Somatic cell nuclear transfer (SCNT)・ Oocytes Nuclear Transfer

ECFG21

Electrodes for Oocyte Activation & Nuclear Transfer

Cables, Foot Switch & Others

Nepa Monitoring System

Cell-Culture-Plate Electrodes

Somatic cell nuclear transfer (SCNT)・ Oocytes Nuclear Transfer

APPLICATIONS

Generation of cloned calves and transgenic chimeric embryos from bovine embryonic stem-like cells

Fig. 1. Photographs of calves obtained after nuclear transfer.

A: Two days after birth

B: Four weeks after birth

C: Fingerprinting of DNA from cloned calves, recipient cows, and donor ES-like W3 cells. Electrophoretograms show amplified fragments of DNA derived from leukocytes from recipient cows (panels a, c and e) and cloned calves (panels b, d and f) and from donor ES-like W3 cells (panel g).

Upper and right-side scales indicate the sizes of DNAs (bp) and the intensities of DNA fragments, respectively. Numbers in boxes indicate the sizes of DNAs (upper) and the intensities of DNA fragments (lower). After insertion of donor ES-like cells into the perivitelline space of oocytes, cells and cytoplasts were fused electrically in fusion medium.

(DC: 20V, Pulse length: 50us, Pulse interval: 100ms, 2 Pulses)

Fig. 2.Expression of EGFP in transgenic chimeric embryos derived from ES-like cells.

A： Phase-contrast image of EGFPtransgenic ES-like cells. Magnification: 200x

B： Fluorescence microscopy image of (A)

C： Ten to fifteen ES-like cells being injected into embryos at 8- to 16-cell stage.
(Fluorescence phase-contrast image)

D, E： Micromanipulation procedures for the formation of chimeric embryos generated by EGFP transgenic ES-like cells and recipient embryos by in vitro fertilization. Magnification: 200x

F： Proliferation of transfected ES-like cells after one day in culture following the formation of chimeric embryos. Magnification: 200x

G： EGFP transgenic blastcysts four days after injection of G418-selected ES-like W3 cells that had been transfected with pCX-Neo-EGFP.

H： Fluorescence microscopy image of (G)
Distinct expression of EGFP was apparent in both the ICM and trophectodermal cells.

Dr. Shigeo Saito, Saito Laboratory of Cell Technology
※Biochemical and Biophysical Research Communications,
Volume 309, Issue 1, Pages 104-113, 12 September 2003

PUBLICATIONS

  • Dog

Accomplishment of canine cloning through in vitro matured oocytes: a pioneering milestone

Kukbin Ji, Kangsun Park, Dongern Kim, Eunyoung Kim, Taeyoung Kil , Minkyu Kim

J Anim Sci Technol . 2024 May;66(3):577-586.

  • Pig

Generation by Somatic Cell Nuclear Transfer of GGTA1 Knockout Pigs Expressing Soluble Human TNFRI-Fc and Human HO-1

Kim GA, Lee EM, Cho B, Alam Z, Kim SJ, Lee S, Oh HJ, Hwang JI, Ahn C, Lee BC

Transgenic Res. 2019 Feb;28(1):91-102.

  • Pig

Faster, cheaper, defined and efficient vitrification for immature porcine oocytes through modification of exposure time, macromolecule source and temperature.

Appeltant R, Somfai T, Kikuchi K.

Cryobiology. 2018 Sep 13. pii: S0011-2240(18)30210-4.

  • Pig

The effect of resveratrol on the developmental competence of porcine oocytes vitrified at germinal vesicle stage.

Santos E, Appeltant R, Dang-Nguyen TQ, Noguchi J, Kaneko H, Kikuchi K, Somfai T.

Reprod Domest Anim. 2018 Apr;53(2):304-312.

  • Pig

Timing of CRISPR/Cas9-related mRNA microinjection after activation as an important factor affecting genome editing efficiency in porcine oocytes.

Sato M, Kosuke M, Koriyama M, Inada E, Saitoh I, Ohtsuka M, Nakamura S, Sakurai T, Watanabe S, Miyoshi K

Theriogenology. 2018 Mar 1;108:29-38.

