| Project/Area Number |
09660295
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Applied animal science
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
TAKAHASHI Yoshiyuki Graduate School of Veterinary Medicine, Hokkaido University, Professor, 大学院・獣医学研究科, 教授 (70167485)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1998: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1997: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | cattle / oocytes / in vitro fertilization / in vitro culture / synthetic oviduct fluid medium / 合成卵管液 |
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| Research Abstract |
This study was performed to establish a reliable system for the in vitro production of bovine embryos using in vitro matured, fertilized and cultured ovarian oocytes. Factors that affect the in vitro fertilization and subsequent embryonic development were investigated using the synthetic oviduct fluid (SOF) as a basic culture medium. Results revealed that reducing the oxygen concentration from 20 to 5% during in vitro insemination did not affect the fertilization rate, but improved the subsequent embryonic development in vitro. Optimal concentrations of glucose in the in vitro insemination medium depend on the composition of the basic medium for insemination. Glucose at 0 to 1 mM gave higher fertilization rate when SOF was used, while addition of 5 to 13.9 mM glucose showed higher fertilization rate when BO medium was used. It is worthy to note that SOF requires less concentration of sperm to obtain high and reliable fertilization rate as compared to BO medium. Subsequent experiments for embryo culture demonstrated that amino acid supplementation, especially addition of glycine and/or taurine improved the embryonic development to the blastocyst stage. Amino acids added to the culture medium degraded, and produced ammonium ion. However, an increase in the ammonium concentration did not affect the embryonic development under the present culture conditions. Results also revealed that both insulin and insulin-like growth factor-I (IGF-I) stimulate the in vitro development of bovine embryos by acting synergistically with amino acids through IGF-I receptor. Optimal concentration of glucose during in vitro culture of bovine embryos was around 1 mM when zygotes were cultured throughout 6 days to the blastocyst stage, although glucose requirement for bovine embryos changed between 8- to 16-cell stages. The in vitro culture system established in the present study was successfully applied for the production of clone embryos using nuclear transplantation technique.
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