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Nuclear Transfer and Cloning

  • Ling Liu
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2066)

Abstract

Nuclear transfer (NT) and cloning offers a unique and powerful experimental tool to study the mechanisms of gene reprogramming, to establish embryonic stem cells from somatic cells, and to clone live offspring. The process of NT involves two different cells. The first are oocytes, which are enucleated and provide the cellular components required for gene reprogramming and early embryo development. The second are donor cells, and their nuclei are injected into the enucleated oocytes. The donor cell nuclei are reprogrammed in the oocyte cytoplasm. The reconstructed oocytes are then activated and will begin to cleave. The embryos now have a genome identical to the original donor cells. These embryos can be used for basic research or implanting into foster mothers to develop to term. The protocols for mouse NT were developed in 1998 with assistance of the piezo drill device, and in 2000 without assistance of the piezo drill device. In this chapter, a comprehensive update on the techniques and protocols employed to generate mice by NT cloning are summarized based on the latest publications.

Key words

Nuclear transfer Cloning Somatic cells Oocytes Oocyte activation Embryo transfer Mouse 

Notes

Acknowledgements

The author would like to thank supervisors, Mrs. Jufeng Qian, Northwest Agriculture University; Dr. Mark Westhusin, Texas A&M University; Dr. Colin McKerlie, Toronto Centre for Phenogenomics; and Mr. Steven Sansing, Charles River Laboratory.

