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Recombinant protein production using Daphnia magna eggs

Research Project

Project/Area Number 17K19236
Research Category

Grant-in-Aid for Challenging Research (Exploratory)

Allocation TypeMulti-year Fund
Research Field Agricultural chemistry and related fields
Research InstitutionOsaka University

Principal Investigator

Watanabe Hajime  大阪大学, 工学研究科, 教授 (80212322)

Project Period (FY) 2017-06-30 – 2020-03-31
Project Status Completed (Fiscal Year 2019)
Budget Amount *help
¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000)
Fiscal Year 2018: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Fiscal Year 2017: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Keywordsミジンコ / 組換えタンパク質 / 遺伝子編集 / 蛍光タンパク質 / ビテロジェニン / GFPタンパク質 / 緑色蛍光タンパク質 / CRISPR/Cas9 / トランスジェニック / 卵黄タンパク / ゲノム編集 / タンパク産生
Outline of Final Research Achievements

In the present study, we aimed to develop a system for accumulation of useful protein in place of Daphnia magna's yolk protein by using gene editing technology.
(1) We generated a plasmid containing a secreted GFP fusion gene, in which a secreted signal peptide and GFP were fused, and confirmed that the expressed protein was secreted into the extracellular space.
(2) Furthermore, the vitellogenin receptor binding domain was fused with the gene. This gene was injected into eggs by microinjection the accumulation of GFP fused protein in the eggs was confirmed by fluorescence microscopy.

Academic Significance and Societal Importance of the Research Achievements

抗体やワクチンを始めとする様々な機能性タンパク質のニーズが高まっているが、通常は 培養細胞を用いた系が主流であり、生産の手間とコスト、さらに血清を用いる場合にはそのリスクも問題となっている。一方でバクテリア、カイコ、ニワトリや植物に目的タンパク質を産生させる試みも進められているが、バクテリアでは分子量の大きなタンパク質の産生に問題があり、その他の生物種でも手間と時間をはじめ種々の問題が存在している。こうした中で、ミジンコが安価な物質生産のツールとして利用できる可能性を示したことで、新たに安価、かつ持続可能な組換えタンパク質産生法が選択肢に加わったことになる。

Report

(4 results)
  • 2019 Annual Research Report   Final Research Report ( PDF )
  • 2018 Research-status Report
  • 2017 Research-status Report
  • Research Products

    (58 results)

All 2020 2019 2018 2017 Other

All Int'l Joint Research (2 results) Journal Article (14 results) (of which Int'l Joint Research: 5 results,  Peer Reviewed: 14 results,  Open Access: 11 results) Presentation (39 results) (of which Int'l Joint Research: 6 results,  Invited: 6 results) Remarks (3 results)

  • [Int'l Joint Research] ルーヴェンカトリック大学(ベルギー)

    • Related Report
      2019 Annual Research Report
  • [Int'l Joint Research] 環境影響評価研究所(スペイン)

    • Related Report
      2019 Annual Research Report
  • [Journal Article] In vitro synthesis of the human calcium transporter Letm1 within cell-sized liposomes and investigation of its lipid dependency2019

    • Author(s)
      Okamura Kosuke、Matsushita Shuhei、Kato Yasuhiko、Watanabe Hajime、Matsui Aiko、Oka Toshihiko、Matsuura Tomoaki
    • Journal Title

      Journal of Bioscience and Bioengineering

      Volume: 127 Issue: 5 Pages: 544-548

    • DOI

      10.1016/j.jbiosc.2018.11.003

    • NAID

      40021891579

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Parasite‐mediated selection in a natural metapopulation of Daphnia magna2019

    • Author(s)
      Cabalzar Andrea P.、Fields Peter D.、Kato Yasuhiko、Watanabe Hajime、Ebert Dieter
    • Journal Title

      Molecular Ecology

      Volume: 28 Issue: 21 Pages: 4770-4785

    • DOI

      10.1111/mec.15260

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Class?III Polyphosphate Kinase?2 Enzymes Catalyze the Pyrophosphorylation of Adenosine‐5′‐Monophosphate2019

    • Author(s)
      Ogawa Marin、Uyeda Atsuko、Harada Kazuo、Sato Yu、Kato Yasuhiko、Watanabe Hajime、Honda Kohsuke、Matsuura Tomoaki
    • Journal Title

      ChemBioChem

      Volume: 20 Issue: 23 Pages: 2961-2967

    • DOI

      10.1002/cbic.201900303

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Atrazine exposed phytoplankton causes the production of non-viable offspring on Daphnia magna2019

