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トランスポゾンを用いたニワトリ胚細胞の半永久的遺伝子操作

Research Project

Project/Area Number 18657070
Research Category

Grant-in-Aid for Exploratory Research

Allocation TypeSingle-year Grants
Research Field Developmental biology
Research InstitutionNara Institute of Science and Technology

Principal Investigator

高橋 淑子  Nara Institute of Science and Technology, バイオサイエンス研究科, 教授 (10183857)

Co-Investigator(Kenkyū-buntansha) 齋藤 大介  奈良先端科学技術大学院大学, バイオサイエンス研究科, 助教 (90403360)
Project Period (FY) 2006 – 2007
Project Status Completed (Fiscal Year 2007)
Budget Amount *help
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2007: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2006: ¥1,800,000 (Direct Cost: ¥1,800,000)
Keywordsトランスポゾン / エレクトロポレーション法 / トリ胚 / Tet-on / ゲノム / 発現誘導 / テトラサイクリン
Research Abstract

トリ胚を用いたエレクトロポレーション法は、さまざまな遺伝子を目的とする組織に導入する方法として極めて有効である。しかしながら、従来のエレクトロポレーション法の問題として、導入された遺伝子の発現が2〜3日間しか維持されないことであった。これは導入された遺伝子が核内染色体に組み込まれないために、細胞の増殖に伴って遺伝子コピー数が減少し、やがては消失してしまうことが原因と考えられていた。我々はこれまでに、これらの問題を克服するために、小型魚類で用いられているTol2トランスポゾン法をトリ胚エレクトロポレーション法に応用し、導入遺伝子をトリゲノム内に安定的に組み込むことに成功している。今年度はさらにこの方法を改良し、我々が最近トリ胚で確立したTet-on法とを組み合わせ、ゲノム上に安定的に組み込まれた外来遺伝子の発現時期を人工的にコントロールできる手法を確立した。準備したplasmidは以下の通り:1)目的とする遺伝子(たとえばEGFP)をTRE(Tetracycline-responsive element)によってドライブさせ、さらにこれらの発現カセットをTol2vectorに組み込んだplasmid。2)rtTA-M2(Dox依存的にTREに結合して転写を活性化させる因子をコードする)を組み込んだTol2vector。3)Tol2vector上の遺伝子カセットがゲノムに組み込まれるために必要なトランスポゼースを発現させるCAGGS-TP。これら3種のplasmidを、エレクトロポレーション法を用いてトリ胚内に共同入し、さらに3日後にDoxを注入すると、EGFPの発現が開始された。これらの新規の方法を用いると、これまで不可能であった胚発生の後期過程におこる器官形成における遺伝子の機能の解析が可能となる。

Report

(2 results)
  • 2007 Annual Research Report
  • 2006 Annual Research Report
  • Research Products

    (24 results)

All 2008 2007 2006 Other

All Journal Article (17 results) (of which Peer Reviewed: 8 results) Presentation (7 results)

  • [Journal Article] Exploration of embryonic origins of germline stem cells and neoblasts in Enchytraeus japonensis(Oligochaeta, Annelida).2008

    • Author(s)
      Sugio, M.
    • Journal Title

      GEP 8

      Pages: 227-36

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] L-Maf regulates p27kip1 expression during chick lens fiber differentiation.2007

    • Author(s)
      Reza, H. M.
    • Journal Title

      Differentiation 75

      Pages: 737-744

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Stage-dependent expression of Pax6 in optic vesicle/cup regulates patterning genes through signaling molecules.2007

    • Author(s)
      Reza, H. M.
    • Journal Title

      Differentiation 75

      Pages: 726-736

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Tet-on inducible system combined with in ovo electroporation dissects multiple roles of genes in somitogenesis of chicken embryos.2007

    • Author(s)
      Watanabe, T.
    • Journal Title

      Dev. Biol. 305

      Pages: 625-636

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Stable integration and conditional expression of electroporated transgenes in chicken embryos.2007

    • Author(s)
      Sato, Y.
    • Journal Title

      Dev. Biol.. 305

      Pages: 616-624

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Cadherin is required for denritic morphogenesis and synaptic terminal organization of retinal horizontal cells.2006

    • Author(s)
      Tanabe, K. et al.
    • Journal Title

      Development 133

      Pages: 4085-4096

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Early segregation of germ somatic lineages during gonadal regeneration in the annelid, Enchytraeus japonensis2006

    • Author(s)
      Tanabe, K. et al.
    • Journal Title

      Current Biology 16

      Pages: 1012-1017

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Level-specific role of paraxial mesoderm in regulation of Tbx5/Tbx4 expression and limb initiation.2006

