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Science in China Series C: Life Sciences

, Volume 45, Issue 3, pp 260–267 | Cite as

Cloned goats (Capra hircus) from adult ear cells

  • Guo Jitong
  • An Zhixing
  • Li Yu
  • Li Xuefeng
  • Li Yuqiang
  • Guo Zekun
  • Zhang Yong
Article
  • 15 Downloads

Abstract

The average number of available oocytes recovered per ovary collected during the breeding season in dairy goats was 5.5 (1815/330). 66.17% (1201/1815) of oocytes extruded the first polar body after maturation in vitro for 20 h. 75.44% (906/1201) of matured oocytes with membrane evagination around the M II chromosomes were enucleated. Ear skin fibroblast cells were derived from an adult female Jining Grey goat (C. hircus). The cells were cryopreserved in liquid nitrogen after passage 2. Thawed cells were further cultured for 3–6 passages and were subjected to serum starvation by 0.5% FBS for 2–10 d, then used as donor cells for nuclear transfer. 98.12% (889/906) of the enucleated oocytes were reconstructed by intracytoplasmic injection of karyoplast. The reconstructed embryos were activated by 5 μmol/L ionomycin for 4.5 min and further activated by culturing with 6-dimethylaminopurine (6-DMAP) for 3 h. After 36 h of culture in mCR1aaBF, 76.69% (645/841) of the cloned embryos cleaved. There were no significant differences in development in vitro between the cloned embryos derived from donor cells precooled at 4°C for 24 h and nonprecooled donor cells. The cleavage rates, 4-cell development, and blastocyst development of reconstructed embryos were 72.48% (79/109), 53.16% (42/79), and 19.05% (8/42) in precooled group; 68.5% (211/308), 59.72% (126/211), and 17.46% (22/126) in nonprecooled group, respectively. Eighteen cloned 4-cell embryos derived from precooled donor cells were transferred and one cloned kid was born. Eighty-four cloned 4-cell embryos derived from nonprecooled donor cells were transferred and no offspring were produced. Of 18 cloned morale from nonprecooled donor cells transferred, one kid was born. The results of microsatellite DNA analyses indicated that the two cloned kids were from the same donor fibroblast cell line derived from an adult goat ear skin.

Keywords

goat ear skin fibroblast cells intracytoplasmic injection nuclear transfer cloned goats 

