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Studies on the N_2-laser microbeam system for efficient gene delivery and the utilization of this system for plant protection

Research Project

Project/Area Number 06404009
Research Category

Grant-in-Aid for General Scientific Research (A)

Allocation TypeSingle-year Grants
Research Field 植物保護
Research InstitutionOsaka Prefecture University

Principal Investigator

OZAKI Takeshi  Osaka Pref.Univ.Agriculture Assoc.Prof., 農学部, 助教授 (00081555)

Co-Investigator(Kenkyū-buntansha) FURUKAWA Hajime  Osaka Pref.Univ.Agriculture Res.Assoc., 農学部, 助手 (40240957)
MORIKAWA Toshinobu  Osaka Pref.Univ.Agriculture Assis.Prof., 農学部, 講師 (90145821)
OHKI Satoshi  Osaka Pref.Univ.Agriculture Assis.Prof., 農学部, 講師 (00128761)
MORI Genjiro  Osaka Pref.Univ.Agriculture Assis.Prof., 農学部, 講師 (20081515)
YAMAGUCHI Toshihiko  Osaka Pref.Univ.Agriculture Assoc.Prof., 農学部, 助教授 (30081558)
Project Period (FY) 1994 – 1995
Project Status Completed (Fiscal Year 1995)
Budget Amount *help
¥19,700,000 (Direct Cost: ¥19,700,000)
Fiscal Year 1995: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1994: ¥17,400,000 (Direct Cost: ¥17,400,000)
KeywordsN_2-laser microbeam / Gene delivery / Plant cell / Cucumber mosaic virus / Cucumis figarei / Resistance / Transformation / N_2レーザーマイクロビーム / 直接遺伝子導入 / キユウリモザイクウイルス / ウイルス抵抗性 / PBI121 / CMV‐RNA / Avena macrostachya
Research Abstract

An N_2-laser with a 337.1 nm wavelength and 3 ns pulse width burned localized hole ca.0.5 mum in diameter on the cell wall and cell membrane of cultured cells of melon or tobacco under selected irradiation conditions (1.5-6 muj/spot). No visible damage due to the irradiation was detected under these conditions. Fluotescent-isothyocyanate (FITC) was used to determine the osmotic value of the medium surrounding cultured cells and 0.3 Molarity was the best to get 66.3 % of cells which showed fluorescence. Efficiency of gene delivery into plant cells was tested using this N_2 laser treatment. Mature pollen of lmpatiens balsamina, I.walleriana and Lycoris albiflora was irradiated with a laser at 3muj/spot in medium containing 0.3 M mannitol and plasmid PBI221 harboring beta-glucuronidase. The maximum frequency of transient gene expression was 24% in these treatments. It is concluded that the osmotic value of medium surrounding the pollen grains is extremely important for the GUS gene epression to use N_2-laser microbeam and it is able to introduce any gene into pollen exactly one by one. When the melon cotyledons were treated with a laser in the presence of cucumber mosaic virus RNA,virus multiplication was observed in the epidermal and around the cells 24 hr after irradiation. These results suggest that the present laser method is useful for the transformation of plant cells. Resistance in the African wild plant, Cucumis figarei Des.et.Noude to CMV was found to related to restricted virus movement, and the rates of cell-to-cell and long distance movement in C.figarei is regulated by sequences in CMV RNA 3. A cDNA clone encoding the 3a and coat protein gene of CMV (pepo strain), linked to the cauliflower mosaic virus 35S promotor, was introdeced into tobacco by N_2-laser microbeam and the transgenic tobacco plants expressing the CP gene were protected from infection with CMV.

