Placenta is the main nutrition source for the fetus during pregnancy. Thus, it has a pivotal function in the pregnant process. Many functions of the placenta have been elucidated. An abnormal placenta is associated with a high rate of pregnancy failure in somatic cloned bovine. Differentially expressed genes (DEGs) were examined in a comparison between normal and cloned bovine placenta using annealing control primer (ACP)-based GeneFishing PCR. Using 120 ACPs, nearly 80 genes were identified and the fragments of 42 DEGs were sequenced. 38 of these genes were known genes and four were unknown. To determine the DEGs result, six target clones expressing on one-side of a normal and a clone placenta were selected. Through an analysis of the target genes using the real-time PCR, the expressing pattern was found to be somewhat different from the DEGs. Additionally, several genes appeared with the same expression pattern. Taken together, this suggests that the target genes would be essential for research into what influences the placental formative mechanisms during fetal development.

The objective of this study was to analyzed pattern of proteins expression abnormally in cloned bovine placenta. TIMP-2 protein whose function is related to extracellular matrix degradation and tissue remodeling processes was one of differentially up-regulated proteins in SCNT placenta. And one of down-regulated protein in SCNT placenta was identified as vimentin protein that is presumed to stabilize the architecture of the cytoplasm. The expression patterns of these proteins were validated by Western blotting. To evaluate how regulatory loci of TIMP-2 and vimentin genes was programmed reprogramming in cloned placenta, the status of DNA methylation in the promoter region of TIMP-2 and vimentin genes was analyzed by sodium Bisulfite mapping. The DNA methylation results showed that there was not difference in methylation pattern of TIMP-2 and vimentin loci between cloned and normal placenta. Histone H3 acetylation state of the nucleosome was analyzed in the cloned placental and normal placenta by Western blotting. A small portion of the protein lysates were subjected to Western blotting with the antibodies against anti acetyl-Histone H3. Overall histone H3 acetylation state of SCNT placenta was significantly higher than those of normal placenta cells. It is postulated that cloned placenta at the end of gestation seems to be unusual in function and morphology of placenta via improper expression of TIMP-2 and vimentin by abnormal acetylation states of cloned genome.

Offspring have been produced from somatic cells in a number of species. This biotechnology introduced a new phenomenon in reprogramming and differentiation of somatic cell, namely totipotency. However, birth of oversized calves and perinatal abnormalities such as increased gestation length, lack of spontaneous parturition, higher incidence of dystocia, and reduced perinatal viability of offspring are frequently observed in pregnancies of cloned bovine fetuses. Disturbance of feto-placenta has been proposed as likely causes for abnomal growth. However. Little is known the mechanism responsible for the perinatal problems. Therefore, we focused on gestation length in somatic cell nuclear recipient cows. To solve this issues, placental tissues of control and cloned bovine were obtained by a cesarean section (C-section) before 5 days of paturition. Total RNA from control and cloned bovine placenta was extractd by TRIzol reagent. GeneFishing DEG kits (Seegene) were used to identify differentially expression genes. Total RNA (3 ug) were synthesized by M-MLV reverse transcriptase (200 u/ul) with 10 uM dT-annealing control primer (ACP1) at 42C for 90 min. Then, first-strand cDNA (50 ng) was amplified using the 5 uM arbitary ACP (1-20) and 10 uM dT-ACP2 primers. Some specific expression genes were amplified, Now, we are cloning and sequencing. These finding strongly can be support to solve the problems for parturition delay in nuclear transfer cows, suggest that placenta specific proteins are key indicators for the aberration of gestation and placental function in cows.