Grant-in-Aid for Scientific Research (A)
|Allocation Type||Single-year Grants|
Environmental physiology (including Physical medicine and Nutritional physiology)
|Research Institution||Hokkaido University|
HONMA Kenichi Hokkaido Uni.Sch.of Med., Professor, 医学部, 教授 (40113625)
ISHIURA Masahiro Nagoya Uni.Sch of Sci.Associ.Professor, 理学部, 助教授 (20132730)
KONDO Takao Nagoya Uni.Sch of Sci.Professor, 理学部, 教授 (10124223)
SHINOHARA Kazuyuki Yokohama Uni.Sch of Med., Instructor, 医学部, 助手 (30226154)
HONMA Sato Hokkaido Uni.Sch.of Med., Associ.Professor, 医学部, 助教授 (20142713)
安倍 博 北海道大学, 医学部, 助手 (80201896)
勝野 由美子 北海道大学, 医学部, 助手 (80177419)
|Project Period (FY)
1994 – 1996
Completed(Fiscal Year 1996)
|Budget Amount *help
¥11,700,000 (Direct Cost : ¥11,700,000)
Fiscal Year 1996 : ¥1,700,000 (Direct Cost : ¥1,700,000)
Fiscal Year 1995 : ¥2,500,000 (Direct Cost : ¥2,500,000)
Fiscal Year 1994 : ¥7,500,000 (Direct Cost : ¥7,500,000)
|Keywords||Luciferase / Bio-luminescence / Ca ion / Aequorin / NIH3T3 cell / Reporter gene / Gene expression / real-time monitoring / Aequorin / 発光蛋白質 / Caイオン / トランスジェニックマウス / リアルタイム計測 / 遺伝子導入 / ルシフェラーゼ / 神経ペプチド / 細胞培養 / サーカディアンリズム / 形質転換 / トランスフォーマント / フォトカウント|
1) Construction of vector containing a gene for bio-luminescence
Luciferase bio-luminescence system : Luciferase gene was inserted into pBact : STneoB vector at the downstream of Actin promotor, and tansfected to mouse fibroblastoma cells. Aequorin bio-luminescence system : Aequorin gene was inserted into PRK7 vector at the downstream of CMV promotor, and transfected to NIH3T3 cells.
2) Development of high-sensitive detector for bio-luminescence
Bio-luminescence was detected by photomultiplier. In the signal transduction unit, a photon conuter system was introduced. By this method, the sensitivity of the whole system was increased by 10 to 50 times.
3) Development of cell culture system
The disperse and organotypic cell culture systems were developed for analysing the function of rat suprachiasmatic nucleus. The circadian rhythm was detected in AVP and VIP secretions in the culture medium for several cycles.
4) Transfestion to cultured cells
Transfection to NIH3T3 cell was successfully done. However, that to the neurons was somewhat difficult. Further studies are needed.
5) Gene expression and bio-luminescence
NIH3T3 cells containing aequorin gene showed bio-luminescence when stimulated by KCl. Gene expression was successfully detected by the photometric method.