    • Author(s)
      Religia Pijar、Kato Yasuhiko、Fukushima Ery Odette、Matsuura Tomoaki、Muranaka Toshiya、Watanabe Hajime
    • Journal Title

      Marine Environmental Research

      Volume: 145 Pages: 177-183

    • DOI

      10.1016/j.marenvres.2019.02.007

    • Related Report
      2018 Research-status Report
    • Peer Reviewed
  • [Journal Article] Tryptophan hydroxylase (TRH) loss of function mutations induce growth and behavioral defects in Daphnia magna2018

    • Author(s)
      Rivetti Claudia、Campos Bruno、Pina Benjamin、Raldua Demetrio、Kato Yasuhiko、Watanabe Hajime、Barata Carlos
    • Journal Title

      Scientific Reports

      Volume: 8 Issue: 1 Pages: 1518-1518

    • DOI

      10.1038/s41598-018-19778-0

    • Related Report
      2018 Research-status Report 2017 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Microbiota inoculum composition affects holobiont assembly and host growth in Daphnia2018

    • Author(s)
      Martijn Callens、Hajime Watanabe、Yasuhiko Kato、Jun Miura、Ellen Decaestecker
    • Journal Title

      Microbiome

      Volume: 6 Issue: 1 Pages: 56-56

    • DOI

      10.1186/s40168-018-0444-1

    • Related Report
      2018 Research-status Report 2017 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Genomic integration and ligand-dependent activation of the human estrogen receptor α in the crustacean Daphnia magna2018

    • Author(s)
      Toerner Kerstin、Nakanishi Tsuyoshi、Matsuura Tomoaki、Kato Yasuhiko、Watanabe Hajime
    • Journal Title

      PLOS ONE

      Volume: 13 Issue: 6 Pages: e0198023-e0198023

    • DOI

      10.1371/journal.pone.0198023

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Generation of white-eyed Daphnia magna mutants lacking scarlet function2018

    • Author(s)
      Ismail Nur Izzatur Binti、Kato Yasuhiko、Matsuura Tomoaki、Watanabe Hajime
    • Journal Title

      PLOS ONE

      Volume: 13 Issue: 11 Pages: e0205609-e0205609

    • DOI

      10.1371/journal.pone.0205609

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] A 5′ UTR-Overlapping LncRNA Activates the Male-Determining Gene doublesex1 in the Crustacean Daphnia magna2018

    • Author(s)
      Kato Yasuhiko、Perez Christelle Alexa G.、Mohamad Ishak Nur Syafiqah、Nong Quang D.、Sudo Yuumi、Matsuura Tomoaki、Wada Tadashi、Watanabe Hajime
    • Journal Title

      Current Biology

      Volume: 28 Issue: 11 Pages: 1811-1817

    • DOI

      10.1016/j.cub.2018.04.029

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Monitoring ecdysteroid activities using genetically encoded reporter gene in Daphnia magna2018

    • Author(s)
      Adhitama Nikko、Matsuura Tomoaki、Kato Yasuhiko、Watanabe Hajime
    • Journal Title

      Marine Environmental Research

      Volume: 140 Pages: 375-381

    • DOI

      10.1016/j.marenvres.2018.07.003

    • Related Report
      2018 Research-status Report
    • Peer Reviewed
  • [Journal Article] A murine model of atopic dermatitis can be generated by painting the dorsal skin with hapten twice 14 days apart2018

    • Author(s)
      Kitamura Ayaka、Takata Ryohei、Aizawa Shin、Watanabe Hajime、Wada Tadashi
    • Journal Title

      Scientific Reports

      Volume: 8 Issue: 1 Pages: 5988-5988

    • DOI

      10.1038/s41598-018-24363-6

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] CRISPR/Cas-mediated knock-in via non-homologous end-joining in the crustacean Daphnia magna2017

    • Author(s)
      Kumagai Hitoshi、Nakanishi Takashi、Matsuura Tomoaki、Kato Yasuhiko、Watanabe Hajime
    • Journal Title

      PLOS ONE

      Volume: 12 Issue: 10 Pages: e0186112-e0186112

    • DOI

      10.1371/journal.pone.0186112

    • Related Report
      2017 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Development of a bicistronic expression system in the branchiopod crustacean Daphnia magna2017

    • Author(s)
      Kumagai Hitoshi、Matsuura Tomoaki、Kato Yasuhiko、Watanabe Hajime
    • Journal Title

      genesis

      Volume: 55 Issue: 12

    • DOI

      10.1002/dvg.23083

    • Related Report
      2017 Research-status Report
    • Peer Reviewed
  • [Journal Article] Mapping the expression of the sex determining factor Doublesex1 in Daphnia magna using a knock-in reporter2017

