| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2006: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2005: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Research Abstract |
The present study investigated 1) to examine effects of cysteamine or p-mercaptoethanol in a defined maturation medium for porcine ooctytes, 2) to establish a vitrification method of porcine oocytes, 3) to establish a Novel method for intracytoplasmic sperm injection (ICSI) to in vitro matured porcine oocytes and to produce blastocysts in a serum-free, defined culture medium, and 4) to examine a serum-free vitrification solution for porcine oocytes. The following results were obtained.
1) The addition of 100 μM cysteamine (37%) or p-mercaptoethanol (27%) resulted in significantly higher blastocyst rates following ICSI than that of the PVA containing control medium (16%), and were similar to that using a medium containing follicular fluid (31%).
2) For ICSI to in vitro matured porcine oocytes, a Novel method is effective without a high polyspermic fertilization and produce an efficient rate (over 30%) of blastocysts following ICSI.
3) For vitrification of porcine immature oocytes, a solution of 30% ethylene glycol (EG) + 0.5 M sucrose using a Cryotop method produced 37% of in vitro matured oocytes after in vitro maturation culture, and a blastyoicyst rate of 14% was obtained following ICSI.
4) For vitrification of porcine immature oocytes, there was a significant correlation in the proportion of normal morphological oocytes between equilibration times and the presence or absence of cumulus cells; 4 min equilibration for cumulus-enclosed oocytes and 1 min for cumulus-denuded oocytes.
5) For embryonic development of porcine oocytes following ICSI, the Well of the Well (WOW) culture system using 1 mm in diameter was effective in a defined culture medium.
6) It was shown that in vitro matured porcine oocytes were obtained by a vitrification solution without serum and followed by in vitro maturation using a defined maturation medium, although the maturation rates was low (less than 10%).10.
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