• Title/Summary/Keyword: Nuclear Transfer

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Regulation of S100G Expression in the Uterine Endometrium during Early Pregnancy in Pigs

  • Choi, Yo-Han;Seo, Hee-Won;Shim, Jang-Soo;Kim, Min-Goo;Ka, Hak-Hyun
    • Asian-Australasian Journal of Animal Sciences
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    • v.25 no.1
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    • pp.44-51
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    • 2012
  • Calcium ions play an important role in the establishment and maintenance of pregnancy, but molecular and cellular regulatory mechanisms of calcium ion action in the uterine endometrium are not fully understood in pigs. Previously, we have shown that calcium regulatory molecules, transient receptor potential vanilloid type 5 (TRPV6) and calbindin-D9k (S100G), are expressed in the uterine endometrium during the estrous cycle and pregnancy in a pregnancy status- and stage-specific manner, and that estrogen of conceptus origin increases endometrial TRPV6 expression. However, regulation of S100G expression in the uterine endometrium and conceptus expression of S100G has been not determined during early pregnancy. Thus, we investigated regulation of S100G expression by estrogen and interleukin-$1{\beta}$ (IL1B) in the uterine endometrium and conceptus expression of S100G during early pregnancy in pigs. We obtained uterine endometrial tissues from day (D) 12 of the estrous cycle and treated with combinations of steroid hormones, estradiol-$17{\beta}$ ($E_2$) and progesterone ($P_4$), and increasing doses of IL1B. Real-time RT-PCR analysis showed that $E_2$ and IL1B increased S100G mRNA levels in the uterine endometrium, and conceptuses expressed S100G mRNA during early pregnancy, as determined by RT-PCR analysis. To determine if endometrial expression of S100G mRNA during the implantation period was affected by the somatic cell nuclear transfer (SCNT) procedure, we compared S100G mRNA levels in the uterine endometrium from gilts with SCNT-derived conceptuses with those from gilts with conceptuses derived from natural mating on D12 of pregnancy. Real-time RT-PCR analysis showed that levels of S100G mRNA in the uterine endometrium from gilts carrying SCNT-derived conceptuses was significantly lower than those from gilts carrying conceptuses derived from natural mating. These results showed that S100G expression in the uterine endometrium was regulated by estrogen and IL1B of conceptus origin, and affected by the SCNT procedure during early pregnancy. These suggest that conceptus signals regulate S100G, an intracellular calcium transport protein, for the establishment of pregnancy in pigs.

The Feasibility of Natural Ventilation in Radioactive Waste Repository Using Rock Cavern Disposal Method (동굴처분 방식을 사용하는 방사성 폐기물 처분장의 자연 환기 타당성 평가)

  • Kim Jin;Kwon Sang Ki
    • Journal of Nuclear Fuel Cycle and Waste Technology(JNFCWT)
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    • v.3 no.3
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    • pp.183-192
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    • 2005
  • Natural ventilation in radioactive waste repositories is considered to be less efficient than mechanically forced ventilation for the repository working environment and hygiene & safety of the public at large, for example, controlling the exposure of airborne radioactive particulate matter. It is, however, considered to play an important role and may be fairly efficient for maintaining environmental conditions of the repository over the duration of its lifetime, for example, moisture content and radon (Rn) gas elimination in repository. This paper describes the feasibility of using natural ventilation which can be generated in the repository itself, depending on the conditions of the natural environment during the periods of repository construction and operation. Evidences from natural cave analogues, actual measurements of natural ventilation pressures in mountain traffic tunnels with vertical shafts, and calculations of airflow rates with given natural ventilation pressures indicate possible benefits from passive ventilation for the prospective Korean radioactive waste repository. Natural ventilation may provide engineers with a cost-efficient method for heat and moisture transfer, and radon (Rn) gas elimination in a radioactive waste repository. The overall thermal performance of the repository may be improved. The dry-out period may be extended, and the seepage flux likely would be decreased.

