Korean Journal of Plant Tissue Culture (식물조직배양학회지)

The Korean Society of Plant Biotechnology (한국식물생명공학회)
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The Filter Membrane Culture Procedure with Feeder Cells in Rice Protoplast Culture

Filter membrane과 feeder세포를 이용한 벼의 원형질체 배양

  • LEE, Sung-Ho (Plant Molecular Biology and Biotechnology Research Center) ;
  • SHON, Young Geol (Department of Horticulture, Gyeongsang National University) ;
  • Lee, Soo In (Plant Molecular Biology and Biotechnology Research Center) ;
  • DAVEY Micheal R. (Plant Genetic Manipulation Group, Department of Life Science, The University of Nottingham) ;
  • COCKING Edward C. (Plant Genetic Manipulation Group, Department of Life Science, The University of Nottingham) ;
  • CHO, Moo Je (Plant Molecular Biology and Biotechnology Research Center)
  • Published : 1997.09.01
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Abstract

To investigate the response on feeder cell cultures, protoplasts isolated from cell suspensions initiated from mature seed scutellum-derived callus of the Japonica rice variety Taipei 309, were cultured on filter membranes under various conditions. The effects of various factors, such as gelling agents, feeder cell and protoplast densities, species of feeder cells and heat shock treatment, have been investigated to improve protoplast plating efficiencies on filter membranes. Maximum protoplast plating efficiencies were obtained when protoplasts were cultured on KPR medium semi-solidified with Sea Plaque agarose at a density of $5\;\times\;10^{5}\;ml^{-1}$ protoplasts in the presence of Lolium multiflorum as feeder cells (0.5 ml pcv per 10 ml of protoplast culture medium). Pre-culture heat shock treatments for 1 min. and 5 min. to the protoplasts did not give any appreciable increase on the plating efficiency of protoplasts in the presence of feeder cells. Maltose-supplemented medium was superior for plant regeneration from protoplast-derived colonies compared with medium containing only sucrose. The plants were transferred to the glasshouse, flowered and were fertile.

Japonica 벼 품종 Taipei 309 성숙 종자의 배반에서 유도된 캘러스로부터 유기 시킨 세포 현탁 배양체에서 원형질체를 분리하여 filter membrane과 feeder 세포를 이용한 여러가지 조건에서 배양하였다. 이러한 조건들은 gelling agents, feeder 세포와 원형질체 밀도, feeder 세포의 종류 및 heat shock 처리 등이며 이들이 filter membrane 배양 방법에서 원형질체 평판 효율에 미치는 효과들을 조사하였다. 원형질체 평판 효율은, Lolium multiflorum을 feeder 세포로 사용하고 (10 mL의 원형질체 배양 배지당 0.5 mL pcv) 원형질체를 mL 당 $5\;\times\;10^{5}$개로 하여 Sea Plague agarose 배지에 원형질체를 배양 했을때 최고치를 얻었다. 원형질체에 heat shock 처리를 했을 때 원형질체 평판 효율은 변화가 없었다. carbohydrate source로서 sucrose로서 sucrose 대신에 maltose를 사용했을 때 식물체 재분화율이 높았으며 원형질체로부터 재분화된 이들 식물체들은 임성을 나타내었다.

Keywords

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