References

  1. 1.
    Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH (1997) Viable offspring derived from fetal and adult mammalian cells. Nature 385(6619):810–813CrossRefGoogle Scholar
  2. 2.
    McCreath KJ, Howcroft J, Campbell KH, Colman A, Schnieke AE, Kind AJ (2000) Production of gene-targeted sheep by nuclear transfer from cultured somatic cells. Nature 405(6790):1066–1069CrossRefGoogle Scholar
  3. 3.
    Loi P, Ptak G, Barboni B, Fulka J Jr, Cappai P, Clinton M (2001) Genetic rescue of an endangered mammal by cross-species nuclear transfer using post-mortem somatic cells. Nat Biotechnol 19(10):962–964CrossRefGoogle Scholar
  4. 4.
    Cibelli JB, Stice SL, Golueke PJ, Kane JJ, Jerry J, Blackwell C, Ponce de Leon FA, Robl JM (1998) Cloned transgenic calves produced from nonquiescent fetal fibroblasts. Science 280(5367):1256–1258CrossRefGoogle Scholar
  5. 5.
    Kato Y, Tani T, Sotomaru Y, Kurokawa K, Kato J, Doguchi H, Yasue H, Tsunoda Y (1998) Eight calves cloned from somatic cells of a single adult. Science 282(5396):2095–2098CrossRefGoogle Scholar
  6. 6.
    Kubota C, Yamakuchi H, Todoroki J, Mizoshita K, Tabara N, Barber M, Yang X (2000) Six cloned calves produced from adult fibroblast cells after long-term culture. Proc Natl Acad Sci U S A 97(3):990–995CrossRefGoogle Scholar
  7. 7.
    Lanza RP, Cibelli JB, Blackwell C, Cristofalo VJ, Francis MK, Baerlocher GM, Mak J, Schertzer M, Chavez EA, Sawyer N, Lansdorp PM, West MD (2000) Extension of cell life-span and telomere length in animals cloned from senescent somatic cells. Science 288(5466):665–669CrossRefGoogle Scholar
  8. 8.
    Baguisi A, Behboodi E, Melican DT, Pollock JS, Destrempes MM, Cammuso C, Williams JL, Nims SD, Porter CA, Midura P, Palacios MJ, Ayres SL, Denniston RS, Hayes ML, Ziomek CA, Meade HM, Godke RA, Gavin WG, Overstrom EW, Echelard Y (1999) Production of goats by somatic cell nuclear transfer. Nat Biotechnol 17(5):456–461CrossRefGoogle Scholar
  9. 9.
    Keefer CL, Baldassarre H, Keyston R, Wang B, Bhatia B, Bilodeau AS, Zhou JF, Leduc M, Downey BR, Lazaris A, Karatzas CN (2001) Generation of dwarf goat (Capra hircus) clones following nuclear transfer with transfected and nontransfected fetal fibroblasts and in vitro-matured oocytes. Biol Reprod 64(3):849–856CrossRefGoogle Scholar
  10. 10.
    Keefer CL, Keyston R, Lazaris A, Bhatia B, Begin I, Bilodeau AS, Zhou FJ, Kafidi N, Wang B, Baldassarre H, Karatzas CN (2002) Production of cloned goats after nuclear transfer using adult somatic cells. Biol Reprod 66(1):199–203CrossRefGoogle Scholar
  11. 11.
    Polejaeva IA, Chen SH, Vaught TD, Page RL, Mullins J, Ball S, Dai Y, Boone J, Walker S, Ayares DL, Colman A, Campbell KH (2000) Cloned pigs produced by nuclear transfer from adult somatic cells. Nature 407(6800):86–90CrossRefGoogle Scholar
  12. 12.
    Onishi A (2002) Cloning of pigs from somatic cells and its prospects. Cloning Stem Cells 4(3):253–259CrossRefGoogle Scholar
  13. 13.
    Lee BC, Kim MK, Jang G, Oh HJ, Yuda F, Kim HJ, Hossein MS, Kim JJ, Kang SK, Schatten G, Hwang WS (2005) Dogs cloned from adult somatic cells. Nature 436(7051):641CrossRefGoogle Scholar
  14. 14.
    Hossein MS, Jeong YW, Park SW, Kim JJ, Lee E, Ko KH, Hyuk P, Hoon SS, Kim YW, Hyun SH, Shin T, Hwang WS (2009) Birth of beagle dogs by somatic cell nuclear transfer. Anim Reprod Sci 114(4):404–414CrossRefGoogle Scholar
  15. 15.
    Hossein MS, Jeong YW, Park SW, Kim JJ, Lee E, Ko KH, Kim HS, Kim YW, Hyun SH, Shin T, Hawthorne L, Hwang WS (2009) Cloning Missy: obtaining multiple offspring of a specific canine genotype by somatic cell nuclear transfer. Cloning Stem Cells 11(1):123–130CrossRefGoogle Scholar
  16. 16.
    Shin T, Kraemer D, Pryor J, Liu L, Rugila J, Howe L, Buck S, Murphy K, Lyons L, Westhusin M (2002) A cat cloned by nuclear transplantation. Nature 415(6874):859CrossRefGoogle Scholar
  17. 17.
    Chesne P, Adenot PG, Viglietta C, Baratte M, Boulanger L, Renard JP (2002) Cloned rabbits produced by nuclear transfer from adult somatic cells. Nat Biotechnol 20(4):366–369CrossRefGoogle Scholar
  18. 18.
    Wakayama T, Perry AC, Zuccotti M, Johnson KR, Yanagimachi R (1998) Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394(6691):369–374CrossRefGoogle Scholar
  19. 19.
    Wakayama T, Yanagimachi R (1999) Cloning of male mice from adult tail-tip cells. Nat Genet 22(2):127–128CrossRefGoogle Scholar
  20. 20.
    Wakayama T, Shinkai Y, Tamashiro KL, Niida H, Blanchard DC, Blanchard RJ, Ogura A, Tanemura K, Tachibana M, Perry AC, Colgan DF, Mombaerts P, Yanagimachi R (2000) Cloning of mice to six generations. Nature 407(6802):318–319CrossRefGoogle Scholar
  21. 21.
    Kafri T, Ariel M, Brandeis M, Shemer R, Urven L, McCarrey J, Cedar H, Razin A (1992) Developmental pattern of gene-specific DNA methylation in the mouse embryo and germ line. Genes Dev 6(5):705–714CrossRefGoogle Scholar
  22. 22.
    Ogura A, Inoue K, Wakayama T (2013) Recent advancements in cloning by somatic cell nuclear transfer. Philos Trans R Soc Lond Ser B Biol Sci 368(1609):20110329CrossRefGoogle Scholar
  23. 23.
