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Recent Research on Production of Cloned Animals by Nuclear Transplantation (핵이식에 의한 복제동물 생산의 최근 연구 성과)

  • 박충생
    • Journal of Embryo Transfer
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    • v.8 no.1
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    • pp.37-44
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    • 1993
  • Nuclear transplantation techinque has been found to be the most potential and efficient method for producing a large number of genetically identical animals from a single embryo. The technical development of nuclear transplantation in mammals and its application to the production of cloned animals were reviewed. For the efficient and successful production of cloned embryos by nuclear transplantation, selection and micromanipulation of recipient eggs or embryos as capacious recipient cytoplasm, and benefitial preparation of multiple totipotent embryonic cells as donor nuclei, and also fusion technique are very critical. Recent works approaching to these critical points were introduced and discussed.

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Application of Nuclear Transplantation to Production of Cloned Animals (복제동물 생산을 위한 핵이식기법의 응용)

  • 이효충
    • Journal of Embryo Transfer
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    • v.6 no.2
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    • pp.18-29
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    • 1991
  • Nuclear transplantation technique is known as the most potential and efficient method for producing a large number of genetically identical animals from a single embryo. The technical development of nuclear transplantation in mammals and its application to the production of cloned animals are described. For the efficient and successful production of cloned embryos by nuclear transplantation, the right selection and micromanipulation of recipient eggs or embryos as capacious recipient cytoplasm, the adequate and benefitlal preparation of multiple totipotent embryonic cells as donor nuclei, and also the fusion technique are very critical. Recent studies approaching to these critical points are introduced and discussed. Up to date, the overall efficiency of production of cloned embryos and offspring in livestock is estimated to be low. Further technical development of nuclear transplantation will enable large-scale production of cloned livestock and in near future the commercial cloning of animals will become a reality.

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Current Status and Prospects of Nuclear Transplantation Technology for Production of Cloned Animals (복제동물 생산을 위한 핵이식기술의 개발 현황과 전망)

  • 이효종
    • Journal of Veterinary Clinics
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    • v.16 no.1
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    • pp.163-176
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    • 1999
  • The nuclear transplantation technique is known as the most potential and efficient method for producing large numbers of genetically identical animals from a single embryo and somatic cells. After Dolly was introduced in 1997, many scientists were amazed. A possibility came to a reality that live offspring could be produced with differentiated somatic cells from an adult animal. On the other side, many in the press and the sensationalists focused on the socially, ethically and scientifically unacceptable sides of the technology. In this article, the history, current status and prospects of the technological development of nuclear transplantation in mammals and its application to the production of cloned animals are described. For the efficient and successful production of cloned embryos by nuclear transplantation, the right selection, preactivation and micromanipulation of oocytes as capacious recipient cytoplasm, the adequate and benefitial preparation of multiple totipotent embryonic and somatic cells as donor nuclei, fusion of them and in vitro production of cloned embryos are very critical. Recently the overall efficiency of production of cloned embryos and offspring in livestock has been much improved. Cloning will also be a more efficient, faster and useful way of creating transgenic fetuses for gene therapies, gene pharming, organs for xenotransplantation by preselection and mass production of transgenic embryos and consequently improving the production efficiency in transgenic animals. Further technical development of nuclear transplantation will enable large-scale production of cloned livestock and in near future the commercial cloning of animals will become a reality.

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Production of cloned Rabbits by Nuclear Transplantation (핵이식에의한 복제토끼 생산)

  • 이효정;전병균;윤희준;이경미;송상현;공일근;노규진;최민철;최상용
    • Journal of Embryo Transfer
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    • v.9 no.2
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    • pp.161-166
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    • 1994
  • This experiment was carried out to produce cloned aniraals by nuclear transplantation in rabbits. The ovulated oocytes were collected from the oviducts between 14 and 15 hours after hGG injection. The denuded oocytes were used as nuclear recipient cytoplasm following enucleation by micromanipulation. The blastomeres separated from the 8-cell embryos were used as nuclear donor. The nucleated oocytes receiving a blastomere in the perivitelline space were electrically fused in the 0.28 M mannitol solution at 1.5 kV /cm, 60$\mu$sec for three times. The nuclear transplant embryos which were used and developed to 2- to 4-cell stage in vitro were transferred into the oviducts of synchronized recipient does. A total of 64 nuclear transplant embryos were transferred to 7 recipient does and produced three offspring(4.7%) from a foster mother 31 days after embryo transfer.

