Skip to main content
SpringerLink
Log in

Effects of chemical activation and season on birth efficiency of cloned pigs

  • Article
  • Published:
Science in China Series C: Life Sciences Aims and scope Submit manuscript

Abstract

The effects of chemical activation on birth efficiency of cloned pigs were studied by investigating the developmental process from porcine oocyte activation to birth of cloned pigs. Three different activation methods were used: (i) Electroporation (Ele); (ii) Ele followed by incubation with 6-dimethylaminopurine (6-DMAP); and (iii) Ele followed by a treatment with cycloheximide (CHX). In experiment 1, the rates of cleavage, developmental rates and cell number of porcine parthenogenetic (PA) embryos were investigated in the three treatment groups. In experiment 2, NT embryos produced by the three different activation treatments were compared for the rates of cleavage, development and cell number. Finally, the effects of Ele and Ele+CHX activation methods on birth efficiency of cloned pigs were compared. The activated oocytes treated by combination activation generally showed a higher (P<0.05) blastocyst rate and produced more expanded blastocysts than oocytes activated with Ele. The rates of cleavage and total cell number of parthenotes were not significantly different. Parthenogenetic embryos activated with 6-DMAP developed into blastocyst and expanded blastocyst stages at a significantly (P<0.05) higher rate than those treated with Ele, but the developmental capability was dramatically decreased in NT embryos. With the CHX activation method, the NT embryo blastocyst rate was substantially (P<0.05) increased although the production of expanded blastocysts was not significantly different from that by the other two methods. The birth rate of cloned pigs increased in the CHX group, though the rate was not significantly different from Ele. The effects of season on developmental rate of the porcine PA embryos and birth rate of cloned pigs were also examined in our study. Porcine oocytes collected in the spring had higher developmental capabilities than those collected in the winter. However, no difference in birth rate of the cloned pigs was found between the oocytes collected in the two seasons. The results obtained from PA and NT embryos, following different activation methods, were inconsistent, suggesting that activation mechanisms are dissimilar in PA and NT embryos. Although the chemical activation in our study leads to an elevation of the blastocyst rate, it does not improve the oocyte’s molecular programming and so does not significantly improve the efficiency of producing cloned pig births.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Nanassy L, Lee K, Javor A, et al. Effects of activation methods and culture conditions on development of parthenogenetic porcine embryos. Anim Reprod Sci, 2008, 104: 264–274, 17320316, 10.1016/j.anireprosci.2007.01.019, 1:CAS:528:DC%2BD1cXhsVOmsb4%3D

    Article  PubMed  CAS  Google Scholar 

  2. 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: 1089–1092, 11778012, 10.1126/science.1068228, 1:CAS:528:DC%2BD38Xht1Gqtbw%3D

    Article  PubMed  CAS  Google Scholar 

  3. Onishi A, Iwamoto M, Akita T, et al. Pig cloning by microinjection of fetal fibroblast nuclei. Science, 2000, 289: 1188–1190, 10947985, 10.1126/science.289.5482.1188, 1:CAS:528:DC%2BD3cXmt1WhsL8%3D

    Article  PubMed  CAS  Google Scholar 

  4. Polejaeva I A, Chen S H, Vaught T D, et al. Cloned pigs produced by nuclear transfer from adult somatic cells. Nature, 2000, 407: 86–90, 10993078, 10.1038/35024082, 1:STN:280:DC%2BD3cvkt12rug%3D%3D

    Article  PubMed  CAS  Google Scholar 

  5. Naruse K, Quan Y S, Kim B C, et al. Brief exposure to cycloheximide prior to electrical activation improves in vitro blastocyst development of porcine parthenogenetic and reconstructed embryos. Theriogenology, 2007, 68: 709–716, 17604096, 10.1016/j.theriogenology.2007.05.065, 1:CAS:528:DC%2BD2sXos1Sgsrs%3D

    Article  PubMed  CAS  Google Scholar 

  6. Lai L, Prather R S. Production of cloned pigs by using somatic cells as donors. Cloning Stem Cells, 2003, 5: 233–241, 14733743, 10.1089/153623003772032754, 1:CAS:528:DC%2BD2cXislaqtw%3D%3D

    Article  PubMed  CAS  Google Scholar 

  7. Im G S, Seo J S, Hwang I S, et al. Development and apoptosis of pre-implantation porcine nuclear transfer embryos activated with different combination of chemicals. Mol Reprod Dev, 2006, 73: 1094–1101, 16736528, 10.1002/mrd.20455, 1:CAS:528:DC%2BD28XnvFahsb0%3D

