• 제목/요약/키워드: Hypocotyl protoplast

검색결과 6건 처리시간 0.016초

The effects of cytokinin and plating density on protoplast culture of sunflower

  • Chitpan Kativat;Witsarut Chueakhunthod;Piyada Alisha Tantasawat
    • Journal of Plant Biotechnology
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    • 제49권4호
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    • pp.331-338
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    • 2022
  • Sunflower (Helianthus annuus L.) protoplasts were isolated from seven-day-old etiolated hypocotyls of 10 A line and four-week-old fully expanded young leaves of PI 441983 line in vitro seedlings using an enzymatic method. Purified protoplasts were collected by filtration and floatation in sucrose solution. Semi-solid protoplast culture was performed using the L4 regeneration protocol with various culture media and plating densities to achieve the highest efficiencies for protoplast culture of hypocotyl and mesophyll protoplasts of 10 A and PI 441983 lines, respectively. The concentrations in liquid L'4M medium and different plating densities were evaluated in two types of cytokinins, the adenine-type 6-benzyladenine (BA) and the phenylurea-type thidiazuron (TDZ). The highest colony formation was achieved in both sunflower lines when 0.5 mgL-1 BA and 0.5 mgL-1 TDZ were applied with a high plating density (3 × 105 protoplasts mL-1). These conditions led to 38.45% and 39.40% colony formation for hypocotyl protoplasts of the 10 A line and mesophyll protoplasts of the PI 441983 line, respectively. Moreover, many hypocotyl protoplast-derived colonies developed into micro-calli. In addition, superior development of both sunflower protoplasts was observed with all plating densities when BA was used in combination with TDZ. This finding will be applicable to future sunflower hybrid production via somatic hybridization.

양배추 배축 원형질체로부터 식물체 재분화 (Plant Regeneration from Hypocotyl-Derived Protoplasts of Brassica oleracea var. capitata)

  • 이연희;조현석;서석철;김호일
    • 식물조직배양학회지
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    • 제22권1호
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    • pp.7-11
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    • 1995
  • 종자발아 후 5일된 양배추 그린챌린져 배축으로부터 분리된 원형질체로부터 완전한 식물체를 재분화 시켰다. Kao배지와 B5 배지를 기본으로 한 기존의 몇가지 배지에서 원형질체를 배양한 결과 변형시킨 BS배지에서 Plating efficiency는 낮았으나 식물체 재분화율은 가장 높게 나타나 거의 원형질체 배양배지 조성, 생장조절제 농도 등이 재분화에 상당한 영향을 주는 것으로 나타났다. 또한 사용된25가지 종류의 재분화 배지중에서 MS배지에 l% sucrose zeatin 3 mg/L, 1.6% T.C. agar가 첨가된 배지에서 재분화율이 가장 좋았다. 식물체 재분화에 미치는 PVP (polyvinyl Polypyrrolidone)와 agar 농도의 영향으로는 PVP는 큰 효과가 없었고 agar농도를 1.6%로 2배 증가시켰을 때 재분화율이 다소 높아졌다. 현재까지 재분화된 식물체는 150여 개체로서 포장에서 재배중에 있다.

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강남콩(Phaseolus vulgaris L.) Callus의 원형질체 유이 및 배양 (Isolation and Culture of Phaseolus vulgaris L. Callus Protoplasts)

  • 김상구
    • Journal of Plant Biology
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    • 제26권4호
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    • pp.191-196
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    • 1983
  • The isolatin and culture of protoplasts from hypocotyl originated callus of Phaseolus vulgaris cv. Damyang were carried out. The maximum protoplast yield of 4.6$\times$105 per gram fresh callus, using the 13-day-old callus, was obtained by digeston for 6 hours in the enzyme solution. After 10 day-culture of the isolated callus protoplsts, plating efficiency was 50%. Thereafter, cell cluster medium, and followed by leading to callus formation on an agar medium after 3 weeks of the liquid culture.

