Manipulation of chicken primordial germ cells and its application to producing transgenic chickens

• Project/Area Number: 20380156
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Applied animal science
• Research Institution: National Institute of Agrobiological Sciences
• Principal Investigator: NAITO Mitsuru 独立行政法人農業生物資源研究所, 動物発生分化研究ユニット, 上級研究員 (70355733)
• Project Period (FY): 2008 – 2012
• Project Status: Completed (Fiscal Year 2012)
• Budget Amount: ¥18,720,000 (Direct Cost: ¥14,400,000、Indirect Cost: ¥4,320,000); Fiscal Year 2012: ¥2,730,000 (Direct Cost: ¥2,100,000、Indirect Cost: ¥630,000); Fiscal Year 2011: ¥2,730,000 (Direct Cost: ¥2,100,000、Indirect Cost: ¥630,000); Fiscal Year 2010: ¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000); Fiscal Year 2009: ¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000); Fiscal Year 2008: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000)
• Keywords: 始原生殖細胞 / 外来遺伝子 / トランスフェクション / GFP遺伝 / ニワトリ初期胚 / 生殖系列キメラ / 品種識別 / 体外培養 / GFP遺伝子

Research Abstract

In order to develop a technique producing transgenic chickens, we attempted to culture primordial germ cells isolated from early-stage embryos in vitro for long-term. The cultured primordial germ cells retained the ability to migrate to gonads after transfer into recipient embryos and also confirmed the ability of differentiating to functional gametes in recipient gonads. Introduction of GFP gene into the cultured primordial germ cells was performed and cultured primordial germ cells expressing GFP gene were successfully entered gonads of recipient embryos.

Research Products

• [Journal Article] Expression of GFP Gene in Cultured PGCs Isolated from Embryonic Blood and Incorporation into Gonads of Recipient Embryos (2012)
  • Author(s): Naito M, Harumi T and Kuwana T
  • Journal Title: The Journal of Poultry Science Volume: 49 Issue: 2 Pages: 116-123
  • DOI: 10.2141/jpsa.011094
  • NAID: 130004446358
  • ISSN: 1346-7395, 1349-0486
  • Peer Reviewed
• [Journal Article] Spermatogonial stem Cells (2011)
  • Author(s): Naito M
  • Journal Title: Avian Biology Research Volume: 4 Issue: 2 Pages: 66-69
  • DOI: 10.3184/175815511x13069461860174
  • Peer Reviewed
• [Journal Article] In vitro culture of testicular and ovarian gonocytes obtained from 19-day incubated chicken embryos and subsequent colonization into gonads of recipient embryos. (2011)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 48 Pages: 112-118
  • NAID: 10028047602
  • Peer Reviewed
• [Journal Article] Allele-specific PCR typing and sequencing of the mitochondrial D-loop region in four layer breeds (2011)
  • Author(s): Harumi T, Sano A, Minematsu T, Naito M
  • Journal Title: Animal Science Journal Volume: 82 Pages: 223-226
  • NAID: 10030376136
  • Peer Reviewed
• [Journal Article] A new method for isolating viable gonadal germ cells from 7-day-old chick embryos (2011)
  • Author(s): Nakajima Y, Minematsu T, Naito M, Tajima A
  • Journal Title: Journal of Poultry Science e Volume: 48 Pages: 106-111
  • NAID: 10028047579
  • Peer Reviewed
• [Journal Article] In vitro culture of testicular and ovarian gonocytes obtained from 19-day incubated chicken embryos and subsequent colonization into gonads of recipient embryos (2011)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science Volume: 48 Pages: 112-118