    • Author(s)
      Nong Quang Dang、Mohamad Ishak Nur Syafiqah、Matsuura Tomoaki、Kato Yasuhiko、Watanabe Hajime
    • Journal Title

      Scientific Reports

      Volume: 7 Issue: 1 Pages: 13521-13521

    • DOI

      10.1038/s41598-017-13730-4

    • Related Report
      2017 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Presentation] Molecular mechanism of adaptive sex ratio adjustment in the water flea Daphnia magna2020

    • Author(s)
      Yasuhiko Kato, Quang Dang Nong, Christelle Alexa Garcia Perez, Nur Syafiqah Mohamad Ishak, Hajime Watanabe
    • Organizer
      EMBO | EMBL Symposia The Organism and its Environment
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Monitoring of hormonal activities in the water flea Daphnia magna using genome editing tools2020

    • Author(s)
      Yasuhiko Kato, Ayano Noda, Yudai Yamamoto, Yuto Yamazaki, Nikko Adhitama, Hajime Watanabe
    • Organizer
      EMBO | EMBL Symposia The Organism and its Environment
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research
  • [Presentation] 生殖戦略を決定する化学コミュニケーションの解明2019

    • Author(s)
      西上旺希、加藤泰彦、渡邉肇
    • Organizer
      新学術領域研究「化学コミュニケーションのフロンティア」第5回公開シンポジウム
    • Related Report
      2019 Annual Research Report
  • [Presentation] オオミジンコにおける長鎖ノンコーディングRNAによるdoublesex1遺伝子の活性化2019

    • Author(s)
      加藤 泰彦,Perez Christelle Alexa,Nong Quang,Mohamad Ishak Nur Syafiqah,和田 忠志,渡邉 肇
    • Organizer
      日本動物学会 第90回 大阪大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] Atrazine exposed phytoplankton causes the production of non-viable offspring on Daphnia magna.2019

    • Author(s)
      Pijar Religia, Yasuhiko Kato, Ery Odette Fukushima, Tomoaki Matsuura, Toshiya Muranaka, Hajime Watanabe
    • Organizer
      日本動物学会 第90回 大阪大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] 環境応答により雌雄を産み分けるミジンコの性スペクトラム2019

    • Author(s)
      加藤 泰彦,Perez Christelle Alexa,Nong Quang,渡邉 肇
    • Organizer
      新学術領域研究「性スペクトラム _連続する表現型としての雌雄」第3回領域会議
    • Related Report
      2019 Annual Research Report
  • [Presentation] 遺伝子組み換え技術を応用したオオミジンコにおける幼若ホルモン活性の可視化2019

    • Author(s)
      野田彩乃、加藤泰彦、松浦友亮、渡邉肇
    • Organizer
      第71回日本生物工学会大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] ビテロジェニン輸送経路を利用した組換えタンパク質のミジンコ卵への蓄積系2019

    • Author(s)
      辻 勇祐, 加藤 泰彦, 渡邉 肇
    • Organizer
      第42回日本分子生物学会年会
    • Related Report
      2019 Annual Research Report
  • [Presentation] Development of a reporter system to visualize juvenile hormone activity in Daphnia magna2019

    • Author(s)
      野田彩乃、加藤泰彦、松浦友亮、渡邉肇
    • Organizer
      第42回日本分子生物学会年会
    • Related Report
      2019 Annual Research Report
  • [Presentation] ミジンコにおける生存戦略を決定する化学コミュニケーションの解明2019

    • Author(s)
      西上旺希、加藤泰彦、渡邉肇
    • Organizer
      「化学コミュニケーションのフロンティア」 第 6 回公開シンポジウム
    • Related Report
      2019 Annual Research Report
  • [Presentation] ミジンコにおけるオス決定因子Dsx1発現細胞のトランスクリプトーム解析2019

    • Author(s)
      加藤泰彦, Quang Dang Nong, 渡邉肇
    • Organizer
      「先進ゲノム支援」2019年度拡大班会議
    • Related Report
      2019 Annual Research Report
  • [Presentation] ミジンコにおける性特異的な長鎖ノンコーディングRNAの探索2019

    • Author(s)
      加藤泰彦, Christelle Alexa Gracia Perez, 渡邉肇
    • Organizer
      「先進ゲノム支援」2019年度拡大班会議
    • Related Report
      2019 Annual Research Report
  • [Presentation] 生殖戦略を決定する化学コミュニケーションの解明2018