    • Author(s)
      Saito, D. et al.
    • Journal Title

      Developmental Biology 292

      Pages: 79-89

    • Related Report
      2006 Annual Research Report
  • [Journal Article] 発生過程における細胞移動2006

    • Author(s)
      大畑絵美, 高橋淑子
    • Journal Title

      蛋白質核酸酵素(共立出版) 51

      Pages: 742-746

    • Related Report
      2006 Annual Research Report
  • [Journal Article] からだ作りにみる上皮-間充織転換2006

    • Author(s)
      吉野剛史, 仲矢由紀子, 高橋淑子
    • Journal Title

      実験医学(羊土社) 24

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Epimorphin acts extracellulary to promote cell-sorting and aggregation during the condensation of vertebral cartilage.2006

    • Author(s)
      Oka, Y.et al.
    • Journal Title

      Developmental Biology 291

      Pages: 25-37

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Embryonic stem cells that differentiate into RPE cell precursors in vitro develop into RPE cell monolayers in vivo.2006

    • Author(s)
      Aoki, H. et al.
    • Journal Title

      Experimental Eye Research 82

      Pages: 265-274

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Notch signaling_mediates the_segmental specification of angio blasts in somites and their directed migration toward the dorsal aorta in avian embryos.

    • Author(s)
      Sato, Y.
    • Journal Title

      Developmental Cell (In press)

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Transposon-mediated stable integration and tetracycline-inducible expression of electroporated transgenes in chicken embryos.

    • Author(s)
      Takahashi, Y.
    • Journal Title

      Methods in Avian Embryology 87(in press)

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Somitogenesis as a model to study the formation of morphological boundaries and cell epithelialization.

    • Author(s)
      Takahashi, Y.
    • Journal Title

      Dev. Growth Differ. (in press)

    • NAID

      10024399769

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Tet-on inducible system combined with in ovo electroporation dissects multiple roles of genes in somitogenesis of chicken embryos.

    • Author(s)
      Watanabe, T. et al.
    • Journal Title

      Developmental Biology (in press)

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Stable integration and conditional expression of electroporated transgenes in chicken embryos.

    • Author(s)
      Sato, T. et al.
    • Journal Title

      Developmental Biology (in press)

    • Related Report
      2006 Annual Research Report
  • [Presentation] Neural crest cell migration to the adrenal gland: CXCR4/SDF1 and BMP mediate guidance, while SF-1 is involved in target recognition.2008

    • Author(s)
      Takahashi, Y.
    • Organizer
      NAIST GCOE International Symposium Developmental Biology
    • Place of Presentation
      Nara, Japan
    • Related Report
      2007 Annual Research Report
  • [Presentation] バイオ研究の楽しさとは 〜細胞の挙動を探る〜2007

    • Author(s)
      高橋淑子
    • Organizer
      東洋紡バイオ財団設立25周年記念シンポジウム
    • Place of Presentation
      大阪
    • Year and Date
      2007-06-01
    • Related Report
      2007 Annual Research Report
  • [Presentation] 体作りにおける細胞の社会〜遺伝子から個体まで〜.2007

    • Author(s)
      高橋淑子
    • Organizer
      日本醸造学会大会(招待講演)
    • Place of Presentation
      東京
    • Related Report
      2007 Annual Research Report
  • [Presentation] 血管の発生とパターニング.2007

    • Author(s)
      高橋淑子
    • Organizer
      第5回血液・血管オルビスシンポジウム(特別指名講演)
    • Place of Presentation
      東京
    • Related Report
      2007 Annual Research Report
  • [Presentation] Somitic contribution to the formation of dorsal aorta involves cell migration regulated by Notch and Ephrin.2007

    • Author(s)
      高橋淑子
    • Organizer
      第40回発生生物学会第59回細胞生物学会合同大会
    • Place of Presentation
      福岡
    • Related Report
      2007 Annual Research Report
  • [Presentation] Cell migration and vascular patterning in the body.2007

    • Author(s)
      Takahashi, Y.
    • Organizer
      4th NIBB-EMBL MEETING"CELL & DEVELOPMENTAL BIOLOGY"
    • Place of Presentation
      Okazaki, Japan
    • Related Report
      2007 Annual Research Report
  • [Presentation] STABLE INTEGRATION AND TET-ON INDUCIBLE EXPRESSION OF ELECTROPORATED TRANSGENES IN CHICKEN EMBRYOS.2007

    • Author(s)
      Takahashi, Y.
    • Organizer
      First International Meeting"The Chick as a model organism: genes, development and function"
    • Place of Presentation
      Barcelona, Spain
    • Related Report
      2007 Annual Research Report

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Published: 2006-04-01   Modified: 2016-04-21  

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