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Production of Transgenic Chimeric Chickens Using Blastodermal Cells

  • Yan, Haifeng (Lab for Molecular Biology Applied to Animal Production, Hunan Institute of Animal and Veterinary Science) ;
  • Lee, Chaeyoung (Ilsong Institute of Life Science, Hallym University) ;
  • Xiao, Bingnan (Lab for Molecular Biology Applied to Animal Production, Hunan Institute of Animal and Veterinary Science) ;
  • Trefil, Pavel (Research Institute of Biopharmacy and Veterinary Drugs) ;
  • Liu, Shixun (Lab for Molecular Biology Applied to Animal Production, Hunan Institute of Animal and Veterinary Science) ;
  • Kim, Younyoung (Ilsong Institute of Life Science, Hallym University) ;
  • Wu, Xiaolin (Lab for Molecular Biology Applied to Animal Production, Hunan Institute of Animal and Veterinary Science)
  • Received : 2004.04.09
  • Accepted : 2004.09.06
  • Published : 2005.02.01

Abstract

A practical approach was proposed to produce transgenic chimeric chickens using blastodermal cells (BCs). The chicken BCs were mechanically dissociated and transferred into the recipient eggs that had been exposed to 500 rads irradiation of$^{60}Co$ and windowed on the equatorial plane. Chimeric chickens were generated using two models: the crosses (MXL) from Black Minors (ii,EE,b/b) ♂${\times}$Barred Leghorns (ii,ee,B/-) ♀ as donors and White Leghorns (WL, II) as acceptors (Model 1), or the Black Heifengs (BH, ii,EE,bb) as donors and Hua-xing white (HW, II) as recipients (Model 2). The treated eggs were incubated in their original shells in normal conditions until hatching. Green fluorescent protein (GFP) gene was transferred into the BCs derived from MXL and BH via lipofectamine and the pEGFP-C1, and transfection efficiency into the BCs was examined under a fluorescent microscope. Potential transgenic chimeras were selected based on the proposed methods in this study. Using the fresh BCs, the best rate of phenotypic chimeras was 6.7% and 26.0% in model-1 groups, and model-2 groups, respectively. We also described the optimized conditions for transfection. Although 30% of the BCs transfected in vitro emitted green light under an inverted fluorescent microscope, no embryos injected with the transfected BCs expressed foreign GFP gene at 3-4 days.

Keywords

Chimeric Chickens;Blastodermal Cells;Transfection;GFP

Acknowledgement

Supported by : Hallym University

References

  1. Etches, R. J., M. E. Clark, A. Toner, G. Liu and A. M. Gibbins. 1996. Contributions to somatic and germline lineages of chicken blastodermal cells maintained in culture. Mol. Reprod. Dev. 45:291-298 https://doi.org/10.1002/(SICI)1098-2795(199611)45:3<291::AID-MRD5>3.0.CO;2-N
  2. Jeong, D. K. and J. Y. Han. 2002. Migration activity of chicken gonadal primordial germ cells (gPGCs) and post-transfer localization of LacZ-transfected gPGCs in the embryonic gonads. Asian-Aust. J. Anim. Sci. 15:1227-1231
  3. Naito, M., M. Sakurai and T. Kuwana. 1998. Expressing of exogenous DNA in the gonads of chimeric chicken embryos produced by transfer of primordial germ cells transfected in vitro and subsequent fate of the introduced DNA. J. Reprod. Fertil. 113:137-143 https://doi.org/10.1530/jrf.0.1130137
  4. Naito, M., A. Sano, Y. Matsubara, T. Harumi, T. Tagami, M. Sakurai and T. Kuwana. 2001. Localization of primordial germ cells or their precursors in stage${\times}$blastoderm of chickens and their ability to differentiate into functional gametes in opposite-sex recipient gonads. Reproduction 121:547-552
  5. Speksnijder, G., R. J. Etches and A. M. Verrinder Gibbins. 1999. Germline chimeric chickens from FACS- sorted donor cells. Mol. Reprd. Dev. 52:33-42
  6. Speksnijder, G. and R. Ivarie. 2000. A modified method of shell windowing for producing somatic or germline chimeras in fertilized chicken eggs. Poult. Sci. 79:1430-1433
  7. Petitte, J. N., M. E. Clark, G. Liu, A. M. Verrinder Gibbins and R. J. Etches. 1990. Production of somatic and germline chimeras in the chicken by transfer of early blastodermal cells. Development 108:185-189
  8. Lee, C. K. and J. A. Piedrahita. 2003. Transgenesis and germ cell engineering in domestic animals. Asian-Aust. J. Anim. Sci. 16:910-927 https://doi.org/10.5713/ajas.2003.910
  9. Qian, F. and C. Z. Xiao. 1999. Comparison of Lipofection and Electroporation Gene Transfer into Mammalian Cells. Prog. Biochem. Biophys. 26:289-291
  10. Trefil, P. 1997. Chicken somatic and germline chimeras. World Poultry-Misset 13:10
  11. Yang, C. C., H. S. Chang, C. J. Lin, C. C. Hsu, J. I. Cheung, L. Hwu and W. T. K. Cheng. 2004. Cock spermatozoa serve as the gene vector for generation of transgenic chicken (Gallus gallus). Asian-Aust. J. Anim. Sci. 17:885-891 https://doi.org/10.5713/ajas.2004.885
  12. Kino, K., B. Pain, S. P. Leibo, M. Cochran, M. E. Clark and R. J. Etches. 1997. Production of chicken chimeras from injection of frozen-thawed Blastodermal cells. Poult. Sci. 76:753-761 https://doi.org/10.1093/ps/76.5.753
  13. Bednarczyk, M., P. Lakota and M. Siwek. 2000. Improvement of hatchability of chicken eggs injected by blastoderm cells. Poult. Sci. 79:1823-1828
  14. Pain, B., M. E. Clark, M. Shen, H. Nakazawa, M. Sakurai, J. Samarut and R. J. Etches. 1996. Long-term in vitro culture and characterisation of avian embryonic stem cells with multiple morphogenetic potentialities. Development 122:2339-2348
  15. Thoraval, P., F. Lasserre, F. Coudert and G. Dambrine. 1994. Somatic and germline chicken chimeras obtained from Brown and White Leghorns by transfer of early blastodermal cells. Poult. Sci. 73:1897-1905
  16. Brazolot, C. L., J. N. Petitte, R. J. Etches and A. M. Gibbins. 1991. Efficient transfection of chicken cells by lipofection, and introduction of transfected blastodermal cells into the embryo. Mol. Reprod. Dev. 30:304-312 https://doi.org/10.1002/mrd.1080300404