  • NAID: 10028047602
  • Peer Reviewed
• [Journal Article] Long Term in vitro Culture of Chicken Primordial Germ Cells Isolated from Embryonic Blood and Incorporation into Germline of Recipient Embryo (2010)
  • Author(s): Naito M, Harumi T and Kuwana T
  • Journal Title: The Journal of Poultry Science Volume: 47 Issue: 1 Pages: 57-64
  • DOI: 10.2141/jpsa.009058
  • NAID: 10026253120
  • ISSN: 1346-7395, 1349-0486
  • Peer Reviewed
• [Journal Article] Long term in vitro culture of chicken primordial germ cells isolated from embryonic blood and incorporation into germline of recipient embryo. (2010)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 47 Pages: 57-64
  • NAID: 10026253120
  • Peer Reviewed
• [Journal Article] A method for cryopreserving semen from Yakido Roosters using N-methylacetamide as a cryoprotective agent. (2010)
  • Author(s): Sasaki K, Tatsumi T, Tsutsui M, Niinomi T, Imai T, Naito M, Tajima A, Nishi Y
  • Journal Title: Journal of Poultry Science Volume: 47 Pages: 297-301
  • NAID: 10027656361
  • Peer Reviewed
• [Journal Article] Long term in vitro culture of chicken primordial germ cells isolated from embryonic blood and incorporation into germline of recipient embryo (2010)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 47 Pages: 57-64
  • NAID: 10026253120
  • Peer Reviewed
• [Journal Article] Morphological characteristics of symmetrically developed ovaries in a 17-day incubated chicken embryo. (2010)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 47 Pages: 85-88
  • NAID: 10026253262
  • Peer Reviewed
• [Journal Article] Preferential Migration of Transferred Primordial Germ Cells to Left Germinal Ridge of Recipient Embryos in Chickens (2009)
  • Author(s): Naito M, Minematsu T, Harumi T and Kuwana T
  • Journal Title: The Journal of Poultry Science Volume: 46 Issue: 1 Pages: 40-45
  • DOI: 10.2141/jpsa.46.40
  • NAID: 10024798951
  • ISSN: 1346-7395, 1349-0486
  • Peer Reviewed
• [Journal Article] Preferential migration of transferred primordial germ cells to left germinal ridge of recipient embryos in chickens. (2009)
  • Author(s): Naito M, Minematsu T, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 46 Pages: 40-45
  • NAID: 10024798951
  • Peer Reviewed
• [Journal Article] Migratory ability of chick primordial germ cells transferred into quail embryos. (2009)
  • Author(s): Ishiguro S, Minematsu T, Naito M, Kanai Y, Tajima A
  • Journal Title: Journal of Reproduction and Development 55 Pages: 183-186
  • NAID: 10024941745
  • Peer Reviewed
• [Journal Article] Preferential migration of transferred primordial germ cells to left germinal ridge of recipient embryos in chickens (2009)
  • Author(s): Naito M, Minematsu T, Harumi T, Kuwana T
  • Journal Title: Journal of Poultry Science 46 Pages: 40-45
  • NAID: 10024798951
  • Peer Reviewed
• [Journal Article] Quantitative genotyping by amplifying the polymorphic sequences of Pre-Melanosomal Protein (PMEL17) gene using real-time polymerase chain reaction in chickens. (2008)
  • Author(s): Minematsu T, Harumi T and Naito M
  • Journal Title: British Poultry Science Volume: 49(5) Issue: 5 Pages: 542-549
  • DOI: 10.1080/00071660802298310
  • Peer Reviewed
• [Journal Article] Germ cell-specific expression of GFP gene induced by chicken vasa homologue (Cvh) promoter in early chicken embryos (2008)
