This study was performed to comprehend the developmental characteristics of cloned embryos knocked out (KO) of α-1,3-galactosyltransferase (GalT) gene. Immature oocytes were collected and cultured for 40 hrs (1-step) or 20hrs (with hormone) + 20hrs (without hormone) (2-step). The embryos transferred with miniature pig ear fibroblast cell were used as control. The reconstructed embryos were cultured in PZM-3 with 5% CO2 in air at 38.5℃ for 6 days. To determine the quality of the blstocysts, TUNEL and quantitative realtime RT-PCR were performed. The embryos were transferred to a surrogate (Landrace) at an earlier stage of the estrus cycle. The maturation rate was significantly higher in 2-step method than that of 1-step (p<0.05). The blastocyst development of GalT KO embryos was significantly lower than that of normal cloned embryos (p<0.05). The total and apoptotic cell number of GalT KO blastocysts was not different statistically from control. The relative abundance of Bax-α/Bcl-xl ratio was significantly higher in both cloned blastocysts than that of in vivo blastocysts (p<0.05). Taken together, it can be postulated that the lower developmental potential and higher expression of apoptosis related genes in GalT KO SCNT embryos might be a cause of a low efficiency of GalT KO cloned miniature pig production.

We have previously produced transgenic (TG) mice expressing the human lactoferrin (hLF), interleukin-10 (hIL-10), and thrombopoietin (hTPO) proteins in the milk. In this study, we examined whether simple crossbreeding between two kids of a single transgenic mouse can produce double transgenics co-expressing two human proteins.. The hLF male, and the hIL-10 male were crossbred with the hIL-10 and hTPO females, and the hTPO female, respectively. PCR analysis for genotyping showed 32%, 23% and 24% double transgenic rates for hLF/hIL-10, hLF/hTPO, and hIL-10/hTPO transgenes, respectively. We analyzed the expression levels of the human proteins from double transgenic mice and compared those with their single transgenic siblings. All double transgenic co-expressed two human proteins at comparable levels to singles', unless hTPO was not co-expressed: for hLF, 1.1 mg/ml in hLF/hIL-10, whereas 0.5 mg/ml in hLF/hTPO; for hIL-10, 4.1 mg/ml in hIL-10/hLF, whereas 1.4 mg/ml in hIL-10/hTPO. Ihe downregulation of hTPO to half level of singles' was observed in double transgenic mice. The possible reason why hTPO co-expressed might lead to down-regulation of another human protein was discussed. These results suggested that double transgenic generated by crossbreeding between two singles' could be useful system for bioreactor.

Most studies on transgenic bioreactors have focused on expression levels of interest genes. In this study we examined whether transgenic bioreactors would inherit expression level of the Oansgene to long-term generations independently of transgene sources. We employed three transgenic mice, which were separately reported, carrying different transgenes and copy numbers, 27 kb of hLF and 22 kb of hIL-10 genomic sequences, and 1.3 kb of hTPO cDNA, respectively. Three females of the transgenic lineages crossbred with a wild-type male up to 20 generations to test transgenic frequencies of their progenies and to determine expression levels of the transgenes. Ultimately, transmission rates of kLF, hIL-10, and hTPO were 64.3+-7.0, 59.3+-9.8, and 56.1+-9.7, respectively, appeared following Mendelian pattern of inheritance. Notably, we found that levels of expressions of hLF, hIL-10, and hTPO in milk were sustained to high numbers of generations. No transgene silencing of expression was observed in every generations of all transgenic mice. In conclusion, we suggest that once established animal bioreactors could consistently transmit the transgene to continual generations, without loss of expressional activity, independently of transgene sources.

This paper describes the traditional Ondol medical care, the specific combination of ancient and contemporary examples to be articulated. As a traditional Korean culture, to be extracted from the historical perspective, it described the relationship and origins between them, From a health perspective, in accordance with the logic and pharmacologyit introduced Ondol as the important principle of health care equipment. The modern architecture of some common harmful to the body, from the perspective of Ondol, it should be improved and based on scientific proof to the theory.

In order to generate transgenic goats expressing human growth hormone (hGH) in their mammary glands, goat β-casein/hGH hybrid gene was introduced into goat zygotes by pronuclear microinjection. DNA-injected embryos were transferred to the oviduct of recipients at 2-cell stage or to the uterus at morula/blastocyst stage after cultivation in glutathione-supplemented mSOF medium in vitro. Pregnancy and survival rate were not significantly different between 2-cell embryos and morula/blastocysts transferred to oviduct and uterus, respectively. One transgenic female goat was generated from 153 embryos survived from DNA injection. Southern blot analysis revealed that the transgenic goat harbored single-copy transgene with a partial deletion in its sequences. Despite of the partial sequence deletion, the transgene was successfully expressed hGH at the level of 72.1±15.1 μg/ml in milk throughout lactation period, suggesting that the sequence deletion had occurred in non-essential part of the transgene for the transgene expression. Unfortunately, however, the transgene was not transmitted to her offspring during three successive breeding seasons. These results demonstrated that goat β-casein/hGH gene was integrated into the transgenic goat genome in a mosaic fashion with a partial sequence deletion, which could result in a low level expression of hGH and a failure of transgene transmission.

