In the present study, we investigated the effects of genotypes on in vitro maturation and fertilization in porcine fresh/frozen-thawed oocytes. The porcine cumulus-oocyte complexes (COCs) were divided into four groups according to whether they were: (1) in vitro matured; (2) cryopreserved and in vitro matured; (3) in vitro fertilized and (4) cryopreserved, and in vitro fertilized. Maturation of porcine COCs was accomplished by incubation in NCSU23 medium. Immature oocytes were cryopreserved by Open Pulled Straws (OPS) method according to Vajta et al., (1998). Oocytes stained by Acetic-Orcein method were observed under the microscope. DNA extracted from the ovaries was analyzed by RAPD (random amplified polymorphic DNA) and SSCP (single strand conformational polymorphisrrt) method. The rates of oocytes maturation and fertilization were significantly high in AA genotype. The results indicated that in vitro maturation and fertilization in porcine fresh/frozen-thawed oocytes may be affected by genotypes in pigs.

This study was carried out to evaluate the effects of glucose and sodium phosphate on in vitro development of porcine oocytes matured and fertilized in vitro. When the culture medium was supplemented with various concentrations of glucose, the higher proportions (23 and 26%) of oocytes developed to morular or blastocyst stages were at the concentrations of 2.78 and 5.56 mM than 0 (9%; P<0.05) and 11.12 mM (18%). In experiment to evaluate effect of sodium phosphate during in vitro development of porcine oocytes, a significantly (P<0.05) higher proportions of embryos developed to morular or blastocyst stages was obtained with sodium phosphateof 0.28 (25%) and 0.53 (27％) mM than 0 (15%), 1.05 (19%) and 2.10 (10%) mM. On the other hand, when oocytes were cultured in medium with (0.53 mM) sodium phosphate, the proportions of developed embryos were significantly (P<0.05) higher in medium without (29%) that than with (14%) 5.56 mM glucose. However, a higher proportion of embryos developed to morular or blastocyst stages were obtained in medium with (23%) that than without (8%) glucose (P<0.05). The minimum essential medium (MEM) added to the culture medium were higher regardless of presence of sodium phosphate and glucose on the development of embryos. Although sodium phosphate and glucose could support morular and blastocyst development to a limited extend (10∼24%), significantly higher proportion (36%) at morular or blastocyst stages was obtained by MEM adding in the medium with sodium phosphate and glucose. These results suggest that the early development of in vitro fertilized porcine oocytes can be maintained efficiently by glucose and sodium phosphate when they were cultured in medium with MEM.

This study was carried out to investigate the effect of cysteamine addition during in vitro maturation, fertilization and culture of porcine oocytes. Oocytes were matured for the first 22 h in mTCM -199 media supplemented with or without 150 μM cysteamine. They then were matured for an additional 22 h in mTCM-199 media without hormones supplemented with or without 150 μM cysteamine. When cumulus-oocyte complexes (COCs) were matured in the mTCM-199 media supplemented with cysteamine, the rates of GVBD and maturation (metaphase Ⅱ) were enhanced as compared to the media without the addition of cysteamine. Also, when COCs were matured in the mTCM-199 media supplemented with cysteamine, the rates of sperm penetration, male pronucleus formation, cleavage and blastocyst formation after in vitro fertilization were enhanced as compared to the media without the addition of cysteamine. In conclusion, it was suggested that oocytes matured for the first 22 h in mTCM-199 media supplemented with 150 μM cysteamine increased the rates of metaphase Ⅱ, sperm penetration, male pronucleus and blastocyst formation were higher as compared to the media without addition of cysteamine.