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References

  1. 1.
    Wilmut, I., Schnieke, A. E., McWhir, J. etal., Viable offspring derived from fetal and adult mammalian cells, Nature, 1997, 385:810.PubMedCrossRefGoogle Scholar
  2. 2.
    Wakayama, T., Perry, A. C., Zuccotti, M. et al., Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei, Nature, 1998, 394:369.PubMedCrossRefGoogle Scholar
  3. 3.
    Kato, Y., Tani, T., Sotomaru, Y. et al., Eight calves cloned from somatic cells of a single adult, Science, 1998, 282:2095.PubMedCrossRefGoogle Scholar
  4. 4.
    Wells, D. N., Misica, P. M., Tervit, H. R. et al., Adult somatic cell nuclear transfer is used to preserve the last surviving cow of the Enderby Island cattle breed, Reprod. Fertil. Dev., 1998, 10: 369.PubMedCrossRefGoogle Scholar
  5. 5.
    Zakhartchenko, V., Alberio, R., Stojkovic, M. et al., Adult cloning in cattle: potential of nuclei from a permanent cell line and from primary cultures, Mol. Reprod. Dev., 1999, 54: 264.PubMedCrossRefGoogle Scholar
  6. 6.
    Shiga, K., Fujita, T., Hirose, K. et al., Production of calves by transfer of nuclei from cultured somatic cells obtained from Japanese black bulls, Theriogenology, 1999, 52: 527.PubMedCrossRefGoogle Scholar
  7. 7.
    Schnieke, A. E., Kind, A. J., Ritchie, W. A. et al., Human factor IX transgenic sheep produced by transfer of nuclei from transfected fetal fibroblasts, Science, 1997, 278: 2130.PubMedCrossRefGoogle Scholar
  8. 8.
    Cibelli, J. B., Stice, S. L., Golueke, P. J. et al., Cloned transgenic calves produced from nonquiescent fetal fibroblasts, Science, 1998, 280: 1256.PubMedCrossRefGoogle Scholar
  9. 9.
    Zakhartchenko, V., Durcova-Hills, G., Stojkovic, M. et al., Effects of serum starvation and re-cloning on the efficiency of nuclear transfer using bovine fetal fibroblasts, J. Reprod. Fertil., 1999, 115: 325.PubMedGoogle Scholar
  10. 10.
    Vignon, X., LeBourhis, D., Chesne, P. et al., Development of bovine nuclear transfer embryos reconstituted with quiescent and proliferative skin fibroblasts, Theriogenology, 1999, 51: 216.CrossRefGoogle Scholar
  11. 11.
    Baguisi, A., Behboodi, E., Melican, D. T. et al., Production of goats by somatic cell nuclear transfer, Nat. Biotechnol., 1999, 17: 456.PubMedCrossRefGoogle Scholar
  12. 12.
    Onishi, A., Iwamoto, M., Akita, T. et al., Pig cloning by microinjection of fetal fibroblast nuclei, Science, 2000, 289: 1188.PubMedCrossRefGoogle Scholar
  13. 13.
    Kubota, C., Yamakuchi, H., Todoroki, J. et al., Six cloned calves produced from adult fibroblast cells after long-term culture, Proc. Natl. Acad. Sci. USA, 2000, 97: 990.PubMedCrossRefGoogle Scholar
  14. 14.
    Wang, Y., Zou, X., Liu, J. et al., Cloned goats (Capra hircus) from foetal fibroblast cell lines, Chinese Science Bulletin, 2000, 45(1): 34.CrossRefGoogle Scholar
  15. 15.
    Keefer, C. L., Baldassarre, H., Keyston, R. et al., Generation of dwarf goat (Capra hircus) clones following nuclear transfer with transfected and nontransfected fetal fibroblasts and in vitro-matured oocytes, Biol, Reprod., 2001, 64: 849.CrossRefGoogle Scholar
  16. 16.
    Guo, J. T., Li, Y. Q., Zhang, Y., Production of recipient oocytes from slaughter house ovaries for nuclear transfer in goat, Acta Agriculturae Boreali-occidentalis Sinica (in Chinese), 1998, 7 (5): 127.Google Scholar
  17. 17.
    Li, Y., Zhang, Y., Guo, J. et al., Dissociation and culture of goat skin cells, J. Agricul. Biotechnol. (in Chinese), 2000, 8(2): 133.Google Scholar
  18. 18.
    Darling, D. C., Morgan, S. J., Animal Cells: Culture and Media, New York: John Wiley and Sons, Inc., 1994.Google Scholar
  19. 19.
    Goto, K., Tanaka, M., Ookutsu, S. et al., Production of a nuclear transferred calf by the intracytoplasmic injection of donor cells, J. Reprod. Dev., 1997, 43: 257.CrossRefGoogle Scholar
  20. 20.
    Rosenkrans, C. F., First, N. L., Culture of bovine zygotes to the blastocyst stage: effects of amino acids and vitamins, Theriogenology, 1991, 35:266.CrossRefGoogle Scholar
  21. 21.
    Arevalo, E., Holder, D. A., Derr, J. N. et al., Caprine microsatellite dinucleotide repeat polymorphisms at the SR-CRSP-1, SR-CRSP-2, SR-CRSP-3, SR-CRSP-4, and SR-CRSP-5 loci, Anim. Genet., 1994, 25: 202.PubMedGoogle Scholar
  22. 22.
    Bhebhe, E., Kogi, J., Holder, D. A. et al., Caprine microsatellite dinucleotide repeat polymorphisms at the SR-CRSP-6, SR-CRSP-7, SR-CRSP-8, SR-CRSP-9, and SR-CRSP-10 loci, Anim. Genet., 1994, 25: 203.PubMedCrossRefGoogle Scholar
  23. 23.
    Yeh, C. C., Kogi, J. K., Holder, M. T. et al., Caprine microsatellite dinucleotide repeat polymorphisms at the SR-CRSP21, SR-CRSP22, SR-CRSP23, SR-CRSP24, SR-CRSP25, SR-CRSP26 and SR-CRSP27 loci, Anim. Genet., 1997, 28: 380.PubMedGoogle Scholar
  24. 24.
    Li, D., Zhou, X., Xu, J. et al., Gene Engineering Manipulation Techniques (in Chinese), Shanghai: Shanghai Science and Technology Press, 1996.Google Scholar
  25. 25.
    Yang, X., Jiang, S., Farrell, P. et al., Nuclear transfer in cattle: effect of nuclear donor cells, cytoplast age, co-culture, and embryo transfer, Mol. Reprod. Dev., 1993, 35:29.PubMedCrossRefGoogle Scholar
  26. 26.
    Liu, L., Ju, J. C., Yang, X., Parthenogenetic development and protein patterns of newly matured bovine oocytes after chemical activation, Mol. Reprod. Dev., 1998, 49: 298.PubMedCrossRefGoogle Scholar
  27. 27.
    Wells, D. N., Misica, P. M., Tervit, H. R., Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells, Biol. Reprod., 1999, 60: 996.PubMedCrossRefGoogle Scholar
  28. 28.
    Kato, Y., Yabuuchi, A., Motosugi, N. et al., Developmental potential of mouse follicular epithelial cells and cumulus cells after nuclear transfer, Biol. Reprod., 1999, 61: 1110.PubMedCrossRefGoogle Scholar

Copyright information

© Science in China Press 2002

Authors and Affiliations

  • Guo Jitong
    • 1
  • An Zhixing
    • 2
  • Li Yu
    • 1
  • Li Xuefeng
    • 2
  • Li Yuqiang
    • 2
  • Guo Zekun
    • 2
  • Zhang Yong
    • 2
  1. 1.College of Life ScienceInner Mongolia UniversityHuhhotChina
  2. 2.Institute of BioengineeringNorthwest Sci-tech University of Agriculture and ForestryYanglingChina

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