    Article  PubMed  CAS  Google Scholar 

  8. Varga E, Pataki R, Lorincz Z, et al. Parthenogenetic development of in vitro matured porcine oocytes treated with chemical agents. Anim Reprod Sci, 2008, 105: 226–233, 17391876, 10.1016/j.anireprosci.2007.03.004, 1:CAS:528:DC%2BD1cXjtVWiu78%3D

    Article  PubMed  CAS  Google Scholar 

  9. Lee J W, Tian X C, Yang X. Optimization of parthenogenetic activation protocol in porcine. Mol Reprod Dev, 2004, 68: 51–57, 15039947, 10.1002/mrd.20043, 1:CAS:528:DC%2BD2cXivFyku7Y%3D

    Article  PubMed  CAS  Google Scholar 

  10. Yi Y J, Park C S. Parthenogenetic development of porcine oocytes treated by ethanol, cycloheximide, cytochalasin B and 6-dimethylaminopurine. Animal Reproduction Science, 2005, 86: 297–304, 15766807, 10.1016/j.anireprosci.2004.07.007, 1:CAS:528:DC%2BD2MXit1Cjtbs%3D

    Article  PubMed  CAS  Google Scholar 

  11. Kim N H, Moon S J, Prather R S, et al. Cytoskeletal alteration in aged porcine oocytes and parthenogenesis. Mol Reprod Dev, 1996, 43: 513–518, 9052943, 10.1002/(SICI)1098-2795(199604)43:4<513::AID-MRD14>3.0.CO;2-#, 1:CAS:528:DyaK28XitVGjsLw%3D

    Article  PubMed  CAS  Google Scholar 

  12. Im G S, Samuel M, Lai L, et al. Development and calcium level changes in pre-implantation porcine nuclear transfer embryos activated with 6-DMAP after fusion. Mol Reprod Dev, 2007, 74: 1158–1164, 17366547, 10.1002/mrd.20492, 1:CAS:528:DC%2BD2sXpt1altrw%3D

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  13. Che L, Lalonde A, Bordignon V. Chemical activation of parthenogenetic and nuclear transfer porcine oocytes using ionomycin and strontium chloride. Theriogenology, 2007, 67: 1297–1304, 17350088, 10.1016/j.theriogenology.2007.02.006, 1:CAS:528:DC%2BD2sXjs1OjsL8%3D

    Article  PubMed  CAS  Google Scholar 

  14. Kim Y S, Lee S L, Ock S A, et al. Development of cloned pig embryos by nuclear transfer following different activation treatments. Mol Reprod Dev, 2005, 70: 308–313, 15625691, 10.1002/mrd.20211, 1:CAS:528:DC%2BD2MXhtVShu70%3D

    Article  PubMed  CAS  Google Scholar 

  15. Young, L E, Sinclair K D, Wilmut I. Large offspring syndrome in cattle and sheep. Rev Reprod, 1998, 3: 155–163, 9829550, 10.1530/ror.0.0030155, 1:CAS:528:DyaK1cXntlaltL8%3D

    Article  PubMed  CAS  Google Scholar 

  16. Petters, R M, Wells K D. Culture of pig embryos. J Reprod Fertil, 1993 (Suppl), 48: 61–73, 1:STN:280:DyaK2c7psVCktQ%3D%3D

    CAS  Google Scholar 

  17. 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 C-Life Sci, 2006, 49: 164–171, 16704120, 10.1007/s11427-006-0164-9, 1:CAS:528:DC%2BD28XltFamtbc%3D

    Article  PubMed  CAS  Google Scholar 

  18. Zhang Y H, Pan D K, Sun X Z, et al. In vitro developmental competence of pig nuclear transferred embryos: effects of GFP transfection, refrigeration, cell cycle synchronization and shapes of donor cells. Zygote, 2006, 14: 239–247, 16822335, 10.1017/S0967199406003716, 1:CAS:528:DC%2BD28Xms1elsL8%3D

    Article  PubMed  CAS  Google Scholar 

  19. Zhang K, Wang S H, MaY F, et al. Effects of leptin supplementation in in vitro maturation medium on meiotic maturation of oocytes and preimplantation development of parthenogenetic and cloned embryos in pigs. Anim Reprod Sci, 2007, 101: 85–96, 10.1016/j.anireprosci.2006.08.021, 1:CAS:528:DC%2BD2sXntFyhsLo%3D