Report

(3 results)
  • 1995 Annual Research Report   Final Research Report Summary
  • 1994 Annual Research Report
  • Research Products

    (25 results)

All Other

All Publications (25 results)

  • [Publications] 尾崎 武司: "Cucumis melsならびに同属異種野生植物の数種ウイルスに対する反応" 関西病虫研報. 36. 13-20 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 佐古 勇: "株分けネギの病原ウイルスとウイルス再感染による生育・収量への影響" 関西病虫研報. 36. 21-27 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 尾崎 武司: "N_2レーザーマイクロビームによる植物細胞への直接遺伝子導入の試み" 植物組織培養. 12. 73-78 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 尾崎 武司: "メロンと同属異種野生植物Cucumis figareiに含まれる抗ウイルス活性物質の部分精製と作用機構" Appl.Biol.Sci.1. 41-52 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Yonaka: "Pepper vein yellows virus,a novel luleo virus from fell pepper plants in Japan" Ann.Phytopath.Soc.Japan. 61. 178-184 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 田部井 豊: "ジゴキシゲニン(DIG)を利用した高感度RAPD法:品種識別への応用" 育雑. 46. 307-308 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 尾崎 武司 (分担): "植物病理学事典" 養覧堂, 1220 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Ohki, S.T.: "Virus-Taxonomy-Sixth report of the international committee of taxonomy of viruses" Springer-Verlag,

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Osaki, T.: "Reaction of Cucumis melo and wild relatives to several viruses" Proc.Kansai Pl.Prot.36. 13-20 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Sako, I.: "Occurence of garlic latent virus and onion yellow virus in seedless welsh onion and influence of viruses reinfection on their field." Proo, Kansai Pl.Prot.36. 21-27 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Osaki, T.: "Direct gene introduction into plant cells using an N_2 laser microbeam." Plant Cell and Tissue Culture. 12. 73-78 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Osaki, T.: "Partial purification and antiviral activitiy of a plant virus inhibiting protein found in the African wild plant, Cucumis figarei." Appl.Biol.Sci.1. 41-52 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Yonaha, T.: "Pepper vein yellows virus, a novel luteovirus from bell pepper plants in Japan." Ann.Phytopath.Soc.Japan. 61. 178-184 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] "Development of highly sensitive RAPD method bu Digoxigenin incorporated PCR ; Application for identification of cultivars." Breeding Science. 46. 307-308 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Morikaw, T.: "Transfer of mildew resistance from the wild oat Avena prostrata into the cultivated oat." Bull.Univ.Osaka Pref.Ser.B. 47. 1-10 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Mayama, S.: "Association between avenalumin accumulation, infection hypha length and infection type in oat crosses segregating for resistance to Puccinia coronata f.sp.avenae race 226." Physiol.Mol.Plant Pathol.46. 255-261 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 尾崎武司: "N_2レーザーマイクロビームによる植物細胞への直接遺伝子導入" 植物組織培養. 12. 73-78 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] 尾崎武司: "メロンと同属異種野性植物Cucumis figareiに含まれる 抗ウイルス活性物質の部分精製と作用機構" Appl.Biol.Sci.1. 41-52 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Yonaha,T.: "Pepper vein yellows virus,a novel luteo virus from bell pepper plants in Japan" Ann.Phytopath Soc Japan. 61. 178-184 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] 尾崎武司: "ヒガンバナ 科および ユリ科球根花卉類のりん 葉切片からの 子球形成とウイルス 無毒化" 植物組織培養. 13(印刷中). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Yanagawa,T.: "In vitro propagation and elimination of virus by bulb scale cultures of Hippeastrum bulbs" Plant Tissue Culture Letter. 13(印刷中). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] 尾崎武司: "植物病理学事典" 養賢堂, 1220 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] 大木理: "植物病理学事典" 養賢堂, 1220 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] 尾崎武司: "N_2レーザーマイクロビームによる植物細胞への直接遺伝子導入" 植物組織培養. 12(印刷中). (1995)

    • Related Report
      1994 Annual Research Report
  • [Publications] 尾崎武司: "メロンと同属異種野生植物Cucumis figareiに含まれる抗ウイルス活性物質の部分精製と作用機構" Appl.Biol.Sci.1(印刷中). (1995)

    • Related Report
      1994 Annual Research Report

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Published: 1994-04-01   Modified: 2016-04-21  

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