    • Author(s)
      渡邉肇
    • Organizer
      新学術領域研究「化学コミュニケーションのフロンティア」第3回公開シンポジウム
    • Related Report
      2018 Research-status Report
  • [Presentation] A 5´UTR-overlapping LncRNA Activates the Male-determining Gene doublesex1 in the Crustacean Daphnia magna2018

    • Author(s)
      Yasuhiko Kato, Christelle Alexa G. Pelez, Nur Syafiqah Mohamad Ishak, Quang D. Nong, Tomoaki Matsuura, Tadashi Wada, Hajime Watanabe
    • Organizer
      第 20 回 日本 RNA 学会年会
    • Related Report
      2018 Research-status Report
  • [Presentation] オオミジンコにおける毒性影響可視化のための遺伝子工学的手法の開発2018

    • Author(s)
      渡邉肇
    • Organizer
      第45回日本毒性学会学術年会
    • Related Report
      2018 Research-status Report
  • [Presentation] Monitoring of chemical and metal contaminants using engineered Daphnia magna2018

    • Author(s)
      Yasuhiko Kato, Hajime Watanabe
    • Organizer
      Second Interdisciplinary and Research Alumni Symposium iJaDe2018
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research
  • [Presentation] オオミジンコにおける内分泌かく乱物質検出のためのGAL4/UASシステムの確立2018

    • Author(s)
      加藤泰彦、山崎勇人、松浦友亮、渡邉肇
    • Organizer
      第70回日本生物工学会大会
    • Related Report
      2018 Research-status Report
  • [Presentation] Visualization of ecdysteroid activities using genetically encoded reporter gene in Daphnia magna2018

    • Author(s)
      Nikko Adhitama, Yasuhiko Kato, Tomoaki Matsuura, Hajime Watanabe
    • Organizer
      第70回日本生物工学会大会
    • Related Report
      2018 Research-status Report
  • [Presentation] 気液界面培養下の気道上皮細胞への排ガス曝露影響評価法の検討-(1)送気流量の検討-2018

    • Author(s)
      伊藤剛、村木直美、田村久美子、利根川義男、石井幸雄、酒井康行、渡邉肇、高野裕久
    • Organizer
      第59回大気環境学会年会
    • Related Report
      2018 Research-status Report
  • [Presentation] 気液界面培養下の気道上皮細胞への排ガス曝露影響評価法の検討-(2)線維芽細胞共培養の有効性の検討-2018

    • Author(s)
      村木直美、伊藤剛、田村久美子、利根川義男、石井幸雄、酒井康行、渡邉肇、高野裕久
    • Organizer
      第59回大気環境学会年会
    • Related Report
      2018 Research-status Report
  • [Presentation] 再構成型無細胞翻訳系を用いたGタンパク質共役型受容体の安定化変異体の取得2018

    • Author(s)
      仲井宏紀、一色衣香、渡邉肇、村田武士、松浦友亮
    • Organizer
      「細胞を創る」研究会11.0
    • Related Report
      2018 Research-status Report
  • [Presentation] In vitro evolution of E. coli multidrug efflux transporter EmrE by using liposome display2018

    • Author(s)
      内田紗衣、植田淳子、渡邊肇、松浦友亮
    • Organizer
      「細胞を創る」研究会11.0
    • Related Report
      2018 Research-status Report
  • [Presentation] オオミジンコの遺伝子操作法の開発と応用2018

    • Author(s)
      加藤泰彦、志賀靖弘、時下進一、山形秀夫、渡邉肇
    • Organizer
      東京薬科大学生命科学部25周年記念シンポジウム
    • Related Report
      2018 Research-status Report
  • [Presentation] 環境応答により雌雄を産み分けるミジンコの性スペクトラム2018

    • Author(s)
      Yasuhiko Kato、Christelle Alexa G. Perez、Quang D. Nong、Hajime Watanabe
    • Organizer
      新学術領域研究 性スペクトラム 第二回 領域班会議
    • Related Report
      2018 Research-status Report
  • [Presentation] 多剤排出トランスポーターEmrEを検出素子とした細胞毒性化合物検出技術の開発2018

    • Author(s)
      服部将人, 渡邉肇, 松浦友亮
    • Organizer
      第13回無細胞生命科学研究会
    • Related Report
      2018 Research-status Report
  • [Presentation] A 5' UTR-Overlapping LncRNA Activates the Male- Determining Gene doublesex1 in the Crustacean Daphnia magna2018

