The Journal of Reproduction and Development Supplement

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The clones produced by somatic cell nuclear transfer (SCNT) technique had the identical genetic background from donors except for cytoplasmic transmission molecules originated from recipient oocytes. Nuclear microsatellite (MS) genotypes and mitochondrial DNA (mtDNA) sequence polymorphisms were analyzed in twenty-one SCNT cloned minipigs generated from three somatic cell presenting donors. The MS genotyping results showed that all cloned minipigs derived from each donor had identical chromosomal genotypes comparing to those of donors. However, mtDNA sequences were polymorphic even within the identical donor-derived progenies strongly suggesting their different origins of enucleated oocytes. Sequencing results using hair root DNA samples showed that there were three or four mtDNA haplotypes found in three donor's cloned minipig populations: twelve cloned minipigs produced from donor D1 had four mtDNA haplotypes, and four from D2 and five from D3 had three mtDNA haplotypes, respectively. On the other hand, among all cloned minipigs nine individuals had heteroplasmic mtDNA sequences mixed two mtDNA haplotypes which transmitted from the enucleated recipient oocytes and nucleus presenting donor cells. Heteroplasmic mtDNAs were observed in three all cloned minipig populations in spite of different frequencies. The results of the present study suggested that the origins of recipient oocytes may be originated from four lineages for production of SCNT cloned minipigs. These studies will further clarify the role of mtDNA heteroplasmy components on the efficiency of nuclear transfer minipigs and on growth and metabolism of the offspring.

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It has been proposed that imprints in uniparental mouse embryos, for some genes, can be appropriately adjusted by dosage compensation or counting mechanisms. We were interested to investigate a possible relationship between imprint expression and ploidy in pig embryos. To gain insight into this phenomenon, we compared the expression patterns of eight imprinted genes and the methylation status of IGF2/H19 DMR3 between haploid and diploid porcine parthenogenetic (PG) blastocysts. The results demonstrated that IGF2, NNAT and PEG10 did not display detectable levels of expression in haploid PG blastocysts while GRB10, H19, IGF2R, PEG1 and XIST were expressed at lower levels in haploid PG blastocysts in comparison to diploid their counterparts. These results indicate that the paternally expressed genes, with the exception of PEG1, were activated in diploid PG blastocysts, but not in haploid PG blastocysts. Moreover, the methylation status of IGF2/H19 DMR3 in haploid and diploid PG blastocysts was investigated using the COBRA assay. The results showed that the IGF2/H19 DMR3 was partially digested in diploid PG blastocysts but this region in haploid PG blastocysts remained undigested. Moreover, the observed digestion pattern in diploid PG blastocysts was somewhat different from that of the IVF blastocysts, indicating that the methylation status of this region in diploid PG blastocysts was partly altered. These findings appear to be evidence that there exists a process to adjust imprinted expression of certain genes, even in diploid PG embryos. This phenomenon may be associated with altered methylation at an imprinting control region.

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Hematopoietic stem cells (HSCs) are cells with self-renewal and multilineage differentiation potential. Recent studies suggest that epigenetic modification plays an importance role in controlling stem cell differentiation and cell fate decisions. In this report, to determine whether DNA Methylation is involved in the regulation of factors related HSCs maintenance, we investigated the DNA methylation status of promoter locus of these factors and expression in prospectively purified mouse hematopoietic stem cell and progenitor cells. There was tissue-dependently differentially methylated region (T-DMR) in the 5’region of the genes related HSCs self-renewal. Here we propose that transcriptional regulation of Bmi1, Mel18 and P21 during HSC differentiation involves epigenetic system by DNA methylation.

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This study was conducted to investigate the variation of growth characteristics and reproductive physiology in cloned Hanwoo male calves during growing stage. The hematological parameters, body weight, and plasma hormonal levels, birth to 12 months, were analyzed in the cloned calves (n=3). The hematological parameters, such as white blood cell, red blood cell, and platelet, were not different in both normal and cloned calves. The difference, however, was significantly higher in the cloned calves, 5-6 months (p<0.05) and 7-12 months (p<0.01), than that of the comparators, respectively. The plasma IGF-1 level was statistically significant in the cloned calves, 5-10 months, compared to that of the normal calves (p<0.05). However, the plasma testosterone level was not different in both normal and clone calves according to growing stage. Taken together, the cloned Hanwoo male calves are growing faster and maintaining a normal reproductive physiology.

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