    Article  CAS  Google Scholar 

  20. Pan D K, Zhang Y H, SUN X Z, et al. Cloned pigs derived from somatic cell nuclear transfer embryos cultured in vitro at low oxygen tension. Chinese Sci Bull, 2006, 51: 839–844, 10.1007/s11434-006-0839-4, 1:CAS:528:DC%2BD28XjtVKmtLY%3D

    CAS  Google Scholar 

  21. Liu L, Moor R M. Factors affecting electrical activation of porcine oocyte matured in vitro. Anim Reprod Sci, 1997, 48: 67–80, 9412734, 10.1016/S0378-4320(97)00044-4, 1:STN:280:DyaK1c%2FnsVaitg%3D%3D

    Article  PubMed  CAS  Google Scholar 

  22. Whitaker M. Calcium at fertilization and in early development. Physiol Rev, 2006, 86: 25–88, 16371595, 10.1152/physrev.00023.2005, 1:CAS:528:DC%2BD28XhtlKnsbY%3D

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  23. Zimmermann U, Vienken J. Electric field-induced cell-to-cell fusion. J Membr Biol, 1982, 67: 165–182, 7050391, 10.1007/BF01868659, 1:CAS:528:DyaL38XktlGntLk%3D

    Article  PubMed  CAS  Google Scholar 

  24. Grupen C G, Mau J C, McIlfatrick S M, et al. Effect of 6-dimethylaminopurine on electrically activated in vitro matured porcine oocytes. Mol Reprod Dev, 2002, 62: 387–396, 12112604, 10.1002/mrd.10126, 1:CAS:528:DC%2BD38XktlWrsr8%3D

    Article  PubMed  CAS  Google Scholar 

  25. Cha S K, Kim N H, Lee S M, et al. Effect of cytochalasin B and cycloheximide on the activation rate, chromosome constituent and in vitro development of porcine oocytes following parthenogenetic stimulation. Reprod Fertil Dev, 1997, 9: 441–446, 9402254, 10.1071/R96078, 1:CAS:528:DyaK2sXnslyksLY%3D

    Article  PubMed  CAS  Google Scholar 

  26. Li G P, Seidel G E, Squires E L. Intracytoplasmic sperm injection of bovine oocytes with stallion spermatozoa. Theriogenology, 2003, 59: 1143–1155, 12527063, 10.1016/S0093-691X(02)01179-2

    Article  PubMed  Google Scholar 

  27. Li R, Lai L, Wax D, et al. Cloned transgenic swine via in vitro production and cryopreservation. Biol Reprod, 2006, 75: 226–230, 16672718, 10.1095/biolreprod.106.052514, 1:CAS:528:DC%2BD28XnsVWgsLk%3D

    Article  PubMed  CAS  Google Scholar 

  28. Shard M A. Resumption of meiosis: mechanism involved in meiotic progression and its relation with developmental competence. Theriogenology, 2001, 55: 1241–1254, 10.1016/S0093-691X(01)00480-0

    Article  Google Scholar 

  29. Le Beux G, Richard F J, Sirard M A. Effect of cycloheximide, 6-DMAP, roscovitine and butyrolactone I on resumption of meiosis in porcine oocytes. Theriogenology, 2003, 60: 1049–1058, 12935845, 10.1016/S0093-691X(03)00124-9

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing, 100193, China

    YuFang Ma, HengXi Wei, Rui Fang, Kun Zhang, Kai Xue, YanKun Lou & Ning Li

  2. College of Animal Science and Technology, China Agricultural University, Beijing, 100193, China

    YuFang Ma & LinSheng Lian

  3. Beijing Genprotein Biotechnology Company, Beijing, 100193, China

    Yan Li, QiuYan Li, Rui Zhao & YunPing Dai

Authors
  1. YuFang Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yan Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. HengXi Wei
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. QiuYan Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Rui Fang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Rui Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Kun Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Kai Xue
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. YanKun Lou
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. YunPing Dai
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. LinSheng Lian
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Ning Li
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ning Li.

Additional information

Supported by National Key Basic Research and Development Program (China of Grant No. G200000161).

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ma, Y., Li, Y., Wei, H. et al. Effects of chemical activation and season on birth efficiency of cloned pigs. SCI CHINA SER C 52, 657–664 (2009). https://doi.org/10.1007/s11427-009-0087-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11427-009-0087-3

Keywords

Search

Navigation