• Title/Summary/Keyword: Ginseng cotyledon

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Effects of Growth Regulators on Somatic Embryogenesis from Ginseng Zygotic Embryos (인삼 접합자배로부터 체세포배의 발생에 미치는 생장조절제의 영향)

  • Yong Eui CHOI;Woong Young SOH
    • Korean Journal of Plant Tissue Culture
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    • v.22 no.3
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    • pp.157-164
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    • 1995
  • Intact mature zygotic embryos or their excised cotyledons of ginseng, were cultured on media containing various growth regulators such as auxin (2,4D, IAA) and cytokinin(BAP kinetin). In the culture of intact zygotic embryos, auxin inhibited germination but cytokinin did not Somatic embryogenesis occurred only from those of ungerminated embryos. In the culture of cotyledon segment, medium without growth regulators was the most appropriate to somatic embryogenesis. Somatic embryos were produced sporadically over the surfaces of zygotic embryos on medium containing auxin, while on medium without growth regulators, or media containing cytokinin, somatic embryos formed only on the proximal region of cotyledon. on medium containing 2,4-D, somatic embryos originated from multiple cells which comprised epidermal and subepidermal layers of cotyledon, which resulted in poly-somatic embryogenesis. When these somatic embryos were cultured on the same medium, the primary somatic embryos procured secondary embryos, which arose from epidermal or subepidermal single cells.

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REDIFFERENTIATION FROM TISSUE CULTURE AND ISOLATION OF VIABLE PROTOPLASTS IN PANAX GINSENG C.A. MEYER (고려인삼의 조직배양에 의한 기관형성과 원형질체배양에 관한 연구)

  • Choi Kwang-Tae;Yang Deok-Chun;Kim Nam-Won;Ahn In-Ok
    • Proceedings of the Ginseng society Conference
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    • 1984.09a
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    • pp.1-11
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    • 1984
  • Ginseng cotyledon calli were cultured on 1/2MS media supplemented with combination of various growth regulators to induce more embryoids and plantlets in a short period. And tissues of ginseng root and calli were also incubated under various factors or conditions to establish methods for the isolation of viable protoplasts in Panax ginseng C.A. Meyer. The calli derived from cotyledon produced numerous embryoids in 1/2MS media containing 0.5mg/$\ell$ 2,4-D and 0.5mg/$\ell$ kinetin after 2 months' culture. But only shoot formation was less frequent. Further development of these embryoids occurred on 1/2MS medium supplemented with the same concentration of BA and GA. Viable protoplasts were isolated from the root tissue and callus of ginseng. The specific conditions for the isolation of viable protoplasts were required of ginseng materials, root tissue and callus, being processed. For the production of viable protoplasts from 1-year old ginseng root tissue, an enzyme mixture of $2\%$ cellulase 'Ono-zuka' and $0.5\%$ macerozyme, an enzyme solution pH of 5.2 to 5.8, a 7- to 8- hour incubation period at $28{\pm}1^{\circ}C$, and 0.9M mannitol as osmoticum in the cell enzyme mixture were optimum, while the treatments with an enzyme mixture of $2\%$ cellulase 'Onozuka', $2\%$ macerozyme and $1\%$ driselase, and 25-hour incubation period at $28{\pm}1^{\circ}C$, were more efficient for the production of viable protoplasts from ginseng callus.

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Structural Changes and Histochemical Study of Endosperm on Panax ginseng C.A. Meyer during Embryo Development (인삼(Panax ginseng C.A. Meyer) 종자의 배발달에 따른 배유의 구조변화 및 조직화학적 연구)

  • 유성철;김유갑
    • Journal of Ginseng Research
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    • v.16 no.1
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    • pp.37-43
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    • 1992
  • Structural changes of the endosperm of Panax ginseng C.A. Meyer from fertilization to germination were investigated by light microscope. The endosperm of the ginseng seed is cellular type. Since endosperm cells adjacent embryo continuously breakdown and disappear with the elongation of embryo, the real of endosperm is gradually decreased. As the anatropous ovules of immature seed with green seed coat developes more and more, ovary cells adjacent ovary cavity become abundant by the periclinal division, their size is decreased, hypotrophy of cell wall discern, and they are gradually differentiated in seed coat. Though embryo responds strongly to basic dye at the stage of completion of endosperm formation, tissue of endosperm responds to acidic dye positively Cell wall of embryo and endosperm are composed of primary cell wall not lignified. Endosperm cells adjacent embryo begin to breakdown in the endosperm tissue of indehiscent seed before the beginning of the after-ripening. Dehiscent seed of which seed coat is opened through after-ripening represent the form as a seedling in the result of embryo developments with the formation of organs; radicle, cotyledon, plumule. Umbilifom layer represents strong positive response to the toluidine blue and the basic function. Umbiliform layer that endosperm cells breakdown and disappear is observed clearly at the periphery of the embryo cotylemon, while slightly at the periphery of the radicle.

