Establishment of transgenic reptiles by targeting spermatogonial stem cells

• Project/Area Number: 26650084
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Developmental biology
• Research Institution: Kyoto Prefectural University of Medicine
• Principal Investigator: Nomura Tadashi 京都府立医科大学, 医学(系)研究科(研究院), 准教授 (10323007)
• Co-Investigator(Renkei-kenkyūsha): ONO Katsuhiko 京都府立医科大学, 教養部生物学教室・大学院神経発生生物学, 教授 (30152523) ; GOTOH Hitoshi 京都府立医科大学, 教養部生物学教室・大学院神経発生生物学, 助教 (20462202) ; MURAKAMI Yasunori 愛媛大学, 大学院理工学研究科, 准教授 (50342861)
• Project Period (FY): 2014-04-01 – 2016-03-31
• Project Status: Completed (Fiscal Year 2015)
• Budget Amount: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000); Fiscal Year 2015: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000); Fiscal Year 2014: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: トランスジェニック動物 / 爬虫類 / 精巣 / 電気穿孔法 / 発現ベクター / トランスポゾン / 精子幹細胞 / トランスジェニック / モデル生物 / 遺伝子導入

Outline of Final Research Achievements

The aim of this project is the establishment of transgenic reptiles as a new experimental model of life sciences. To achieve this purpose, we targeted adult testicular stem cells and introduced expression vectors for fluorescent proteins (a linearized RFP expression vector and GFP expression vector with transposon sequences) into the testis of adult Madagascar ground geckoes. At 10 days after in vivo electroporation, we confirmed RFP and GFP expression in the testicular cells in seminiferous tubes. Furthermore, we detected RFP transgene in 1 of 8 embryos by genomic PCR analysis. These data suggest that making transgenic reptiles by targeting adult testicular stem cells is highly feasible, although further technical improvements are required to increase the efficiency of transgenesis.

Research Products

• [Int'l Joint Research] Center for Regenerative Therapy, Dresden(Germany)
• [Journal Article] The evolution of basal progenitors in the developing non-mammalian brain. (2016)
  • Author(s): NOmura, T., Ohtaka-Maruyama, C., Yamashita, W., Wakamatsu, Y., Murakami, Y., Calegari, F., Suzuki, K., Gotoh, H., Ono, K.
  • Journal Title: Development Volume: 143 Issue: 1 Pages: 66-74
  • DOI: 10.1242/dev.127100
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] The neocortex and dorsal ventricular ridge: functional convergence and underlying developmental mechanisms (2016)
  • Author(s): Wataru Yamashita, Tadashi Nomura
  • Journal Title: Brain Evolution by Design Volume: 1
  • Peer Reviewed
• [Journal Article] Genetic manipulation of reptilian embryos: toward an understanding of cortical development and evolution (2015)
  • Author(s): Nomura T, Yamashita W, Gotoh H, Ono K
  • Journal Title: Frontiers in Neuroscience Volume: 9 Pages: 1-11
  • DOI: 10.3389/fnins.2015.00045
  • NAID: 120005947014
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Reconstruction of ancestral brains: Exploring the evolutionary process of encephalization in amniotes. (2014)
  • Author(s): Tadashi Nomura, Yasaunori Murakami, Hiroshi Goto, Katsuhiko Ono.
  • Journal Title: Neurosci Res. Volume: in press Pages: 25-36
  • DOI: 10.1016/j.neures.2014.03.004
  • Peer Reviewed / Open Access
• [Journal Article] 哺乳類大脳皮質の拡大をもたらした分子機構の解明：進化発生学、幹細胞生物学生物学を基盤とした研究戦略の試み (2014)
  • Author(s): 野村真
  • Journal Title: 京都府立医科大学雑誌 Volume: 123 Pages: 607-616
  • Open Access
• [Journal Article] 細胞内エネルギー代謝と生理機能 (2014)
  • Author(s): 後藤仁志、野村真、小野勝彦
  • Journal Title: STUDIA HUMANA et NATURALIA Volume: 48 Pages: 63-70
  • NAID: 120006328746
• [Presentation] Changes in progenitor dynamics during amniote evolution (2015)
  • Author(s): Tadashi Nomura
  • Organizer: The 7th Research Conference of Developmental Neurobiologists in Korea
  • Place of Presentation: 韓国
  • Year and Date: 2015-12-18
  • Int'l Joint Research / Invited
• [Presentation] 胚操作技術を用いた羊膜類大脳皮質進化過程の解明 (2015)
  • Author(s): 野村真
  • Organizer: 細胞を創る会8.0
  • Place of Presentation: 大阪
  • Year and Date: 2015-11-13
  • Invited
• [Presentation] Evolution of cortical structures in mammalian and non-mammalian lineages (2015)
  • Author(s): Tadashi Nomura
  • Organizer: 第38回日本神経科学会
  • Place of Presentation: 神戸
  • Year and Date: 2015-07-28
  • Invited
• [Presentation] Evolution of progenitor dynamics and transcriptional regulation in amniote brains (2015)
  • Author(s): Tadashi Nomura
  • Organizer: 第48回日本発生生物学会大会
  • Place of Presentation: 筑波
  • Year and Date: 2015-06-05
  • Invited
• [Presentation] Convergent evolution of pallial basal progenitors in amniotes (2015)
  • Author(s): Tadashi Nomura, Chiaki Ohtaka-Maruyama, Wataru Yamashita, Yoshio Wakamatsu, Yasunori Murakami, Federico Calegari, Kunihiro Suzuki, HItoshi Gotoh, Katsuhiko Ono
  • Organizer: Cortical Evolution Conference 2015
  • Place of Presentation: Spain
  • Year and Date: 2015-05-18
  • Int'l Joint Research
• [Presentation] Evolution of transcriptional regulations underlying species-specific neuronal diversities in amniotes. (2015)
  • Author(s): Tadashi Nomura, Hitoshi Gotoh, Katsuhiko Ono.
  • Organizer: CDB symposium "Time in Development"
  • Place of Presentation: 神戸
  • Year and Date: 2015-03-23
• [Presentation] Regulation of neural stem cells and brain evolution in amniotes (2014)
  • Author(s): Tadashi Nomura
  • Organizer: Neural Stem Cell in Evolution
  • Place of Presentation: Dresden, Germany
  • Year and Date: 2014-07-08
  • Invited
• [Presentation] Regulation of neural stem cells and brain evolution in amniotes (2014)
  • Author(s): Tadashi Nomura, Federico Calegari, Yasunori Murakami, HItoshi Gotoh, Katsuhiko Ono.
  • Organizer: 47th Annual Meeting of JSDB cosponsored by APDBN
  • Place of Presentation: 名古屋
  • Year and Date: 2014-05-28