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Studies on Nuclear Transplantation in Mouse Embryos III. Production of Nuclear Transplanted Mice (생쥐배의 핵치환에 관한 연구 III. 핵치환 생쥐의 생산)

  • 박용석;정형민;박세필;이상진;정병현;정길생
    • Korean Journal of Animal Reproduction
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    • v.13 no.2
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    • pp.93-97
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    • 1989
  • These experiments were carried out to develop the technique of nuclear transplantation necessary for elevating utilization efficiency of high quality embryos and the production of clone animals. Embryos of pronucleus stages were obtained from ICR mice. Removal of pronuclei and their transfer to recipient embryos were carried out by micromanipulation and virus mediation. The results obtained in these experiments were summarized as follows ; 1. Total 337 pairs of pronuclear stage embryos were subjected to nuclear transplantatin and 247 pairs(73.3%) of them were successfully transplanted and the number of fused embryos between transplanted nucleus and cytoplasm was 188 pairs(55.8%). 2. Of the 188 fused embryos cultured in vitro, 174(92.4%), 131(69.7%) and 117(62.2%) embryos were developed to 2-cell, morula and blastocyst stages, respectively. 3. When total 104 nuclear transplanted embryos were transferred to uteri of recipient mice on day 2-3 of pseudopregnancy, 4 of 12 recipient mice were pregnant and the number of embryos developed to young was 28(26.9%).

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Cloning of Transgenic Rabbit Embryos Expressing Green Fluorescent Protein Gene by nuclear Transplantation (Green Fluorescent Protein 발현 토끼 수정란의 핵이식에 의한 복제)

  • Kang, T. Y.;Yin, X. J.;Rho, G. J.;Lee, H.;Chae, Y. J.;Lee, H. J
    • Journal of Embryo Transfer
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    • v.15 no.2
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    • pp.167-173
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    • 2000
  • The principal objective of this study was to clone transgenic embryos in order to improve the efficiency of transgenic animal production by the combination of microinjection and nuclear transplantation techniques. Mature female New Zealand White rabbits were superovulated by eCG and hCG treatments, fllowed by natural mating. Zygotes were collected from the oviducts at 18∼22 h after hCG injection by flushing with D-PBS containing 5% fetal calf serum(FCS). Two to three picoliters of green fluorescent protein(GFP) gene wa microinjected into male pronucleus. The foreign gene-injected zygotes were cultured in TCM-199 or RD medium containing 10% FCS with a monolayer of rabbit oviductal epithelial cells in a 5% CO2 incubator. The morulae expressing GFP gene were selected and their blastomeres were separated for the use of nuclear donor. Following nuclear transplantation of fluorescence-positive morula stage blastomeres, 13 (21.3%) out of 61 fused oocytes developed to blastocyst stage and all of the cloned blastocysts expressed GFP. The results indicate that the screening of transgene in rabbit embryos by GFP detection could be a promisible method for the preselection of transgenic embryos. Also the cloning of preselected transgenic embryos by nuclear transplantatin could be efficiently applied to the multiple production of transgenic animals.

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Nuclear Transplantation of Bovine IVF Embryos by Cell Cycle Control of Recipient Cytoplasm (수핵란 세포질의 세포주기 조절에 의한 소 체외수정란의 핵이식)

  • 정희태;임석기;박춘근;양부근;김정익
    • Korean Journal of Animal Reproduction
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    • v.20 no.3
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    • pp.307-313
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    • 1996
  • This study was conducted to investigate the effect of S-phase synthronized nuclear transfer on the development of nuclear transplant bovine embryos. A blastomere derived from the 16~32 cell stage bovine embryos was transferred into an enucleated metaphase II(MII) oocytes or activated S-phase eggs. From the MII-phase and S-phase nuclear transfer, 6.3%(4/63) and 13.8%(9/65) of nuclear transplant embryos developed to the blastocyst stage, respectively. In the S-phase nuclear transfer, maximal proportion of embryos developed to the blastocyst stage(16.6%) was obtained after the recipient cell was activated 8 h prior to receving a donor nucleus. MII-phase nuclear transplant embryos showed the PCC state of their nuclear at 1.5~2 h after fusion, whereas, S-phase nuclear transplant embryos did not undergo PCC. The result of this study suggests that if blastomeres of unknown cell-cycle-stage are used, S-phase nuclear transplantation through the activation of enucleated oocytes prior to fusion enhances development of nuclear transplant embryos. This result also suggests that the interval time from oocyte activation to cell fusion may affect the development of nuclear transplant embryos.