  • Pig

Combined Treatment with Demecolcine and 6-Dimethylaminopurine during Postactivation Improves Developmental Competence of Somatic Cell Nuclear Transfer Embryos in Pigs.

Lee J, You J1, Lee GS, Lee ST, Hyun SH, Lee E.

Anim Biotechnol. 2018 Jan 2;29(1):41-49.

  • Pig

In vitro oocyte maturation in a medium containing reduced sodium chloride improves the developmental competence of pig oocytes after parthenogenesis and somatic cell nuclear transfer.

Lee J, Lee H, Lee Y, Park B, Elahi F, Lee ST, Park CK, Hyun SH, Lee E.

Reprod Fertil Dev. 2017 Aug;29(8):1625-1634.

  • Pig

Investigation of the Developmental Potential and Developmental Kinetics of Bovine Parthenogenetic and Somatic Cell Nuclear Transfer Embryos Using a Time-Lapse Monitoring System.

Park MJ, Kim EY, Kang MJ, Lee JB, Jeong CJ, Park SP

Cell Reprogram. 2017 Aug;19(4):245-254.

  • Pig

Improvement of the developmental competence of porcine oocytes collected from early antral follicles by cytoplast fusion.

Dang-Nguyen TQ, Appeltant R, Somfai T, Ishihara S, Men NT, Santos EC, Noguchi J, Kaneko H, Kikuchi K

J Reprod Dev. 2017 Feb 16;63(1):59-65.

  • Pig

Production of human apolipoprotein(a) transgenic NIBS miniature pigs by somatic cell nuclear transfer.

Shimatsu Y, Horii W, Nunoya T, Iwata A, Fan J, Ozawa M.

Exp Anim. 2016;65(1):37-43.

  • Pig
  • Dog

Absence of nucleolus formation in raccoon dog-porcine interspecies somatic cell nuclear transfer embryos results in embryonic developmental failure.

Jeon Y, Nam YH, Cheong SA, Kwak SS, Lee E, Hyun SH.

J Reprod Dev. 2016 Aug 25;62(4):345-50.

  • Pig

Cell Synchronization by Rapamycin Improves the Developmental Competence of Porcine SCNT Embryos.

Hyun H, Lee SE, Son YJ, Shin MY, Park YG, Kim EY, Park SP.

Cell Reprogram. 2016 Jun;18(3):195-205.

  • Dog

Propagation of elite rescue dogs by somatic cell nuclear transfer.

Oh HJ, Choi J, Kim MJ, Kim GA, Jo YK, Choi YB, Lee BC.

Anim Sci J. 2016 Jan;87(1):21-6.

  • Bovine

Effective Oocyte Vitrification and Survival Techniques for Bovine Somatic Cell Nuclear Transfer.

Park MJ, Lee SE, Kim EY, Lee JB, Jeong CJ, Park SP

Cell Reprogram. 2015 Jun;17(3):199-210.

  • Pig

Nuclear transfer and transgenesis in the pig.

Kurome M, Kessler B, Wuensch A, Nagashima H, Wolf E.

Methods Mol Biol. 2015;1222:37-59.

  • Pig

Pig oocytes with a large perivitelline space matured in vitro show greater developmental competence after parthenogenesis and somatic cell nuclear transfer.

Lee J, You J, Lee GS, Hyun SH, Lee E.

Mol Reprod Dev. 2013 Sep;80(9):753-62.

  • Pig

Production of cloned NIBS (Nippon Institute for Biological Science) and α-1, 3-galactosyltransferase knockout MGH miniature pigs by somatic cell nuclear transfer using the NIBS breed as surrogates.

Shimatsu Y, Yamada K, Horii W, Hirakata A, Sakamoto Y, Waki S, Sano J, Saitoh T, Sahara H, Shimizu A, Yazawa H, Sachs DH, Nunoya T.

Xenotransplantation . 2013 May-Jun;20(3):157-64.

  • Bovine

Forced collapse of the blastocoel enhances survival of cryotop vitrified bovine hatching/hatched blastocysts derived from in vitro fertilization and somatic cell nuclear transfer.