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Development of IoT-based real-time Toxic Chemical management System (IoT 기반의 실시간 유해 화학물 관리 시스템 개발)

  • Kang, Min-Soo;Ihm, Chunhwa;Jung, Yong-Gyu;Lee, Minho
    • The Journal of the Institute of Internet, Broadcasting and Communication
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    • v.16 no.5
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    • pp.143-149
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    • 2016
  • Recent accidents caused by toxic chemicals and the social problems caused by frequent. As of 2010, there are more than 100,000 types of deadly toxic chemicals being distributed throughout Korea, and severely intoxicated patients along with an enormous number of patients can be induced at the time of an accident involving deadly toxic chemicals. Internationally, the seriousness of large-scale disasters due to a NBC disaster (nuclear, biologic and chemical disaster) is being highlighted as well. So, we obtain the information of the RFID tag attached to a glass bottle with containing the toxic chemical to transfer the data to the smart device has been studied a system that can monitor the status of the toxic chemical in real time. The proposed system is the information was sent to the main system using a zigbee communication by recognizing the tag vial containing the toxic chemical with the 13.56MHz bandwidths good permeability. User may check the information in real time by utilizing the smart device. However, the error of the system for managing the toxic chemical generates a result that can not be predicted. Failure of the system was detecting the error by using a comparator as this can cause an error. And the detected error proposed a duplex system so that they do not affect the overall system.

Effects of Cell Cycle Regulators on the Cell Cycle Synchronization of Porcine induced Pluripotent Stem Cells

  • Kwon, Dae-Jin;Hwang, In-Sul;Kwak, Tae-Uk;Yang, Hyeon;Park, Mi-Ryung;Ock, Sun-A;Oh, Keon Bong;Woo, Jae-Seok;Im, Gi-Sun;Hwang, Seongsoo
    • Development and Reproduction
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    • v.21 no.1
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    • pp.47-54
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    • 2017
  • 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, $100{\mu}M$), roscovitine (ROSC, $10{\mu}M$), or olomoucine (OLO, $200{\mu}M$) for 12 h. Flow Cytometry (FACs) data showed that piPSCs in control were in G1 ($37.5{\pm}0.2%$), S ($34.0{\pm}0.6%$) and G2/M ($28.5{\pm}0.4%$). 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.

Knock-in Somatic Cells of Human Decay Accelerating Factor and α1,2-Fucosyltransferase Gene on the α1,3-Galactosyltransferase Gene Locus of Miniature Pig (α1,3-Galactosyltransferase 유전자 위치에 사람 Decay Accelerating Factor와 α1,2-Fucosyltransferase 유전자가 Knock-in된 미니돼지 체세포)

  • Kim, Ji Woo;Kang, Man-Jong
    • Reproductive and Developmental Biology
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    • v.39 no.3
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    • pp.59-67
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    • 2015
  • Galactose-${\alpha}1,3$-galactose (${\alpha}1,3$-Gal) epitope is synthesized at a high concentration on the surface of pig cells by ${\alpha}1,3$-galactosyltransferase gene (GGTA1). The ${\alpha}1,3$-Gal is responsible for hyperacute rejection in pig-to-human xenotransplantation. The generation of transgenic pigs as organ donors for humans is necessary to eliminate the GGTA1 gene that synthesize $Gal{\alpha}$(1,3)Gal. To prevent hyperacute graft rejection in pig-to-human xenotransplantation, previously, we developed ${\alpha}1,3$-galactosyltransferase gene-knock-out somatic cell by homologous recombination. In this study, we established cell lines of ${\alpha}1,3$-GT knock-out expressing hDAF and hHT gene from minipig fibroblasts to apply somatic cell nuclear transfer. The hDAF and hHT mRNA were expressed in the knock-in somatic cells and ${\alpha}1,3$-GT mRNA was suppressed. However, the knock-in somatic cells were increased resistance to human serum-mediated cytolysis.