    • Author(s)
      Yasuhiko Kato、Christelle Alexa G. Perez、Quang D. Nong、Hajime Watanabe
    • Organizer
      JAJ RNA 2018
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research / Invited
  • [Presentation] オオミジンコにおけるHSP70プロモーターを利用した熱誘導型遺伝子発現系の開発2018

    • Author(s)
      加藤泰彦、清水里奈、渡邉肇
    • Organizer
      第41回日本分子生物学会年会
    • Related Report
      2018 Research-status Report
  • [Presentation] 環境応答により雌雄を産み分けるミジンコの性スペクトラム2018

    • Author(s)
      加藤泰彦、Quang Dang Nong、渡邉肇
    • Organizer
      「先進ゲノム支援」2018年度拡大班会議
    • Related Report
      2018 Research-status Report
  • [Presentation] Alterations in the activity of the sex determination factor dsx1 resulted in different intersex phenotypes in Daphnia magna2018

    • Author(s)
      Quang Dang Nong , Yasuhiko Kato , Hajime Watanabe
    • Organizer
      「先進ゲノム支援」2018年度拡大班会議
    • Related Report
      2018 Research-status Report
  • [Presentation] Possibility of chemical communication among Daphnia individuals for reproductive strategy2018

    • Author(s)
      Yasuhiko Kato, Hajime Watanabe
    • Organizer
      Frontier Research on Chemical Communications (ISCC2019)
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research
  • [Presentation] 生態学と分子生物学の接点としてのミジンコ2018

    • Author(s)
      渡邉肇
    • Organizer
      日本生態学会 第66回大会
    • Related Report
      2018 Research-status Report
    • Invited
  • [Presentation] オオミジンコの周期性単為生殖における環境依存的な性決定メカニズム2017

    • Author(s)
      加藤泰彦、Quang Dang Nong、Nur Syafiqah Mohamad Ishak、渡邉肇
    • Organizer
      日本動物学会第88回大会
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] ミジンコの環境依存的な性決定を制御する長鎖非コードRNA2017

    • Author(s)
      加藤泰彦、Christelle Alexa Pelez、Nong Dang Quang、渡邉肇
    • Organizer
      2017年度生命科学系学会合同年次大会(招待講演)
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] オオミジンコにおけるCas9タンパク質を用いた高効率な遺伝子破壊法と遺伝子導入法の開発2017

    • Author(s)
      熊谷仁志、中西貴士、松浦友亮、加藤泰彦、渡邉肇
    • Organizer
      第2回ゲノム編集学会
    • Related Report
      2017 Research-status Report
  • [Presentation] オオミジンコにおける眼の色素遺伝子 scarletの遺伝子組み換えマーカーとしての検討2017

    • Author(s)
      Nur Izzatur Binti Ismail、加藤泰彦、松浦友亮、渡邉肇
    • Organizer
      第2回ゲノム編集学会
    • Related Report
      2017 Research-status Report
  • [Presentation] TALEN を利用した非相同末端結合によるオオミジンコゲノムへのエストロゲンバイオセンサーの導入2017

    • Author(s)
      Torner Kerstin、加藤泰彦、渡邉肇
    • Organizer
      第2回ゲノム編集学会
    • Related Report
      2017 Research-status Report
  • [Presentation] 重金属曝露を可視化したミジンコの作製2017

    • Author(s)
      荒尾拓斗、加藤泰彦、渡邉肇
    • Organizer
      第44回日本毒性学会学術年会
    • Related Report
      2017 Research-status Report
  • [Presentation] ゲノム編集技術によるミジンコの性決定機構の解析2017

    • Author(s)
      加藤泰彦、渡邉肇
    • Organizer
      日本動物学会 第88回 富山大会
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] Genome editing for visualization of water pollutants using water flea2017

    • Author(s)
      Yasuhiko Kato, Hajime Watanabe
    • Organizer
      19th International Symposium on Pollutant Responses in Marine Organisms
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research / Invited
  • [Remarks] 大阪大学工学研究科生物工学専攻渡邉研究室

    • URL

      http://www.bio.eng.osaka-u.ac.jp/ez/index.html

    • Related Report
      2019 Annual Research Report
  • [Remarks] 大阪大学工学研究科生命先端工学専攻渡邉研究室

    • URL

      http://www.bio.eng.osaka-u.ac.jp/ez/

    • Related Report
      2018 Research-status Report
  • [Remarks] 大阪大学 工学研究科 生命環境システム工学

    • URL

      http://www.bio.eng.osaka-u.ac.jp/ez/index.html

    • Related Report
      2017 Research-status Report

URL: 

Published: 2017-07-21   Modified: 2021-02-19  

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