Min SH, Lee E, Son HH, Yeon JY, Koo DB.

Cryobiology. 2013 Apr;66(2):195-9.

  • Pig

Oxamflatin improves developmental competence of porcine somatic cell nuclear transfer embryos.

Park SJ, Park HJ, Koo OJ, Choi WJ, Moon JH, Kwon DK, Kang JT, Kim S, Choi JY, Jang G, Lee BC

Cell Reprogram. 2012 Oct;14(5):398-406.

  • Bovine

Improved cloning efficiency and developmental potential in bovine somatic cell nuclear transfer with the oosight imaging system.

Kim EY, Park MJ, Park HY, Noh EJ, Noh EH, Park KS, Lee JB, Jeong CJ, Riu KZ, Park SP.

Cell Reprogram. 2012 Aug;14(4):305-11.

  • Pig

Production of cloned NIBS (Nippon Institute for Biological Science) and α-1, 3-galactosyltransferase knockout MGH miniature pigs by somatic cell nuclear transfer using the NIBS breed as surrogates.

Klymiuk N, Böcker W, Schönitzer V, Bähr A, Radic T, Fröhlich T, Wünsch A, Keßler B, Kurome M, Schilling E, Herbach N, Wanke R, Nagashima H, Mutschler W, Arnold GJ, Schwinzer R, Schieker M, Wolf E.

FASEB J. 2012 Mar;26(3):1086-99.

  • Pig

Cleavage pattern and survivin expression in porcine embryos by somatic cell nuclear transfer.

Jeon Y, Jeong SH, Biswas D, Jung EM, Jeung EB, Lee ES, Hyun SH.

Theriogenology. 2011 Oct 15;76(7):1187-96.

  • Human

Human oocytes reprogram somatic cells to a pluripotent state.

Noggle S et al.

Nature. 2011 Oct 5;478(7367):70-5.

  • Bovine

Effect of Cryopreservation and in Vitro Culture of Bovine Fibroblasts on Histone Acetylation Levels and in Vitro Development of Hand-Made Cloned Embryos

Chacón L, Gómez MC, Jenkins JA, Leibo SP, Wirtu G, Dresser BL, Pope CE.

Zygote. 2011 Aug;19(3):255-64

  • Cat

Trichostatin A modified histone covalent pattern and enhanced expression of pluripotent genes in interspecies black-footed cat cloned embryos but did not improve in vitro and in vivo viability.

Gómez MC, Pope CE, Biancardi MN, Dumas C, Galiguis J, Morris AC, Wang G, Dresser BL.

Cell Reprogram. 2011 Aug;13(4):315-29.

  • Dog

Effective donor cell fusion conditions for production of cloned dogs by somatic cell nuclear transfer.

Park J, Oh H, Hong S, Kim M, Kim G, Koo O, Kang S, Jang G, Lee B

Theriogenology. 2011 Mar 1;75(4):777-82.

  • Bovine

Combination of the somatic cell nuclear transfer method and RNAi technology for the production of a prion gene-knockdown calf using plasmid vectors harboring the U6 or tRNA promoter.

Wongsrikeao P, Sutou S, Kunishi M, Dong YJ, Bai X, Otoi T.

Prion. 2011 Jan-Mar;5(1):39-46.

  • Pig

Effects of vascular endothelial growth factor on porcine preimplantation embryos produced by in vitro fertilization and somatic cell nuclear transfer.

Biswas D, Jung EM, Jeung EB, Hyun SH.

Theriogenology. 2011 Jan 15;75(2):256-67.

  • Pig

Enhancement of cytoplasmic maturation of in vitro-matured pig oocytes by mechanical vibration.

Mizobe Y, Yoshida M, Miyoshi K.

J Reprod Dev. 2010 Apr;56(2):285-90.

  • Pig

Latrunculin A dramatically improves the developmental capacity of nuclear transfer embryos derived from gene-modified clawn miniature pig cells.

Himaki T, Mori H, Mizobe Y, Miyoshi K, Sato M, Takao S, Yoshida M.

Cell Reprogram. 2010 Apr;12(2):127-31.