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Systems for Production of Calves after Embryo Transfer of Nuclear Transplant Embryos (소 핵이식 수정란에 의한 산자 생산에 관한 연구)

  • 황우석
    • Journal of Embryo Transfer
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    • v.10 no.1
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    • pp.83-90
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    • 1995
  • Production of calves after transfer of nuclear transplant embryos is the latest technology to be applied in commercial livestock breeding. The objective of this study was to establish an efficient procedure to produce offsprings from nuclear transplant embryos. The fusion rates (72.7% vs. 80.8%), cleavage rates (62.5% vs. 71.4%) and rates of development in vitro (12.0% vs. 15.2%) of nuclear transplant embryos were not significantly different between 30 and 40h maturation age of cytoplast. The in vivo and in vitro-derived embryos as nuclei donor were used in this system of bovine nuclear transplantation. Fusion rates of nuclear transplant embryos were not significantly different between in vivo and in vitro-derived embryos (73.0 and 79.2%, respectively). The percentage of embryos reaching the morulae or blastocysts were 21.8% for in vivo-derived embryos and 11.9% for in vitro-derived embryos (p<0.01). Pregnancy rates after embryo transfer of nuclear transplant embryos were not significantly different between in vivo and in vitro-derived embryos (45.9 and 40.5%, respectively). However, calving rates after embryo transfer of nuclear transplant embryos were significantly higher in the in vivo-derived embryos than in vitro (p<0.01). Further research for age of cytoplast and use of in vitro-derived embryos as nuclei donor is required in this system. In conclusion, these results clearly show that the use of in vitro-derived oocytes as recipient cytoplast can improve the nuclear transplant system for genetic progress in cattle.

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Splenic Hemochromatosis Incidentally Found on Tc-99m MDP Bone Scan in a Chronic Myelogenous Leukemia Patient who Received Bone Marrow Transplantation (골수 이식을 받은 만성 골수성 백혈병 환자의 Tc-99m MDP 골스캔에서 우연히 발견된 비장 Hemochromatosis)

  • Seo, Ji-Hyoung;Bae, Jin-Ho;Jeong, Sin-Young;Lee, Jae-Tae;Lee, Kyu-Bo;Ahn, Byeong-Cheol
    • The Korean Journal of Nuclear Medicine
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    • v.38 no.1
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    • pp.109-110
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    • 2004
  • Purpose: Tc-99m MDP bone scan was performed to evaluate a generalized bone pain in a 24-year-old male chronic myelogenous leukemia patient who received bone marrow transplantation at 7 months ago. The patient had received large amounts of blood transfusion for managing symptoms related to anemia. Bone scan revealed substantial splenic tracer uptake. Magnetic resonance image and laboratory evidence of hemochromatosis suggests that the presence of large quantities of iron in the spleen of this patient may have been responsible for the splenic uptake of the bone scanning agent. The authors report a case of splenic hemochromatosis incidentally found on Tc-99m MDP bone scan.

Studies on cryopreservation of nuclear transplanted mouse embryos (마우스 핵이식란의 동결에 관한 연구)

  • Lee, Byeong-chun;Jo, Choong-ho;Hwang, Woo-suk
    • Korean Journal of Veterinary Research
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    • v.33 no.1
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    • pp.171-178
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    • 1993
  • The present study was carried out to investigate the developmental potency to blastocyst after freezing and thawing of nuclear transplanted 2-cell embryos. The nuclei from 2-, 4- and 8-cell mouse embryos were transferred into enucleated 2-cell embryos, and the reconstituted embryos were submitted to direct current(DC) pulse at output voltage of 2.0 kV/cm for $100{\mu}$ sec to induce cell fusion. The recovery rate and developmental potency to blastocyst after freezing and thawing of nuclear transplanted 2-cell embryos was investigated. 1. The recovery rate of nuclear transplanted 2-cell embryos in normal morphology after freezing and thawing was significantly higher in rapid freezing(DMSO 4.5M) than in slow cooling(p<0.01). 2. When the recovered embryos in normal morphology were cultured in vitro, there were no significant differences in the developmental potency to blastocyst between the freezing methods and the concentrations of cryoprotectant. In summary, these experiments have proved that rapid freezing method(DMSO 4.5M) is effective in nuclear transplanted 2-cell mouse embryos. If improved micromanipulation techniques and freezing are combined, nuclear transplantation technique will contribute to the improvement of productivity in livestock animals.

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