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STUDIES ON THE TISSUE CULTURE OF PANAX GINSENG

  • Harn C
    • Proceedings of the Ginseng society Conference
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    • 1974.09a
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    • pp.9-22
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    • 1974
  • Unlike the tissue culture in animals and human being, in higher plants various parts of the plant are cultured for varied purposes, and they are named variously depending on which parts are used as explants or what purposes they are cultured for. Followings are some of the names of culture used frequently: organ culture, tissue culture, callus culture, single cell culture, meristem culture, mericlone culture, ovary culture, ovule culture, embryo culture, endosperm culture, anther culture, pollen culture, protoplast culture, etc.. As the names of the culture indicate, in some kinds of culture the explants used for culture are actually not tissues, but organs, single cells, or protoplasts. It seems, however, convenient to call all of the above-mentioned cultures grossly as tissue culture. Several kinds of tissue culture were attempted using Panax ginseng as material and some of the results were summarized below. 1. Callus culture After dormancy of the sed was broken, whole embryo or parts (hypocotyl, cotyledon and epicotyl) of partly grown embryo were cultured in the media supplemented with growth regulators. Rapid swelling occurred in a few weeks, but most of the swelling was observed only in the basal part of epicotyl, changes in the other parts of embryo appearing in much later stages. The swelling or increase in size, however, was resulted not from the divisions of cells, but from the mere expansion of cell. Real calli were formed about two months after inoculation of explants. Callus tissues developed from cortex, pith, and vascular bundle in the cases of hypo- and epicotyl, from mesophyl tissue in the case of cotyledon. Shoots developed more easily from cotyledons regardless of whether they are detached from or attached to the embryo proper. 2. Culture in the Knudson C medium When cotyledons, detached from or attached to the embryo proper, were cultured in the growth regulator-free Knudson C medium comprision only several kinds of mineral compounds and sucrose, shoot primordium or callus developed profusely and finally plantlets were produced directly from shoot primordium or indirectly through callus. In this medium epidermal cells as well as mesophyl cells of the cotyledon became meristematic and divided, changing into multinucleate cells or multicellular bodies, developing eventually into either shoot primordia or calli. 3. Anther culture Anthers were cultured in the media supplemented with various growth regulators applied singly or in combinations. Callus was formed mostly in the connective tissue of anther. Cells of anther wall layers changed in appearance, but no division occurred. Microspores of all stages in development were not changed, ruling out the possibility that microspore-originated callus might be formed. 4. Isolation of protoplast Protoplasts were isolated from young root, leaf, and epicotyl, using 0.7M D-mannitols as osmoticum and using macerozyme and cellulase respectively for maceration and digestion of the cell wall. Production in large number of naked intact protoplast was rather difficult as compared with other plant species. Fusion of protoplasts occurred infrequently mainly due to the fewer number of naked protoplasts in the solution.

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Identification and Changes of Physiologically Active Substances During Chilling Storage of Dehisced Ginseng Seeds (저온저장중 개갑인삼종자내의 생리활성물질 동정 및 변화)

  • Kwon, Woo-Saeng;Baek, Nam-In;Lee, Jung-Myung
    • Journal of Ginseng Research
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    • v.21 no.1
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    • pp.13-18
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    • 1997
  • Identification and changes of physiologically active substances during chilling storage of dehisced ginseng (Panax ginseng C. A. Meyer) seeds were analyzed using various preparatory separation methods and purification columns; Dowex 50W and silica gel columns. Seven components with Rf values of 0.20, 0.40, 0.58, 0.66, and 0.70 In solvent system, $CHCl_3$:MeOH=3:1 (v/v), Rf values of 0. 63 and 0.74 in solvent system, $CHCl_3$:MeOH:$H_2O$:=7:3:1 (v/v) were obtained through Dowex 50W and silica gel column chromatographies. Two components with Rf values of 0.20 and 0.63 in the all chilling treatments were detected in the extract obtained through both chromatographies, and only the former component was gradually increased till 4 weeks of chilling storage and then rapidly decreased from 8 to 16 weeks. UV spectra of Rf values of 0.66 and 0.56 were similar to that of cytokinin, but their physiological activities were not found. Rf values of 0.20 showed activity by radish cotyledon expansion bioassay. The component with Rf value of 0.20 was revealed to have a naphthalene in the proposed chemical structure by various NMR techniques.