  • Pig

Valproic acid enhances in vitro development and Oct-3/4 expression of miniature pig somatic cell nuclear transfer embryos.

Miyoshi K, Mori H, Mizobe Y, Akasaka E, Ozawa A, Yoshida M, Sato M.

Cell Reprogram. 2010 Feb;12(1):67-74.

  • Bovine

Production of bovine cloned embryos with donor cells frozen at a slow cooling rate in a conventional freezer (-20 degrees C).

Chacón L, Gómez MC, Jenkins JA, Leibo SP, Wirtu G, Dresser BL, Pope CE.

Zygote. 2009 Nov;17(4):341-51.

  • Pig

Post-activation treatment with demecolcine improves development of somatic cell nuclear transfer embryos in pigs by modifying the remodeling of donor nuclei.

Song K, Hyun SH, Shin T, Lee E.

Mol Reprod Dev. 2009 Jul;76(7):611-9.

  • Cat

Generation of domestic transgenic cloned kittens using lentivirus vectors.

Gómez MC, Pope CE, Kutner RH, Ricks DM, Lyons LA, Ruhe MT, Dumas C, Lyons J, Dresser BL, Reiser J.

Cloning Stem Cells . 2009 Mar;11(1):167-76.

  • Cat

Nuclear transfer of sand cat cells into enucleated domestic cat oocytes is affected by cryopreservation of donor cells.

Gómez MC, Pope CE, Kutner RH, Ricks DM, Lyons LA, Ruhe M, Dumas C, Lyons J, López M, Dresser BL, Reiser J.

Cloning Stem Cells . 2008 Dec;10(4):469-83.

  • Cat

Production of second-generation cloned cats by somatic cell nuclear transfer.

Yin XJ1, Lee HS, Yu XF, Kim LH, Shin HD, Cho SJ, Choi EG, Kong IK.

Theriogenology. 2008 May;69(8):1001-6.

  • Bovine

Cloned Embryos from Semen. Part 2: Intergeneric Nuclear Transfer of Semen-Derived Eland (Taurotragus oryx) Epithelial Cells into Bovine Oocytes

Nel-Themaat L, Gómez MC, Pope CE, Lopez M, Wirtu G, Jenkins JA, Cole A, Dresser BL, Bondioli KR, Godke RA.

Cloning Stem Cells. 2008 Mar;10(1):161-72.

  • Cat

Generation of Cloned Transgenic Cats Expressing Red Fluorescence Protein

Yin XJ, Lee HS, Yu XF, Choi E, Koo BC, Kwon MS, Lee YS, Cho SJ, Jin GZ, Kim LH, Shin HD, Kim T, Kim NH, Kong IK.

Biol Reprod. 2008 Mar;78(3):425-31.

  • Pig

Effects of recloning on the efficiency of production of alpha 1,3-galactosyltransferase knockout pigs

Fujimura T, Murakami H, Kurome M, Takahagi Y, Shigehisa T, Nagashima H.

J Reprod Dev. 2008 Feb;54(1):58-62.

  • Pig

Birth of cloned miniature pigs derived from somatic cell nuclear transferred embryos activated by ultrasound treatment

Miyoshi K, Inoue S, Himaki T, Mikawa S, Yoshida M.

Mol Reprod Dev. 2007 Dec;74(12):1568-74.

  • Bovine

Effect of Cryoprotectant Composition on In Vitro Viability of In Vitro Fertilized and Cloned Bovine Embryos Following Vitrification and In-Straw Dilution

Taniguchi M, Ikeda A, Arikawa E, Wongsrikeao P, Agung B, Naoi H, Nagai T, Otoi T.

Journal of Reproduction and Development, 10 April 2007

  • Pig

Modification of maturation condition improves oocyte maturation and in vitro development of somatic cell nuclear transfer pig embryos

Song K, Lee E.

J Vet Sci . 2007 Mar;8(1):81-7.

  • Boar

A Comparison of Reproductive Characteristics of Boars Generated by Somatic Cell Nuclear Transfer to Highly Related Conventionally Produced Boars

Williams NE, Walker SC, Reeves DE, Sherrer E, Galvin JM, Polejaeva I, Rampacek G, Benyshek L, Christenson RK, Graves WM, Pratt SL.

