1992 Fiscal Year Final Research Report Summary
In-vitro manipulation of mammalian embryos
| Project/Area Number |
02556039
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Applied veterinary science
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| Research Institution | Hokkaido University |
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Principal Investigator |
KANAGAWA Hiroshi Hokkaido University, Veterinary Medicine, Professor, 獣医学部, 教授 (00111162)
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| Co-Investigator(Kenkyū-buntansha) |
ISHIMORI Hisao Embryo Transfer Laboratory, Snow Brand Milk Products Co., Ltd. Researcher, 受精卵移植研究所, 研究員
SEIKE Noboru Embryo Transfer Laboratory, Snow Brand Milk Products Co., Ltd. Chief Researcher, 受精卵移植研究所, 主席研究員
HISHINUMA Mitsugu Hokkaido University, Veterinary Medicine, Instructor, 獣医学部, 助手 (30183578)
TAKAHASHI Yoshiyuki Hokkaido University, Veterinary Medicine, Associate Professor, 獣医学部, 助教授 (70167485)
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| Project Period (FY) |
1990 – 1992
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| Keywords | Embryos / Cryopreservation / Vitrification / Fusion of embryos / Nuclear transplantation / 単為発生 |
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| Research Abstract |
In this project, we have performed a lot of studies on cryopreservation and cloning of mammalian embryos including mouse and large domestic animals such as sheep and cattle.
We established a new rapid freezing procedure using a mixture of permeating cryoprotectant such as 3M ethylene glycol and 0.25 M non-permeating sugar (lactose, sucrose or trehalose). This simple freezing procedure was demonstrated to be very effective not only for cryopreservation of mouse embryos and oocytes, but also for sheep embryos. We also developed two types of new vitrification solutions consisted of 25% ethylene glycol and 25% DMSO; and 20% ethylene glycol, 20% DMSO and 10% 1,3-butanediol, respectively. These two vitrification solutions gave not only a high in-vitro survival for mouse and cattle embryos, but also resulted in many offspring, both mice and calves, after the transfer of vitrified-warmed embryos to recipient animals.
Regarding the cloning of embryos, we have done a lot of experiments using mouse oocytes and embryos to determine the factors affecting reprograming of nuclear tranplanted embryos. One of the most salient findings from our results is that the donor cell-cycle stage critically affects the chromatin structure and development of nuclear transplant embryos when the nuclei from donor embryos were transferred to enucleated oocytes. With transplantation of early-stage nuclei, high proportions of development to the blastocyst and of offspring were obtained from nuclear transplant embryos with a nucleus from a 2-, 4- or 8-cell-stage embryos. This is the first time that such result has been achieved.
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