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원형질체 배양을 통한 배추 〔Brassica campestris ssp. perkinensis〕캘러스 형성 및 뿌리분화 (Callus Formation and Rooting of Inbred Lines of Chinese Cabbage (Brassica campestris ssp. perkinensis) Though Protopalst Culture)

  • 염옥희;전익조;김혜진;백남권;임학태
    • 식물조직배양학회지
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    • 제28권3호
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    • pp.153-157
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    • 2001
  • 기내에서 성장한 배추의 자엽, 하배축과 계대배양한 지 2~3주 되는 어린잎이 원형질체 분리재료로 사용되었다. 배추의 원형질체 분리에 가장 적합한 효소의 조합은 0.4M mannitol 을 삼투조절제로 한 1% Cellulysin과 0.5% Macerozyme의 효소조합이 배추 원형질체 분리에 가장 적합한 것으로 나타났으며, 잎조직을 27$\pm$1$^{\circ}C$에서 30rpm의 속도로 12~16시간 동안 효소와 반응시켰을 때 7.6$\times$$10^{5}$ protoplast/g의 가장 많은 원형질체를 얻었다. 세포분열을 유기시키기 위하여 K8p 배지에 5 mg/L 2,4-D와 2 mg/L에 첨가하였을 때 배양 7~10일에 자엽과 하배축에서 분리한 원형질체에서 세포군들이 형성되었다. 분열한 세포가 8~10 세포크기 단계에 이르면 0.2% agarose 반고체 배지에 고정시켜 배양하였다. 얻어진 calli들을 100가지 다양한 재분화 배지에 옮겼으나 식물체의 재분화는= 이뤄지지 않았지만, 간혹 캘러스에서 뿌리가 분화되는 것은 관찰할 수 있었다.

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STUDIES ON THE TISSUE CULTURE OF PANAX GINSENG

  • Harn C
    • 고려인삼학회:학술대회논문집
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    • 고려인삼학회 1974년도 학술대회지
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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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양배추 protoclone의 변이 (Protoclonal variation in cabbage (Brassica oleracea ssp. capitata))

  • 이연희;조현석;김호일;나종현;이인원
    • 식물조직배양학회지
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    • 제24권3호
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    • pp.175-181
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    • 1997
  • 양배추 F$_1$ 종자 그린 챌린저 배축으로부터 분리된 원형질체로부터 재분화된 protoclone들에 대한 형태적 특성, 배수성, 동위효소, RAPD 분석으로 변이 양상을 살펴보았다. 형태적 특성은 초자, 엽형, 엽색, 엽결각, savoy, wave등을 조사한 결과 대부분이 형태적으로 원식물체와 같았으나 3개체는 잎의 결각이 뚜렷하고 1개체는 savoy와 wave가 심하게 나타났다. Flow cytometry system으로 종자에서 정상적으로 발아한 배축, 자엽과 잎의 배수성을 측정한 결과 배측과 자엽에서는 2배체와 4배체 세포가 혼합되어 있었으나 잎에서는 일정하게 정상적인 2배체로 측정되었다. 잎을 재료로 하여 125개체 protoclone들의 배수성을 본 결과 2배체 91개체(72.8%), 4배체 32개체(25.6%), 혼합배수체 2개체(1.6%)로 나타났다. 또한 4배체를 나타내는 R$_{0}$ protoclone 개체를 자가수분시켜 얻은 종자를 발아시켜 배축, 자엽과 잎의 배수성을 측정한 결과 잎에서 4배체, 배축과 자엽에서는 4배체, 8배체 세포가 혼합되어 있었다. PGM과 MDH 동위효소 분석 결과 R$_{0}$ 정상개체 그리고 변이를 나타낸 개체, 배수성이 다른 개체들간의 밴드양상은 같았으며 15종류의 random primer를 사용한 RAPD에서도 차이는 없었다.없었다.

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