Cloning Stem Cells . 2006 Fall;8(3):130-9.

  • Pig

In Vitro Development of Cloned Embryos Derived from Miniature Pig Somatic Cells after Activation by Ultrasound Stimulation

Miyoshi K, Sato K, Yoshida M.

Cloning Stem Cells. 2006 Fall;8(3):159-65.

  • Cat

Chromosomal Aneuploidy in African Wildcat Somatic Cells and Cloned Embryos

Gómez MC, Pope CE, López M, Dumas C, Giraldo A, Dresser BL.

Cloning Stem Cells. 2006 Summer;8(2):69-78.

  • Pig

Optimization of Ca2+ Concentrations in Fusion and Activation Media for Production of Cloned Embryos from Miniature Pig Somatic Cells

Miyoshi K, Yuki Y, Yoshida M.

J Reprod Dev. 2005 Dec;51(6):699-706.

  • Pig

Caffeine promotes premature chromosome condensation formation and in vitro development in porcine reconstructed embryos via a high level of maturation promoting factor activity during nuclear transfer

Kawahara M, Wakai T, Yamanaka K, Kobayashi J, Sugimura S, Shimizu T, Matsumoto H, Kim JH, Sasada H, Sato E.

Reproduction. 2005 Sep;130(3):351-7.

  • Cat

Birth of African Wildcat Cloned Kittens Born from Domestic Cats

Gómez MC, Pope CE, Giraldo A, Lyons LA, Harris RF, King AL, Cole A, Godke RA, Dresser BL.

Cloning Stem Cells. 2004;6(3):247-58.

  • Bovine

Expression of trophoblast cell-specific pregnancy related genes in somatic-cell cloned bovine pregnancies

Patel OV, Yamada O, Kizaki K, Takahashi T, Imai K, Takahashi S, Izaike Y, Schuler LA, Takezawa T, Hashizume K.

Biology of Reproduction, Volume 70, Issue 4, Pages 1114-1120, April 2004

  • Cat

Nuclear Transfer of Synchronized African Wild Cat Somatic Cells into Enucleated Domestic Cat Oocytes

Gómez MC, Jenkins JA, Giraldo A, Harris RF, King A, Dresser BL, Pope CE.

Biol Reprod. 2003 Sep;69(3):1032-41.

  • Pig

Utility of Rapidly Matured Oocytes as Recipients for Production of Cloned Embryos from Somatic Cells in the Pig

Biol Reprod. 2002 Aug;67(2):540-5.

  • Bovine

In Vitro Development of Bovine Nuclear Transfer Embryos from Transgenic Clonal Lines of Adult and Fetal Fibroblast Cells of the Same Genotype

Arat S, Gibbons J, Rzucidlo SJ, Respess DS, Tumlin M, Stice SL.

Biol Reprod. 2002 Jun;66(6):1768-74.

  • Bovine

Enhanced Survivability of Cloned Calves Derived from Roscovitine-Treated Adult Somatic Cells

Gibbons J, Arat S, Rzucidlo J, Miyoshi K, Waltenburg R, Respess D, Venable A, Stice S.

Biol Reprod. 2002 Apr;66(4):895-900.

  • Pig

Development of porcine embryos reconstituted with somatic cells and enucleated metaphase I and II oocytes matured in a protein-free medium

Miyoshi K, Rzucidlo SJ, Gibbons JR, Arat S, Stice SL.

BMC Dev Biol. 2001;1:12.

  • Pig

Establishment of a Porcine Cell Line from In Vitro-Produced Blastocysts and Transfer of the Cells into Enucleated Oocytes

Miyoshi K, Taguchi Y, Sendai Y, Hoshi H, Sato E.

Biol Reprod. 2000 Jun;62(6):1640-6.

Electroporation

Drug Delivery and Transfection

Electro Cell Fusion

Fluorescent Staining

Single-Cell/Micro-Particle Transfer

Cell Freezing

Mechanical Vibration