyl elongation is associated with the acropetal differentiation of the cambium in the hypocotyl elongation is associated with the acropetal differentiation of the cambium in the hypocotyl. The elongation of procambial cells occurs not only during active elongation but also after cessation of elongation of the hypocotyl. In tangential view, the procambium of the hypocotyl in early stage has homogeneous structure composed of short cells. Subsequently, these procambial cells elongate actively and then become elongated long cells. These long cells eventually become fusiform initials, while some of elongated long cells are transversely divided and then converted into ray initials. The characteristics of the vascular cambium are entirely acquired some time after hypocotyl elongation is completed, and the transitin from procambium to vascular cambium in the hypocotyl is a rather gradual process.

• Journal of Plant Biology
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• v.34 no.4
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• pp.274-281
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• 1991

The relationship between elongation of hypocotyl and its vascular transition was studied with seedling of Glycine max. the hypocotyl elongation proceeded acropetally from the base of hypocotyl toward the cotyledonary node. The vascular transition did not occur in the basal region of the hypocotyl, which did not nearly elongate, with exarch radial vascular bundles. However, the vascular transition was almost completed at the middle part of the hypocoty, more or less elongated, with endarch collateral vascular bundles. Such bundles also appeared in the uppermost region of the hypocotyl, in which the elongation was the most striking. These results suggested that the vascular transition was related to the hypocotyl elongation and that the transition of primary vascular system in Glycine max seedling was established by rather rapid process. Our observations of the serial sections from root to hypocotyl revealed that the vascular system through the root-hypocotyl-cotyledon was a unit, to which one of the epicotyl that did not participate in transition was superimposed.

• Journal of Plant Biotechnology
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• v.36 no.4
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• pp.344-351
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• 2009

Current status of soybean transformation method in Korera was reviewed with recent publications. Most frequently used method for genetic transformation was Agrobacterium-mediated transformation on cotyledonary node which is most popular method used in foreign country. In addition to this, various methods such as sonicationmediated transformation, in planta transformation, and transformation on meristem tissue of germinating seed, have been tried in Korea, even though their efficiencies on repeatability and stability were relatively low. Based on the promising results developed recently by reviewer, several important considerations for successful soybean transformations were suggested. They are 1) proper genotype screening, 2) targeting transformation on exact point, 3) multiple shoot formation, 4) efficient selection pressure, 5) successful shoot elongation, 6) efficient root formation. These are the basic requirements for stable and highly efficient soybean transformation of Korean soybean.

• Korean Journal of Plant Tissue Culture
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• v.22 no.5
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• pp.291-298
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• 1995

This study describes plant regeneration from leaf explant of Bupleurum falcatum through somatic embryogenesis, and the effect of 2,4-dichlorophenoxyacetic acid on somatic embryo abnormalities. The relationship between the cotyledon number of somatic embryo and its germinability is also described. Embryogenic calli were selected from calli formed on explants cultured on MS solid basal medium supplemented with 1 mg/L 2,4-D. Cotyledonary abnormalities were observed in somatic embryos which were developed from calli cultured on MS medium with 1 mg/L 2,4-D for 6-week and then subcultured on 2,4-D free MS medium for 3 weeks. The frequency of abnormalities was as follows: 7% of somatic embryos had one cotyledon, 65% of them had two cotyledons, 25% three cotyledons, 5% four cotyledons, 2% five cotyledons, and 3% trumpet-like cotyledons. The two cotyledon somatic embryos were germinated at a frequency of 80%. However the germination frequency of one cotyledon embryo and multicotyledonary embryo was lower than that of the two cotyledon somatic embryo. All of trumper-like somatic embryos did not germinate. Histological observations of multicotyledon embryo showed circular procambium in the root but pocambial strands in the cotyledonary node or upper hypocotyl. The number of the strands was equal to the cotyledon number.