    Fulka J Jr, Fulka H (2007) Somatic cell nuclear transfer (SCNT) in mammals: the cytoplast and its reprogramming activities. Adv Exp Med Biol 591:93–102CrossRefGoogle Scholar
  24. 24.
    Schnieke AE, Kind AJ, Ritchie WA, Mycock K, Scott AR, Ritchie M, Wilmut I, Colman A, Campbell KH (1997) Human factor IX transgenic sheep produced by transfer of nuclei from transfected fetal fibroblasts. Science 278(5346):2130–2133CrossRefGoogle Scholar
  25. 25.
    Renard JP, Chastant S, Chesne P, Richard C, Marchal J, Cordonnier N, Chavatte P, Vignon X (1999) Lymphoid hypoplasia and somatic cloning. Lancet 353(9163):1489–1491CrossRefGoogle Scholar
  26. 26.
    Wells DN, Misica PM, Tervit HR (1999) Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells. Biol Reprod 60(4):996–1005CrossRefGoogle Scholar
  27. 27.
    Hill JR, Burghardt RC, Jones K, Long CR, Looney CR, Shin T, Spencer TE, Thompson JA, Winger QA, Westhusin ME (2000) Evidence for placental abnormality as the major cause of mortality in first-trimester somatic cell cloned bovine fetuses. Biol Reprod 63(6):1787–1794CrossRefGoogle Scholar
  28. 28.
    De Sousa PA, King T, Harkness L, Young LE, Walker SK, Wilmut I (2001) Evaluation of gestational deficiencies in cloned sheep fetuses and placentae. Biol Reprod 65(1):23–30CrossRefGoogle Scholar
  29. 29.
    Hill JR, Winger QA, Burghardt RC, Westhusin ME (2001) Bovine nuclear transfer embryo development using cells derived from a cloned fetus. Anim Reprod Sci 67(1–2):17–26CrossRefGoogle Scholar
  30. 30.
    Ono Y, Shimozawa N, Ito M, Kono T (2001) Cloned mice from fetal fibroblast cells arrested at metaphase by a serial nuclear transfer. Biol Reprod 64(1):44–50CrossRefGoogle Scholar
  31. 31.
    Wilmut I (2006) Are there any normal clones? Methods Mol Biol 348:307–318CrossRefGoogle Scholar
  32. 32.
    Jaenisch R, Eggan K, Humphreys D, Rideout W, Hochedlinger K (2002) Nuclear cloning, stem cells, and genomic reprogramming. Cloning Stem Cells 4(4):389–396CrossRefGoogle Scholar
  33. 33.
    Jaenisch R (2009) Stem cells, pluripotency and nuclear reprogramming. J Thromb Haemost 7(Suppl 1):21–23CrossRefGoogle Scholar
  34. 34.
    Colman A, Burley J (2014) Human somatic cell reprogramming: does the egg know best? Cell Stem Cell 15(5):531–532CrossRefGoogle Scholar
  35. 35.
    Tiemann U, Wu G, Marthaler AG, Scholer HR, Tapia N (2016) Epigenetic aberrations are not specific to transcription factor-mediated reprogramming. Stem Cell Reports 6(1):35–43CrossRefGoogle Scholar
  36. 36.
    Wan Y, Deng M, Zhang G, Ren C, Zhang H, Zhang Y, Wang L, Wang F (2016) Abnormal expression of DNA methyltransferases and genomic imprinting in cloned goat fibroblasts. Cell Biol Int 40(1):74–82CrossRefGoogle Scholar
  37. 37.
    Houdebine LM (2000) Transgenic animal bioreactors. Transgenic Res 9(4–5):305–320CrossRefGoogle Scholar
  38. 38.
    Houdebine LM (2005) Use of transgenic animals to improve human health and animal production. Reprod Domest Anim 40(4):269–281CrossRefGoogle Scholar
  39. 39.
    Houdebine LM (2009) Applications of genetically modified animals. J Soc Biol 203(4):323–328CrossRefGoogle Scholar
  40. 40.
    Lewandowski J, Kurpisz M (2016) Techniques of human embryonic stem cell and induced pluripotent stem cell derivation. Arch Immunol Ther Exp 64(5):349–370CrossRefGoogle Scholar
  41. 41.
    Niemann H, Petersen B (2016) The production of multi-transgenic pigs: update and perspectives for xenotransplantation. Transgenic Res 25(3):361–374CrossRefGoogle Scholar
  42. 42.
    Rogers CS (2016) Genetically engineered livestock for biomedical models. Transgenic Res 25(3):345–359CrossRefGoogle Scholar
  43. 43.
    Su F, Wang Y, Liu G, Ru K, Liu X, Yu Y, Liu J, Wu Y, Quan F, Guo Z, Zhang Y (2016) Generation of transgenic cattle expressing human beta-defensin 3 as an approach to reducing susceptibility to Mycobacterium bovis infection. FEBS J 283(5):776–790CrossRefGoogle Scholar
  44. 44.
    Kishigami S, Wakayama S, Thuan NV, Ohta H, Mizutani E, Hikichi T, Bui HT, Balbach S, Ogura A, Boiani M, Wakayama T (2006) Production of cloned mice by somatic cell nuclear transfer. Nat Protoc 1(1):125–138CrossRefGoogle Scholar
  45. 45.
    Zhou Q, Boulanger L, Renard JP (2000) A simplified method for the reconstruction of fully competent mouse zygotes from adult somatic donor nuclei. Cloning 2(1):35–44CrossRefGoogle Scholar
  46. 46.
    Whittingham DG (1971) Culture of mouse ova. J Reprod Fertil Suppl 14:7–21PubMedPubMedCentralGoogle Scholar
  47. 47.
    Chatot CL, Ziomek CA, Bavister BD, Lewis JL, Torres I (1989) An improved culture medium supports development of random-bred 1-cell mouse embryos in vitro. J Reprod Fertil 86(2):679–688CrossRefGoogle Scholar
  48. 48.
    Lawitts JA, Biggers JD (1993) Culture of preimplantation embryos. Methods Enzymol 225:153–164CrossRefGoogle Scholar
  49. 49.
    Summers MC, McGinnis LK, Lawitts JA, Raffin M, Biggers JD (2000) IVF of mouse ova in a simplex optimized medium supplemented with amino acids. Hum Reprod 15(8):1791–1801CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Ling Liu
    • 1
  1. 1.IVF Canada Fertility CenterTorontoCanada

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