In order to investigate genetic stability and gene expression profile after cloning procedure, two groups of cloned pigs were used for swine leukocyte antigen (SLA) gene nucleotide alteration and microarray analyses. Each group was consist of cloned pigs derived from same cell line (n=3 and 4, respectively). Six SLA loci were analyzed for cDNA sequences and protein translations. In total, 16 SLA alleles were identified and there were no evidence of SLA nucleotide alteration. All SLA sequences and protein translations were identical among the each pig in the same group. On the other hand, microarray assay was performed for profiling gene expression of the cloned pigs. In total, 43,603 genes were analyzed and 2,150～4,300 reliably hybridized spots on the each chip were selected for further analysis. Even though the cloned pigs in the same group had identical genetic background, 18.6～47.3% of analyzed genes were differentially expressed in between each cloned pigs. Furthermore, on gene clustering analysis, some cloned pigs showed abnormal physiological phenotypes such as inflammation, cancer or cardiomyopathy. We assumed that individual environmental adaption, sociality and rank in the pen might have induced these different phenotypes. In conclusion, the results of the present study indicate that SLA locus genes appear to be stable following SCNT. However, gene expressions and phenotypes between cloned pigs derived from the same cell line were not identical even under the same rearing conditions.

The growing oocytes become progressively capable of resuming meiosis, and full meiotic competence appear when they are about 80% of the size of fully grown oocytes. As hormonal influences vary at different stages of reproductive cycle, the size of oocytes may vary according to the reproductive stages. The present study was designed to compare the diameter between the ovulated and freshly collected immature canine oocytes. The ovulated oocytes were collected 72 hr after ovulation by oviductal tube flushing by laparotomy under general anesthesia. Immature oocytes were collected by ovarian slicing method. Diameter of all oocytes was measured directly using epiflurescence microscope with a calibrated micro-eyepiece micrometer at ×200 magnification. The thickness of zona pellucida and diameter of cytoplasm were measured separately and recorded. A total of 2209 zona intact oocytes were collected, among them 628 from anestrus, 675 from follicular, 838 from luteal and 68 by fallopian tubes flushing methods. The average number of oocytes was 104.7, 168.8, 119.7 and 11.3 for anestrus, follicular, luteal and fallopian tubes flushing methods, respectively. The average diameters of the ooplasm and oocyte were significantly varied in different reproductive stages as well as with ovulated oocytes (P<0.05). The average diameter of ooplasm and oocyte was 115.6 and 127.7, 143.0 and 162.0, 134.6 and 150.6, 159.6 and 185.6 for anestrus, follicular, luteal and ovulated oocytes, respectively. Highest number of oocytes with larger diameter could be collected from the follicular and luteal stages. In conclusion, the follicular and luteal ovaries are the best sources of oocytes for canine IVM.

The objective of this study was to investigate the effects of oxygen tension during in vitro maturation of porcine oocytes on the nuclear maturation and differences in gene expression. Cumulus-oocyte complexes (COCs) were collected from ovaries obtained at a local slaughterhouse, matured for 44 hours in TCM199 supplemented with porcine follicular fluid (pFF) under 5% or 20% oxygen concentration. In results, oxygen tension had no significant effects on nuclear maturation. Relative poly(A) mRNA abundance of MnSOD, CCNB1, LDHA, G6PD, BCL, GPX1, IGFR2, GLUT1, BAX, GREM, PTGS2 was analysed in cumulus cells. GLUT1, G6PD, LDHA were up-regulated in the cumulus cells matured in low oxygen, suggesting a higher glucose uptake and an increase in anaerobic glycolysis, whereas CCNB, MnSOD were up-regulated in the cumulus cells matured in high oxygen, which suggest a higher activity of mitosis-promoting factor and antioxidant response. In conclusion, cumulus cells increase in glucose metabolism via anaerobic glycolysis under low oxygen concentration and show significant change in antioxidant against oxidant damage or apoptotic response under high oxygen concentration. For such an effect of cumulus cells, oocytes could be matured normally regardless of various oxygen concentration.