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The Condition of Regeneration and Antibiotics Concentration for Gene Transformation of Zygotic Embryo in Panax ginseng C. A. Meyer (인삼(Panax ginseng C. A. Meyer)접합자 배의 형질전환을 위한 재분화 및 항생제 농도 조건)

  • 양덕춘;이은경;최원균;김무성
    • Korean Journal of Plant Resources
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    • v.16 no.1
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    • pp.74-81
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    • 2003
  • To obtain multi-shoot using zygotic embryos dissected from ginseng seed, the embryos were cultured on MS medium supplemented with CPA and BA. Effective multi-shoot induction was achieved on 0.5mg/ t CPA and 1.0mg/ t BA treatment. Among the various plant growth regulator treatment, MS basal medium with 1mg/ t 2,4-D and 0.5mg/ t kinetin was more competent and could be induced 4∼6 shoots per one embryo. Also, the best condition for pre-embryoid induction from ginseng cotyledon so as to ginseng transformation appeared to 1mg/ t 2,4-D and 0.5mg/ t kinetin treatment. The kanamycin level to select transformants varied greatly by different explant tyues. The petiole explants with leaf and embryo could survived up to 100$\mu\textrm{g}$ / ml kanamycin concentration where as petiole explants without leaf died all at the level. Conclusionally, our results suggest that optimum kanamycin concentration for ginseng transformation using somatic embryos is about 75∼100$\mu\textrm{g}$ / ml concentration.

Genetic Transformation of Panax ginseng with Herbicide Resistant Gene (제초제 저항성 유전자에 의한 인삼의 형질전환)

  • 양계진
    • Korean Journal of Plant Tissue Culture
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    • v.28 no.6
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    • pp.353-357
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    • 2001
  • Transformation of ginseng plants was achieved by biolistic system with cotyledon explants and callus using phosphinothricin acetyl-transferase (PAT) gene resisting to a herbicide of Bialaphos. The binary vector for transformation was constructed with disarmed Ti-plasmid and with double 355 promoter. The introduced NPT II and PAT genes of the transgenic ginseng plants were successfully identified by the PCR, and the survival test on the medium with basta. The transgenic ginseng plants were propagated using repetitive secondary embryogenesis. The transgenic ginseng plantlets had normal structures of roots and shoots, and dormant buds for new year sprouting. We transferred the transgenic plants to greenhouse and observed the continuing growth until a new year.

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Direct somatic embryogenesis, plant regeneration and genetic transformation of Panax ginseng

  • Park, Yong-Eui;Yang, Deok-Chun;Park, Kwang-Tae;Soh, Woong-Young;Hiroshi Sano
    • Proceedings of the Botanical Society of Korea Conference
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    • 1999.07a
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    • pp.85-89
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    • 1999
  • Somatic embryogendesis is one of good examples of the basic research for plant embryo development as well as an important technique for plant biotechnology. This paper describes the direct somatic embryogenesis from zygotic embryos of Panax ginseng is reversely related to normal axis growth of zygotic embryos by the experiment of various chemical treatments. Under the normal growth condition, the apical tips of embryo axis produced an agar-diffusible substance, which suppressed somatic embryo development from cotyledons. Although the cells of zygotic embryos were released from the restraint of embryo axis, various factors were still involved for somatic embryo development. Electron microscopic observation revealed that the ultrastructure of cells of cotyledon epidermis markedly changed before initiation of embryonic cell division, probably indicating reprogramming events into the cells embryogenically determined state. Polar accumulation of endogenous auxin or cell-cell isolation by plasmolysis pre-treatment is the strong inducer for the somatic embryo development. The cells for the process of somatic embryogenesis might be determined by the physiological conditions fo explants and medium compositions. Direct somatic embryos from cotyledons fo ginseng were originated eithrer from single or multiple cells. The different cellular origin of somatic embryos was originated either from single or multiple cell. The different cellular origin of somatic embryos was depended on various developmental stages of cotyledons. Immature meristematic cotyledons produced multiple cell-derived somatic embryos, which developed into multiple embryos. While fully mature cotyledons produced single cell-derived single embryos with independent state. Plasmolysis pretreatment of cotyledons strongly enhanced single cell-derived somatic embryogenesis. Single embryos were converted into normal plantlets with shoot and roots, while multiple embryos were converted into only multiple shoots. GA3 or a chilling treatment was prerequisite for germination and plant conversion. Low concentration of ammonium ion in medium was necessary for balanced growth of root and shoot of plantlets. Therefore, using above procedures, successful plant regeneration of ginseng was accomplished through direct single embryogenesis, which makes it possible to produce genetically transformed ginseng efficently.

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