  • Author(s): Minematsu T, Harumi T and Naito M
  • Journal Title: Molecular Reproduction and Development Volume: 75(10) Issue: 10 Pages: 1515-1522
  • DOI: 10.1002/mrd.20894
  • Peer Reviewed
• [Journal Article] Quantitative genotyping by amplifying the polymorphic sequences of Pre-Melanosomal Protein (PMEL17) gene using real-time polymerase chain reaction in chickens. (2008)
  • Author(s): Minematsu T, Harumi T, Naito M
  • Journal Title: British Poultry Science 49 Pages: 542-549
  • Peer Reviewed
• [Journal Article] Germ cell-specific expression of GFP gene induced by chicken vasa homologue (Cvh) promoter in early chicken embryos. (2008)
  • Author(s): Minematsu T, Harumi T, Naito M
  • Journal Title: Molecular Reproduction and Development 75 Pages: 1515-1522
  • Peer Reviewed
• [Journal Article] Germ cell-specific expression of GFP gene induced by chicken vasa homologue (Cvh) promoter in early chicken embryos (2008)
  • Author(s): Minematsu T, Harumi T, Naito M
  • Journal Title: Molecular Reproduction and Development 73 Pages: 1515-1522
  • Peer Reviewed
• [Journal Article] Quantitative genotyping by amplifying the polymorphic sequences of Pre-Melanosomal Protein (PMEL17) gene using real-time polymerase chain reaction in chickens (2008)
  • Author(s): Minematsu T, Harumi T, Naito M
  • Journal Title: British Poultry Science 49 Pages: 542-549
  • Peer Reviewed
• [Presentation] PBS[-]法を用いたニワトリ5および9日胚からの生殖巣生殖細胞(GGCs)の分離 (2012)
  • Author(s): 中島友紀, 水野恵理子, 内藤充, 岡田一男, 秋葉よしえ, 田島淳史
  • Organizer: 日本家禽学会2012年度春季大会
  • Place of Presentation: 名古屋大学東山キャンパス
  • Year and Date: 2012-03-30
• [Presentation] 競合法を用いた始原生殖細胞(PGCs)移住能力評価法の開発 (2012)
  • Author(s): 水野恵理子, 桑名貴, 大沼学, 内藤充, 中島友紀, 田島淳史
  • Organizer: 日本家禽学会2012年度春季大会
  • Place of Presentation: 名古屋大学東山キャンパス
  • Year and Date: 2012-03-30
• [Presentation] ニワトリとウズラのPCRによる識別と生殖系列キメラ胚の作出 (2012)
  • Author(s): 内藤充, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2012年度春季大会
  • Place of Presentation: 名古屋大学東山キャンパス
  • Year and Date: 2012-03-30
• [Presentation] PMEL17遺伝子を利用したニワトリ品種識別と生殖系列キメラ判別への応用 (2011)
  • Author(s): 内藤 充・春海 隆・桑名 貴
  • Organizer: 日本家禽学会2011年度春季大会
  • Place of Presentation: 厚木
  • Year and Date: 2011-03-28
• [Presentation] ニワトリ初期胚生殖巣からのCa2+依存性生殖巣生殖細胞(GGCs)遊離に及ぼすMg2+、培養温度およびEDTAの影響 (2011)
  • Author(s): 中島友紀・峰松健夫・内藤充・田島淳史
  • Organizer: 日本家禽学会2011年度春季大会
  • Place of Presentation: 東京農業大学厚木キャンパス(神奈川県厚木市)
  • Year and Date: 2011-03-28
• [Presentation] PMEL17遺伝子を利用したニワトリ品種識別と生殖系列キメラ判別への応用 (2011)
  • Author(s): 内藤充・春海隆・桑名貴
  • Organizer: 日本家禽学会2011年度春季大会
  • Place of Presentation: 東京農業大学厚木キャンパス(神奈川県厚木市)
  • Year and Date: 2011-03-28
• [Presentation] GFP遺伝子を発現するニワトリ始原生殖細胞の作出とレシピエント胚生殖巣への導入 (2011)
  • Author(s): 内藤 充・春海 隆・桑名 貴
  • Organizer: 日本家禽学会2011年度秋季大会
  • Place of Presentation: 十和田
• [Presentation] GFP遺伝子を発現するニワトリ始原生殖細胞の作出とレシピエント胚生殖巣への導入 (2011)
  • Author(s): 内藤充, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2011年度秋季大会
  • Place of Presentation: 北里大学十和田キャンパス
• [Presentation] 新規分離法を用いて回収されたニワトリ生殖巣生殖細胞(GGCs)の移植による生殖系列キメラ個体の作製 (2011)
  • Author(s): 中島友紀, 峰松健夫, 内藤充, 田島淳史
  • Organizer: 日本畜産学会第114回大会
  • Place of Presentation: 北里大学十和田キャンパス
• [Presentation] Spermatogonial stem cells. (2011)
  • Author(s): 内藤充
  • Organizer: First International Symposium on Propagation of Endangered Species of Birds.
