Development of Avian Transgenic System

禽类转基因系统的开发

• 批准号: 04554029
• 负责人: EGUCHI Goro
• 金额: $ 13.63万
• 依托单位: National Institute for Basic Biology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-04554029/
• 关键词: avian chimera; transgenesis; gene transfer; complete culture of avian futilizel eggs; chicken; quail; 形質転換; トランスジェニック・システム; 受精卵完全体外培養; 形質転換系; 鳥類胚完全体外培養

This project started from 1992 in order to establish the avian transgenic system which has been required by researchers in the field of gene expression and regulation in animal in situ, ant the following results have been established through studies conducted for three fiscal years from 1992 to 1994.1.In addition to the complete in vitro culture technique for chicken fertilized eggs, the similar technique for quail fertilized eggs has been established. This technique can give rize to in vitro development of fertilized eggs to adults with more than 25% of efficiency. 2.Using this complete in vitro culture technique for avian fertilized eggs, chimeric chickens with chimeric gonads have been produced by introducing blastoderm cells to other embryos at blastoderm stage. In addition, we have succeeded in producing chimeric F1 chickens. 3.It is highly difficult to observe egg nucleus of avian eggs because of their size and optical density owing to yolk. By combining TV-camera and specifically arranged illumination system, improved method to detect nuclear area of avian fertilized eggs has been established, and now micro-injection of genes to be transfected to in the nuclear area has become possible. 4.To improve the efficiency of recombination of transfected genes to the host genom, it has been repeatedly tried to co-inject the gene recombined with retroviral vector together with the integrase derived from retrovirus, and prospective results have been obtained.Thus the fundamental basis required for establishement of avian transgenesis system has been obtained through this project.

本课题从1992年开始，旨在建立研究者在动物体内原位基因表达和调控领域所需要的禽类转基因系统，经过1992 ~ 1994年三个财政年度的研究，取得了如下成果：1.除了完整的鸡受精卵体外培养技术外，鹌鹑受精卵的类似技术已经建立。该技术可使受精卵体外发育为里泽，有效率达25%以上。2.利用这种完整的鸡受精卵体外培养技术，将胚盘细胞导入胚盘期的其他胚胎中，获得了具有嵌合性腺的嵌合鸡。此外，我们还成功地生产了嵌合F1鸡。3.禽蛋的卵核由于卵黄的存在，其大小和光密度都很大，很难观察到。将电视摄像机与特定的照明系统相结合，建立了检测鸡受精卵细胞核面积的改进方法，使向细胞核内显微注射基因成为可能。4.为了提高转染基因与宿主基因组的重组效率，将重组于逆转录病毒载体的基因与逆转录病毒来源的整合酶共注射进行了多次尝试，取得了预期的效果，为建立禽类转基因系统奠定了基础。

项目成果

Ono,T.,Muto,S.,Mizutani,M.,Agata,K.,Mochii,M.,Kino,K.,Otsuka,K.,Ohta,M.,Yoshida,M.,Eguchi,G.: "Production of quail chimera by transfer of early blastodermal cells and its use for transgenesis" Japanese Poultry Science. 31. 119-129 (1994)

小野，T.，武藤，S.，水谷，M.，阿加塔，K.，持井，M.，木野，K.，大冢，K.，太田，M.，吉田，M.，江口，G.：

G.Eguchi & R.Kodama: "Transdifferentiation" Current Opinion in Cell Biology. 5. 1023-1028 (1993)

江口

Ono.T.,Murakami,T.,Mochii,M.,Agata,K.,Kino,K.,Otsuka,K.,Ohta,M.,Mizutani,M.,Yoshida,M.,Eguchi,G.: "A Complete culture system for avian transgenesis,supporting quail embryos from the single-cell stage to hatching" Developmental Biology. 161. 126-130 (1994)

小野T.、村上T.、持井M.、阿加塔K.、木野K.、大冢K.、太田M.、水谷M.、吉田M.、江口G.：

K.Agata,H.Kobayasgi,G.Eguchi et al.: "Characterization of the multipotent dedifferentiated state of pigmented epithelial cells in vitro" Development. 118. 1025-1030 (1993)

K.Agata、H.Kobayasgi、G.Eguchi 等人：“体外色素上皮细胞多能去分化状态的表征”开发。

Ono, T., Muto, S., Mizutani, M., Agata, K., Mochii, M., Kino, K., Otsuka, K., Ohta, M., Yoshida, M., Eguchi, G.: "Production of quail chimera by transfer of early blastodermal cells and its use for transgenesis" Japanese Poultry Science. 31. 119-129 (1994

小野，T.，武藤，S.，水谷，M.，阿加塔，K.，持井，M.，基诺，K.，大冢，K.，太田，M.，吉田，M.，江口，G.：