| Project/Area Number |
12660250
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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 | Ibaraki University |
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Principal Investigator |
KANAZAWA Takuya Ibaraki Univ., Sch. of Agric., Assist Prof., 農学部, 助手 (70272119)
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| Co-Investigator(Kenkyū-buntansha) |
KOHOMOTO Kaoru Nippon Vet. Anim. Sci. Univ., Dept of Anim Sci., Professor, 獣医畜産学部, 教授 (30011894)
OGAWA Yasuki Ibaraki Univ., Sch. of Agric., Assoc. Prof., 農学部, 助教授 (00302331)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2001: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2000: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Keywords | male germ cell / monoclonal antibody / differential-display / mammal / testes / 家畜 |
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| Research Abstract |
This project was conducted to explore genes which are specifically expressed during growth and differentiation of male germ cells, and to produce monoclonal antibodies that specifically recognize particular types of germ cells in seminiferous tubules of the mammals. First, to produce monoclonal antibodies, cells were isolated after enzymatic digestion of testicular tissues of adult male goat and were used to immunize mice. Hybidoma cells were produced by fusing splenocytes of the immunized mice and mouse myeloma cells. First screening using ELISA for mouse immunoglobulins selected approximately 100 hybridoma cell clones, which produce mouse immunoglobulilns, among a few thousands of hybridoma cells. Second screening using an immunocytochemistry of goat testicular cells limited clones which produced antibodies specific for the cells. Immunohistochemistry of frozen sections prepared from tvarious tissues of adult male goat, including testis, kidney, liver, spleen, epididymis and cardiac, revealed tissue-specificity of antibody-binding of 6 monoclonal antibodies and localization of the relevant antigens in the testes. To explore genes which are responsible for spermatogensis, the differential-display technique was performed. More than 20 cDNA fragments detected using total RNA extracted from pre- and post-spermatogenic goat testis. Each cDNA fragment was isolated and incorporated into cloning vector. Some of cloned cDNA were either sequenced for homology analysis and were labeled with digoxgenin to probe the specific transcripts in the testis.
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