Advertisement

Chinese Science Bulletin

, Volume 53, Issue 7, pp 1035–1039 | Cite as

Green fluorescent protein (GFP) transgenic pig produced by somatic cell nuclear transfer

  • ZhongHua LiuEmail author
  • Jun Song
  • ZhenKun Wang
  • JiangTian Tian
  • QingRan Kong
  • Zhong Zheng
  • Zhi Yin
  • Li Gao
  • HaiKun Ma
  • Shuang Sun
  • YuTian Li
  • HongBin Wang
  • R. S. Prather
Articles Development Biology

Abstract

Transgenic somatic cell nuclear transfer is a very promising route for producing transgenic farm animals. Research on GFP transgenic pigs can provide useful information for breeding transgenic pigs, human disease models and human organ xenotransplantation. In this study, a liposomal transfection system was screened and transgenic embryos were reconstructed by nuclear transfer of GFP positive cells into enucleated in vitro matured oocytes. The development of reconstructed embryos both in vitro and in vivo was observed, and GFP expression was determined. The results showed that porcine fetal-derived fibroblast cells cultured with 4.0 μL/mL liposome and 1.6 μg/mL plasmid DNA for 6 h resulted in the highest transfection rate (3.6%). The percentage of GFP reconstructed embryos that developed in vitro to the blastocyst stage was 10%. Of those the GFP positive percentage was 48%. Reconstructed transgenic embryos were transferred to 10 recipients. 5 of them were pregnant, and 3 delivered 6 cloned piglets in which 4 piglets were transgenic for the GFP as verified by both GFP protein expression and GFP DNA sequence analysis. The percentage of reconstructed embryos that resulted in cloned piglets was 1.0%; while the percentage of piglets that were transgenic was 0.7%. This is the first group of transgenic cloned pigs born in China, marking a great progress in Chinese transgenic cloned pig research.

Keywords

transgenic green fluorescent protein nuclear transfer liposome pig 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Cibelli J B, Stice S L, Golueke P J, et al. Cloned transgenic calves produced from nonquiescent fetal fibroblasts. Science, 1998, 280:1256–1258PubMedCrossRefGoogle Scholar
  2. 2.
    Wall R J, Powell A M, Paape M J, et al. Genetically enhanced cows resist intramammary Staphylococcus aureus infection. Nat Biotechnol, 2005, 23: 445–451PubMedCrossRefGoogle Scholar
  3. 3.
    McCreath K J, Howcroft J, Campbell K H, et al. Production of gene-targeted sheep by nuclear transfer from cultured somatic cells. Nature, 2000, 405: 1066–1069PubMedCrossRefGoogle Scholar
  4. 4.
    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: 2130–2133PubMedCrossRefGoogle Scholar
  5. 5.
    Lai L, Kolber-Simonds D, Park K W, et al. Production of alpha-1, 3-galactosyltransferase knockout pigs by nuclear transfer cloning. Science, 2002, 295: 1089–1092PubMedCrossRefGoogle Scholar
  6. 6.
    Dai Y, Vaught T D, Boone J, et al. Targeted disruption of the alpha 1,3-galactosyltransferase gene in cloned pigs. Nat Biotechnol, 2002, 20: 251–255PubMedCrossRefGoogle Scholar
  7. 7.
    Hao Y H, Yong H Y, Murphy C N, et al. Production of endothelial nitric oxide synthase (eNOS) over-expressing piglets. Transgenic Res, 2006, 15: 739–750PubMedCrossRefGoogle Scholar
  8. 8.
    Phelps C J, Koike C, Vaught T D, et al. Production of α1,3-galactosyltransferase-deficient pigs. Science, 2003, 299: 411–414PubMedCrossRefGoogle Scholar
  9. 9.
    Cheng Y, Wang Y G, Luo J P, et al. Cloned goats produced from the somatic cells of an adult transgenic goat. Chin J Biotechnol (in Chinese), 2002, 18(1): 79–83Google Scholar
  10. 10.
    Gong G C, Dai Y P, Fan B L, et al. Production of transgenic calves by somatic cell nuclear transfer. Chin Sci Bull, 2004, 49(2): 161–166CrossRefGoogle Scholar
  11. 11.
    Park K W, Cheong H T, Lai L, et al. Production of nuclear transfer-derived swine that express the enhanced green fluorescent protein. Anim Biotechnol, 2001, 12: 173–181PubMedCrossRefGoogle Scholar
  12. 12.
    Lai L, Park K W, Cheong H T, et al. Transgenic pig expressing the enhanced green fluorescent protein produced by nuclear transfer using colchicine-treated fibroblasts as donor cells. Mol Reprod Dev, 2002, 62: 300–306PubMedCrossRefGoogle Scholar
  13. 13.
    Hyun S, Lee G, Kim D, et al. Production of nuclear transfer-derived piglets using porcine fetal fibroblasts transfected with the enhanced green fluorescent protein. Biol Reprod, 2003, 69: 1060–1068PubMedCrossRefGoogle Scholar
  14. 14.
    Lee G S, Kim H S, Hyun S H, et al. Production of transgenic cloned piglets from genetically transformed fetal fibroblasts selected by green fluorescent protein. Theriogenology, 2005, 63: 973–991PubMedCrossRefGoogle Scholar
  15. 15.
    Zhang Y H, Pan D K, Sun X Z, et al. Production of porcine cloned transgenic embryos expressing green fluorescent protein by somatic cell nuclear transfer. Sci China Ser C-Life Sci, 2006, 49(2): 164–171CrossRefGoogle Scholar
  16. 16.
    Park K W, Lai L, Cheong H T, et al. Mosaic gene expression in nuclear transfer-derived embryos and the production of cloned transgenic pigs from ear-derived fibroblasts. Biol Reprod, 2002, 66: 1001–1005PubMedCrossRefGoogle Scholar

Copyright information

© Science in China Press 2008

Authors and Affiliations

  • ZhongHua Liu
    • 1
    Email author
  • Jun Song
    • 1
  • ZhenKun Wang
    • 1
  • JiangTian Tian
    • 1
  • QingRan Kong
    • 1
  • Zhong Zheng
    • 1
  • Zhi Yin
    • 1
  • Li Gao
    • 2
  • HaiKun Ma
    • 2
  • Shuang Sun
    • 1
  • YuTian Li
    • 3
  • HongBin Wang
    • 2
  • R. S. Prather
    • 4
  1. 1.Life Science CollegeNorth-east Agricultural UniversityHarbinChina
  2. 2.Animal Medical CollegeNortheast Agricultural UniversityHarbinChina
  3. 3.Harbin Sanyuan Animal Husbandry Co. Ltd.HarbinChina
  4. 4.Division of Animal ScienceUniversity of Missouri-ColumbiaColumbiaUSA

Personalised recommendations