碩士 / 國立宜蘭大學 / 動物科技學系碩士班 / 99 / The freeze-dried semen was lower cost and more convenience, not only for long-term preservation but also for long-distance transportation. Being freeze-dried, the spermatozoa lost their motility and could not be used for in vitro fertilization (IVF) or artificial insemination (AI) for the purpose of fertilization. Only intracytoplasmic sperm injection (ICSI) with embryo transfer could be possible for producing offspring. How to reduce the damage to sperm during the freeze-drying process is the major issue in this research. The objective of the present study was to establish the optimal condition for freeze-drying boar semen and evaluated the effect of the concentration of trehalose (0 M, 0.1 M and 0.2 M), storage temperature (25℃, 4℃ and -80℃) and storage time (1 and 6 months) on the quality of freeze-dried semen, male pronuclear formation (MPN) and embryo development after ICSI. Considering the water content and quality change of boar semen after freeze-dried, we decided the freeze-dried condition with 5 mTorr and -54℃ for freeze-drying 16 hours. The water content of freeze-dried semen was reduced to 3.55%, the quality of freeze-dried semen was not affects for 1 week preservation at 18℃. Compared with fresh semen, the viability, acrosomal integrity and mitochondrial function of freeze-dried semen were significantly decreased and DNA fragmentation index (DFI) was increased significantly. However, the 0.1 M or 0.2 M of trehalose freeze-dried semen possessed higher acrosomal integrity and mitochondrial functional and indicated a significantly lower DNA fragmentation index than that without trehalose. Being stored at -80℃ the viability, acrosomal integrity, mitochondrial functional and DNA integrity of freeze-dried semen were better than that stored at 25℃ or 4℃. Despite that the time in storage did not affect its viability and mitochondrial functional, freeze-dried semen in storage for one month indicated higher acrosomal integrity and lower DFI than that in six months. After the injection of freeze-dried sperm into the oocytes, the male pronuclear formation rate, embryos cleavage and embryo development rate between the treatment groups had no significant difference. Therefore, adding the trehalose can enhance the acrosomal integrity, mitochondrial functional and DNA integrity and effectively reduce the damage to the sperm during the freeze-drying process, but it provides no significant improvement to MPN and embryos development after ICSI.
| Identifer | oai:union.ndltd.org:TW/099NIU07289008 |
| Date | January 2011 |
| Creators | Liu, Ying-Jyug, 劉映均 |
| Contributors | Chen, Ming-Cheng, 陳銘正 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 148 |
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