The Expression of Plasma Progesterone and Estradiol-17$\beta$ Level before Parturition in the Recipients Pregnant by Hanwoo SCNT Embryos (체세포 복제란 이식 한우의 분만 전 혈장 Progesterone과 Estradiol-l7$\beta$ 농도 변화)

  • Hwang, Seong-Soo;Choi, Sun-Ho;Chang, Yoo-Min;Ko, Yeoung-Gyu;Yang, Byong-Chul;Im, Gi-Sun;Min, Kwan-Sik;Seong, Hwan-Hoo
    • Reproductive and Developmental Biology
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    • v.32 no.3
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    • pp.199-203
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    • 2008
  • This study was performed to analyze the characterization of plasma hormonal levels during pregnancy in the Hanwoo recipients pregnant by artificial insemination (AI) or somatic cell nuclear transfer (SCNT) embryos. The synchronized recipients pregnant by SCNT embryos produced by Hanwoo fetal fibroblast cells (n=8) and by AI (control, n=5) were used. The plasma hormonal levels were measured by RIA (P4 and E2) and ELISA (cortisol), respectively. In control, the increase of E2 and the decrease of P4 were occurred immediately before the initiation of parturition. The expression pattern of plasma P4 was similar in both groups from 50 to 10 days before parturition, however, it did not decrease even at the expected date of labor in the SCNT recipients. The plasma cortisol was expressed a lower level during pregnancy in the SCNT recipients. But, the cortisol was increased in the cow aborted around 100 days of pregnancy (n=1). Based on these results, it can be postulated that the failure of the hormonal changes immediately before parturition in the SCNT recipients may be one of the most important reasons for a delayed parturition in clone calving.

Knock-in Efficiency Depending on Homologous Arm Structure of the Knock-in Vector in the Bovine Fibroblasts (체세포에 있어서 Knock-in 벡터 상동영역 구조에 따른 Knock-in 효율)

  • Kim, Se Eun;Park, Da Som;Koo, Deog-Bon;Kang, Man-Jong
    • Reproductive and Developmental Biology
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    • v.41 no.1
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    • pp.7-16
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    • 2017
  • The knock-in efficiency in the fibroblast is very important to produce transgenic domestic animal using nuclear transfer. In this research, we constructed three kinds of different knock-in vectors to study the efficiency of knock-in depending on structure of knock-in vector with different size of homologous arm on the ${\beta}-casein$ gene locus in the somatic cells; DT-A_cEndo Knock-in vector, DT-A_tEndo Knock-in vector I, and DT-A_tEndo Knock-in vector II. The knock-in vector consists of 4.8 kb or 1.06 kb of 5' arm region and 1.8 kb or 0.64 kb of 3' arm region, and neomycin resistance gene(neor) as a positive selection marker gene. The cEndo Knock-in vector had 4.8 kb and 1.8 kb homologous arm. The tEndo Knock-in vector I had 1.06 kb and 0.64 kb homologous arm and tEndo Knock-in vector II had 1.06 kb and 1.8 kb homologous arm. To express endostatin gene as transgene, the F2A sequence was fused to the 5' terminal of endostatin gene and inserted into exon 7 of the ${\beta}-casein$ gene. The knock-in vector and TALEN were introduced into the bovine fibroblast by electroporation. The knock-in efficiencies of cEndo, tEndo I, and tEndo II vector were 4.6%, 2.2% and 4.8%, respectively. These results indicated that size of 3' arm in the knock-in vector is important for TALEN-mediated homologous recombination in the fibroblast. In conclusion, our knock-in system may help to create transgenic dairy cattle expressing human endostatin protein via the endogenous expression system of the bovine ${\beta}-casein$ gene in the mammary gland.

Model for predicting the $^{137}C_s$ contamination of an agricultural plant following a soil deposition (토양침적에 의한 $^{137}C_s$ 농작물 오염평가 모델)