  • Place of Presentation: Abu Dhabi, United Arab Emirates.
• [Presentation] Preferential migration of chicken primordial germ cells to left germinal ridge after transfer into recipient embryos (2011)
  • Author(s): Naito M
  • Organizer: 2nd International Workshop on Preservation of Avian Primordial Germ Cells and Its Usage
  • Place of Presentation: Okinawa, Japan
• [Presentation] A simple recovery of viable gonadal germ cells (GGCs) in domestic chickens (2011)
  • Author(s): Nakajima Y, Minematsu T, Naito M, Tajima A
  • Organizer: 2nd International Workshop on Preservation of Avian Primordial Germ Cells and Its Usage
  • Place of Presentation: Okinawa, Japan
• [Presentation] Spermatogonial stem cells (2011)
  • Author(s): Naito M
  • Organizer: First International Symposium on Propagation of Endangered Species of Birds
  • Place of Presentation: Abu Dhabi, United Arab Emirates
• [Presentation] ニワトリ初期胚血液より採取した始原生殖細胞培養の試み (2010)
  • Author(s): 内藤充・春海隆・桑名貴
  • Organizer: 日本家禽学会2010年度春季大会
  • Place of Presentation: 明治大学駿河台キャンパス、東京都千代田区
  • Year and Date: 2010-03-30
• [Presentation] 孵卵19日目ニワトリ胚より採取した生殖細胞(gonocytes)培養の試み (2010)
  • Author(s): 内藤充, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2010年度春季大会
  • Place of Presentation: 明治大学駿河台キャンパス(東京都千代田区)
  • Year and Date: 2010-03-30
• [Presentation] In vitro culture of chicken primordial germ cells isolated from embryonic blood (2010)
  • Author(s): Naito M, Harumi T and Kuwana T
  • Organizer: XIIIth European Poultry Conference
  • Place of Presentation: Tours, France,
• [Presentation] Basic studies for transgenic chickens. (2010)
  • Author(s): Naito M
  • Organizer: International Workshop on Preservation of Avian Primordial Germ Cells and Its Usage
  • Place of Presentation: Okinawa, Japan,
• [Presentation] In vitro culture of chicken primordial germ cells isolated from embryonic blood. (2010)
  • Author(s): Naito M, Harumi T, Kuwana T.
  • Organizer: XIIIth European Poultry Conference
  • Place of Presentation: Tours, France.
• [Presentation] Basic studies for transgenic chickens. (2010)
  • Author(s): 内藤充.
  • Organizer: International Workshop on Preservation of Avian Primordial Germ Cells and Its Usage.
  • Place of Presentation: Okinawa, Japan.
• [Presentation] Morphological characteristics of symmetrically developed ovaries in a 17-day incubated chicken embryo (2010)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Organizer: XIIIth European Poultry Conference
  • Place of Presentation: Tours, France
• [Presentation] In vitro culture of chicken primordial germ cells isolated from embryonic blood (2010)
  • Author(s): Naito M, Harumi T, Kuwana T
  • Organizer: XIIIth European Poultry Conference
  • Place of Presentation: Tours, France
• [Presentation] Basic studies for transgenic chickens (2010)
  • Author(s): Naito M
