The Thai Journal of Veterinary Medicine


The objective of this study was to determine the optimum condition for porcine oocyte preparation for in vitro fertilization. A total of 1,130 immature oocytes collected from superficial follicles, 3-6 mm in diameter were allocated to 3 experiments. In Experiment 1, the length of time for oocyte maturation were examined (n=469). Immature oocytes were cultivated in TCM199 NaHCO3 supplemented with 10 μg/ml of FSH/LH and 1 μg/ml of Estradiol-17ẞ for 24, 40 or 48 hrs. Approximately 20% of oocytes were fixed and stained with aceto orcein to observe the maturation rate and the remainder were fertilized in vitro by freshly ejaculated sperm. After 4 hrs, the sperm at the concentration of 1x106 spz/ml were introduced to IVF medium containing 20-30 matured oocytes and were incubated for 18 hrs. The oocytes were later cultivated in TCM199 NaHCO3 + 10% FCS for 5 days to observe their cleavage and development. In vitro maturation, fertilization and embryo culture were performed at 38.5°C in an atmosphere of 5% CO2 in air. Higher maturation rates and higher cleavage rates were found in the 40 and 48 hr culture groups (67.6%, 23/34; 45.5%, 55/121 and 70.6%, 24/34; 37.6%, 49/130) than in the 24 hr group (10.3%, 3/29; 5%, 6/121) respectively (P<0.01). In Experiment 2, the oocyte quality was examined (n=400). Oocytes were categorized into 3 groups at collection, CCO (complex cumulus oocyte), P (partial layered cumulus oocyte) and D (denuded oocyte). Cleavage rates in CCO (36.8%, 50/136) and P (31.6%, 48/152) groups were significantly (P<0.05), higher than in the D group (19.6%, 22/112). In Experiment 3, the source of oocytes from sow or gilt ovaries (n=261) were compared. The difference in the cleavage rates was found between the 2 groups (23.4%, 30/128) and (48.1%, 64/133) for sow and gilt ovaries (P<0.05), respectively. Results indicate that a maturation period of 40 to 48 hrs is required for pig oocytes and that cumulus oocytes complexes require at least layer of cumulus cells for maturation and fertilization.



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