  • Jun, In;Keum, Dong-Kwon;Kang, Hee-Seok;Choi, Yong-Ho;Lee, Han-Soo;Lee, Chang-Woo
    • Journal of Nuclear Fuel Cycle and Waste Technology(JNFCWT)
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    • v.4 no.4
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    • pp.365-372
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    • 2006
  • A dynamic compartment model is presented to predict the contamination level of agricultural plant by $^{137}C_s$ as a result of a soil deposition. The model considered the processes of a percolation, soil mixing by a plowing before transplanting, plant uptake, leaching to a deep soil, and fixation to a clay mineral. The effects of the soil properties (pH, clay mineral, organic matter content, and exchangeable K), which are spatially varied, on a plant uptake and the leaching rates of $^{137}C_s$ in a root zone soil were modeled by the Absalom model. To test the validity of the model, the $^{137}C_s$ aggregated transfer factors(TFa) for rice plants were compared with those observed from some simulated $^{137}C_s$ soil deposition experiments, which were carried out with respect to rice plants cultivated in seventeen paddy soils of different properties for two consecutive years. Observed $^{137}C_s$ TFa values of the rice plants did not show an evident trend for the pH and clay content of the soil properties, while they increased with an increasing organic matter content or a decreasing exchangeable K concentration. Predicted $^{137}C_s$ TFa values of the rice plants were found to be comparable with those observed.

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Development of Porcine Parthenogenetic Oocytes Activated with Different Combination of Chemicals (Chemical Agent를 이용한 추가 활성화 처리가 돼지 단위발생란의 발달에 미치는 영향)

  • Seo Jin-Sung;Hwang In-Sun;Kim Se-Woong;Park Hyo-Suk;Kim Dong-Hoon;Yang Byoung-Chul;Kong Il-Keun;Yang Boh-Suk;Im Gi-Sun
    • Reproductive and Developmental Biology
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    • v.30 no.1
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    • pp.1-5
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    • 2006
  • Artificial activation of oocytes is a prerequisite for the successful cloning by nuclear transfer. This study investigated the effect of the different combination of activation agents such as electric pulse (E), thimerosal (Thi) + dithiothreitol (DTT), 6-dimethylaminopurine (6-DMAP) or cycloheximide (CH) on the developmental ability of porcine embryos derived from parthenogenetic activation (PA). PA embryos activated with chemicals showed significantly higher developmental rate to the blastocyst stage compared to the embryos activated with E alone ($21.5{\sim}28.1%$ vs. 18.0%, respectively). Of chemicals, Thi + DTT supported higher development to the blastoryst stage (28.1%). There was no significant difference in 1 pronucleus (PN) formation rate $(59.9{\sim}64.7%)$, but 2PN formation rate was significantly higher in PA embryos with additional activation using chemicals $(7.2{\sim}9.7%)$. In conclusion, this study shows that chemical activation after electric pulse can increase the development of porcine PA embryos.

Studies on Oocyte Activation Regimen for Nuclear Transfer in Hanwoo(Korean Cattle) (핵이식을 위한 한우 난자 활성화 처리방법에 관한 연구)

  • 임기순;양보석;박성재;양병철;장원경;박창식
    • Korean Journal of Animal Reproduction
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    • v.24 no.3
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    • pp.281-288
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    • 2000
  • This experiment was carried out to investigate the optimal activation condition for parthenogenetic development. In order to activate oocytes at 22 h post onset of maturation, the oocytes were subjected to 5 $\mu$M ionomycin(I) for 5 min ,10 $\mu$M calcium ionophore(Ca) for 5 min, 2 mM 6-dimethylamino-purine(DMAP) for 3 h and 10 $\mu\textrm{g}$/$m\ell$ cycloheximide(CH) for 6 h alone or in combination. The activated oocytes were cultured in modified CR$_1$aa at 5% $CO_2$, 5% $O_2$, 90% $N_2$. l. The cleavage rates after 48 h culture of oocytes treated with I, Ca, DMAP and CH were 12.7%, 14.1%, 28.9% and 22.9%, respectively. There was no blastocyst formation. 2. The cleavage rates after 48 h culture of oocytes treated with I + DMAP, I + CH, Ca + DMAP and Ca + CH were 96.9%, 82.1%, 93.1% and 34.7%, respectively. Developmental rates to blastocysts were 10.4%, 5.3%, 17.6% and 7.1 %, respectively. When oocytes were treated with Ior Ca followed by DMAP, the blastocyst formation rate was significantly higher than other groups(P <0.05). 3. According to single activation treatment, pronucleus formation rates were 5.4%, 3.6%, 28.3% and 28.8%, respectively, Whereas, all oocytes treated with the combined activation agents formed 100% pronucleus.

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