μ방lt emitting diodes (LED) devices are commercially introduced as an alternative for low-Ievellaser therapy (11ι，T) ， and it has several advantages over lasers such as a safe, efficient, and less-expensive altemative to treat wounds. And LED irradiation at the same biostimulatory wavelength has similar bíochemical effεαs. In the present study, to asses whether the I핑ht-emitting diode (LED) irradíation can stimulate bone regeneration, irradiated bony defects with or without grafting materials on rat calvaria were compared to corresponding nonirradiated control. Fifty male Sprague-Dawly rats weighing about 150g, were used. Factors for present study were designed as follows, 1) presence or absence of grafting materials, 2) with or without irradiation, and 3) number of irradiation. Two weeks after operation, rats were sacrifìced. Radiologic and 비stomorphologic fmdings were evaluated. Macrospically, there were no incidents of infection, dehiscence, hematoma and necrosis during study. Radiological findings showed greater radiopacity in the graft group and radiopacity increased as the number of irradiation increase. And microscopically, new bαle formation was great in the graft group and increased as the number of irradiation increase, Present study has shown that LED irradiation improved bone regeneration through radiologic and histomorphologic fmdings in rat.

One of the reasons to causing blood coagulation in the tissue of xenografted organs was known to incompatibility of the blood coagulation and anti-coagulation regulatory system between TG pigs and primates. Thus, overexpression of human CD73 (hCD73) in the pig endothelial cells is considered as a method to reduce coagulopathy after pig-to-non-humanprimate xenotransplantation. This study was performed to produce and breed transgenic pigs expressing hCD73 for the studies immune rejection responses and could provide a successful application of xenotransplantation. The transgenic cells were constructed an hCD73 expression vector under control porcine Icam2 promoter (pIcam2-hCD73) and established donor cell lines expressing hCD73. The numbers of transferred reconstructed embryos were 127 ± 18.9. The pregnancy and delivery rate of surrogates were 8/18 (44%) and 3/18 (16%). The total number of delivered cloned pigs were 10 (2 alive, 7 mummy, and 1 died after birth). Among them, three live hCD73-pigs were successfully delivered by Caesarean section, but one was dead after birth. The two hCD73 TG cloned pigs had normal reproductive ability. They mated with wild type (WT) MGH (Massachusetts General Hospital) female sows and produced totally 16 piglets. Among them, 5 piglets were identified as hCD73 TG pigs. In conclusion, we successfully generated the hCD73 transgenic cloned pigs and produced their litters by natural mating. It can be possible to use a mate for the production of multiple transgenic pigs such as α-1,3-galactosyltransferase knock-out /hCD46 for xenotransplantation.

Unlike mouse results, cloning efficiency of nuclear transfer from porcine induced pluripotent stem cells (piPSCs) is very low. The present study was performed to investigate the effect of cell cycle inhibitors on the cell cycle synchronization of piPSCs. piPSCs were generated using combination of six human transcriptional factors under stem cell culture condition. To examine the efficiency of cell cycle synchronization, piPSCs were cultured on a matrigel coated plate with stem cell media and they were treated with staurosporine (STA, 20 nM), daidzein (DAI, 100 μM), roscovitine (ROSC, 10 μM), or olomoucine (OLO, 200 μM) for 12 h. Flow Cytometry (FACs) data showed that piPSCs in control were in G1 (37.5±0.2%), S (34.0±0.6%) and G2/M (28.5±0.4%). The proportion of cells at G1 in DAI group was significantly higher than that in control, while STA, ROSC and OLO treatments could not block the cell cycle of piPSCs. Both of viability and apoptosis were affected by STA and ROSC treatment, but there were no significantly differences between control and DAI groups. Real-Time qPCR and FACs results revealed that DAI treatment did not affect the expression of pluripotent gene, Oct4. In case of OLO, it did not affect both of viability and apoptosis, but Oct4 expression was significantly decreased. Our results suggest that DAI could be used for synchronizing piPSCs at G1 stage and has any deleterious effect on survival and pluripotency sustaining of piPSCs.

Background : Ginseng berry(GB) is useful not only just in growing source but also in functional food source. The ingredients of crops varies with the maturity. So, GB ingredients need to be analyse for optimal harvesting stage of GB against appropriate use. Methods and Results : This study was carried out to determine optimal harvesting stage of GB. GB was harvested 5 day periods from July 12, started harvesting when pollination was 50 days old, until August 1. GB was analysed color, ginsenosides and fatty acids using colorimeter, LC and GC, respectively. As the majority of GB increase, color of freeeze drying GB powder were changed that lightness and yellowness was increased, redness was decreased. Ginsenoside Re, Rb1 and Rb2, major ginsenoside in GB, were increased and Ginsenoside F1, Rk1 and Rg5, minor ginsenoside, were increased for a time and then decreased. Oleic acid, the main fatty acid in GB, was decreased, and linoleic acid and total fatty acid content was increased to July 27 and then decreased. Conclusion : Total ginsenosides content was the highest on August 1 and total fatty acid content was the highest July 27. As the majority of GB increase, ratio of oleic acid on total fatty acid was decreased and linoleic acid was increased. Thus, GB is that the longer a harvest period and the more useful for food source.