  • Organizer: International Workshop on Preservation of Avian Primordial Germ Cells and Its Usage
  • Place of Presentation: Okinawa, Japan
• [Presentation] ウズラ胚に移植されたニワトリPGCsの移住性 (2009)
  • Author(s): 石黒進, 峰松健夫, 内藤充, 金井幸雄, 田島淳史
  • Organizer: 日本家禽学会2009年度春季大会
  • Place of Presentation: 日本大学生物資源科学部(神奈川県藤沢市)
  • Year and Date: 2009-03-28
• [Presentation] ニワトリ初期胚血液より採取した始原生殖細胞培養の試み (2009)
  • Author(s): 内藤充, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2009年度春季大会
  • Place of Presentation: 日本大学生物資源科学部(神奈川県藤沢市)
  • Year and Date: 2009-03-28
• [Presentation] 始原生殖細胞を利用した鳥類の生殖制御法 (2009)
  • Author(s): 内藤 充
  • Organizer: 第102回日本繁殖生物学会大会
  • Place of Presentation: 奈良
• [Presentation] 始原生殖細胞を利用した鳥類の生殖制御法 (2009)
  • Author(s): 内藤充
  • Organizer: 第102回日本繁殖生物学会大会
  • Place of Presentation: 近畿大学農学部、奈良県奈良市
• [Presentation] 始原生殖細胞を利用した鳥類の生殖制御法 (2009)
  • Author(s): 内藤充
  • Organizer: 第102回日本繁殖生物学会大会
  • Place of Presentation: 近畿大学農学部(奈良県奈良市)
• [Presentation] 左右対称に発達した孵卵17日目ニワトリ胚卵巣の形態的特徴 (2009)
  • Author(s): 内藤充, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2009年度秋季大会
  • Place of Presentation: 琉球大学千原キャンパス(沖縄県西原町)
• [Presentation] ニワトリ始原生殖細胞の左右生殖隆起への移住頻度について (2008)
  • Author(s): 内藤充, 峰松健夫, 春海隆, 桑名貴
  • Organizer: 日本家禽学会2008年度秋季大会
  • Place of Presentation: 北里大学獣医学部(青森県十和田市)
  • Year and Date: 2008-08-28
• [Presentation] Allele-specific PCR detection of single nucleotide polymorphisms in the chicken mitochondrial D-loop region (2008)
  • Author(s): Harumi T, Minematsu T, Naito M
  • Organizer: 23rd World's Poultry Congress
  • Place of Presentation: Brisbane, Australia
• [Presentation] Gonocytes obtained from testes or ovary of 20-day incubated chicken embryos contribute to germline lineage after transfer into bloodstream of recipient embryos (2008)
  • Author(s): Naito M, Minematsu T, Harumi T, Kuwana T
  • Organizer: 23rd World's Poultry Congress
  • Place of Presentation: Brisbane, Australia
• [Presentation] 生殖系列キメラにおける生殖巣生殖細胞（GGCs）の移住・増殖能力の評価
  • Author(s): 水野恵理子・大沼学・内藤充・中島友紀・田島淳史
  • Organizer: 日本家禽学会2013年度春季大会
  • Place of Presentation: 安田女子大学
• [Presentation] 希少鳥類種での始原生殖細胞培養技術の検討
  • Author(s): 桑名貴・内藤充
  • Organizer: 日本家禽学会2013年度春季大会
  • Place of Presentation: 安田女子大学
• [Book] 最新畜産ハンドブック（分担執筆） (2013)
  • Author(s): 内藤 充
  • Total Pages: 650
  • Publisher: 講談社
• [Book] 畜産技術発達史 12.鶏の人工孵化と雌雄鑑別技術(分担執筆)(畜産技術協会)(東京) (2011)
  • Author(s): 内藤充
• [Book] 畜産技術発達史 35.採卵鶏の光線管理技術(分担執筆)(畜産技術協会)(東京) (2011)
  • Author(s): 内藤充
• [Book] 畜産技術発達史 12.鶏の人工孵化と雌雄鑑別技術 (2011)
  • Author(s): 内藤充
  • Publisher: 畜産技術協会
• [Book] 畜産技術発達史 35.採卵鶏の光線管理技術 (2011)
  • Author(s): 内藤充
  • Publisher: 畜産技術協会
• [Book] 発生工学日本農学80年史(分担) (2009)
  • Author(s): 内藤 充
  • Publisher: 養賢堂
• [Book] 日本農学80年史第33章家禽学 3.発生工学 (分担執筆) (2009)
  • Author(s): 内藤 充
  • Total Pages: 238
  • Publisher: 養賢堂
• [Book] 日本農学80年史 第33章家禽学(3)発生工学(分担執筆) (2009)
  • Author(s): 内藤充
  • Total Pages: 376
  • Publisher: 養賢堂