Analysis of the sex ratio in preimplantation embryos from B6.K1 and B6.K2 Ped gene congenic mice

  • Michael J. Byrne
  • Judith A. Newmark
  • Carol M. WarnerEmail author
Animal Experimentation


Purpose: The mouse preimplantation embryo development (Ped) gene product, Qa-2, which is the homolog of human HLA-G, influences the rate of preimplantation embryonic development and overall reproductive success. The sex ratio in preimplantation embryos from Ped gene congenic mice was examined in order to determine whether embryo sex is a confounding factor in the control of the rate of preimplantation development.

Methods: B6.K1 (Ped slow) and B6.K2 (Ped fast) congenic mice differ only in the absence (B6.K1) or presence (B6.K2) of the genes encoding Qa-2 protein. We analyzed the sex of B6.K1 (n=221) and B6.K2 (n=260) preimplantation embryos by using Real-Time PCR with primers specific for the X and Y chromosomes.

Results: We found that there was no statistically significant difference in the ratio of male to female preimplantation embryos in either strain.

Conclusions: We conclude that the sex of the embryos is not a confounding factor that affects the Ped gene control of the rate of preimplantation development. Therefore, the Ped gene is entirely responsible for mediating the faster development of B6.K2 embryos compared to B6.K1 embryos.


B6.K1 B6.K2 HLA-G IVF Ped gene Preimplantation embryo Qa-2 Sex ratio 



We thank Marie Amato for helping with the literature search and we thank Michele Mammolenti for managing the mouse husbandry. This study was supported by NIH grant HD39215.


  1. 1.
    Boiso I, Veiga A, Edwards RG. Fundamental of human embryonic growth in vitro and the selection of high-quality embryos for transfer. Reprod BioMed Online 2002;5:328–50PubMedGoogle Scholar
  2. 2.
    Wharf E, Dimirakopoulos A, Khalaf Y, Pickering S. Early embryo development is an indicator of implantation potential. Reprod BioMed Online 2004;8:212–18PubMedCrossRefGoogle Scholar
  3. 3.
    Warner CM, Brenner CA. Genetic regulation of preimplantation embryo survival. Curr Top Dev Biol 2001;52:151–92PubMedGoogle Scholar
  4. 4.
    Warner CM, Newmark JA, Comiskey M, De Fazio SR, O’Malley DM, Rajadhyaksha M et al. Genetics and imaging to assess oocyte and preimplantation embryo health. Reprod Fertil Develop 2004;16:729–41Google Scholar
  5. 5.
    Warner CM, Brownell MS, Rothschild MF. Analysis of litter size and weight in mice differing in Ped gene phenotype and the Q region of the H-2 complex. J Reprod Immunol 1991;19:303–13PubMedCrossRefGoogle Scholar
  6. 6.
    Warner CM, Panda P, Almquist CD, Xu Y. Preferential survival of mice expressing the Qa-2 antigen. J Reprod Fertil 1993;99:145–7PubMedCrossRefGoogle Scholar
  7. 7.
    Exley GE, Warner CM. Selection in favor of the Ped fast haplotype occurs between mid-gestation and birth. Immunogenetics 1999;49:653–9PubMedCrossRefGoogle Scholar
  8. 8.
    Brownell MS, Warner CM. Ped gene expression by embryos cultured in vitro. Biol Reprod 1988;39:806–11PubMedCrossRefGoogle Scholar
  9. 9.
    Wu L, Exley GE, Warner CM. Differential expression of Ped gene candidates in preimplantation mouse embryos. Biol Reprod 1998;59:941–52PubMedCrossRefGoogle Scholar
  10. 10.
    Dey SK, Lim H, Das SK, Reese J, Paria BC, Diakoku T et al. Molecular cues to implantation. Endocr Rev 2004;25(3):341–73PubMedCrossRefGoogle Scholar
  11. 11.
    Newmark JA, Sacher F, Jones GS, Warner CM. Ped gene deletion polymorphism frequency in wild mice. J Exptl Zool 2002;293:179–85CrossRefGoogle Scholar
  12. 12.
    Mardon G, Page DC. The sex-determining region of the mouse Y chromosome encodes a protein with a highly acidic domain and 13 zinc fingers. Cell 1989;56:765–70PubMedCrossRefGoogle Scholar
  13. 13.
    Gubbay J, Collignon J, Koopman P, Capel B, Economou A, Munsterberg A, Vivian N, Goodfellow P, Lovell-Badge R. A gene mapping to the sex-determining region of the mouse Y chromosome is a member of a novel family of embryonically expressed genes. Nature 1990;346:245–50PubMedCrossRefGoogle Scholar
  14. 14.
    Love JM, Knight AM, McAleer MA, Todd JA. Towards construction of a high resolution map of the mouse genome using PCR-analyzed microsatellites. Nucleic Acids Res 1990;8:4123–30Google Scholar
  15. 15.
    Kunieda T, Xian M, Kobayashi E, Imamichi T, Moriwaki K, Toyoda Y. Sexing of mouse preimplantation embryos by detection of Y chromosome-specific sequences using polymerase chain reaction. Biol Reprod 1992;46:692–7PubMedCrossRefGoogle Scholar
  16. 16.
    Greenlee AR, Krisher RL, Plotka ED. Rapid sexing of murine preimplantation embryos using a nested multiplex polymerase chain reaction (PCR). Mol Reprod Dev 1998;14:261–7CrossRefGoogle Scholar
  17. 17.
    Cohen and Cohen. Applied multiple regression/Correlation analysis for the behavioral science. 2nd ed. New Jersey: Lawrence Erlbaum Associates, Inc.; 1983, p. 59–61, 529Google Scholar
  18. 18.
    Tsunoda Y, Tokunaga T, Sugie T. Altered sex ratio of live young after transfer of fast- and slow-developing mouse embryos. Gamete Res 1985;12:301–4CrossRefGoogle Scholar
  19. 19.
    Valdivia RPA, Kunieda T, Azuma S, Toyoda T. PCR sexing and developmental rate differences in preimplantation mouse embryos fertilized and cultured in vitro. Mol Reprod Dev 1993;35:121–6PubMedCrossRefGoogle Scholar
  20. 20.
    Peippo P, Bredbacka P. Sex-related growth rate differences in mouse preimplantation embryos in vivo and in vitro. Mol Reprod Dev 1995;40:56–61PubMedCrossRefGoogle Scholar
  21. 21.
    Avery B, Jorgensen CB, Madison V, Greve T. Morphological development and sex of bovine in vitro-fertilized embryos. Mol Reprod Devel 1992;32:265–70PubMedCrossRefGoogle Scholar
  22. 22.
    Xu KP, Yadav BR, King WA, Betteridge KJ. Sex-related differences in developmental rates of bovine embryos produced and cultured in vitro. Mol Reprod Dev 1992;31:240–52CrossRefGoogle Scholar
  23. 23.
    Bernardi ML, Delouis C. Sex-related differences in the developmental rate of in vitro matured/in vitro fertilized ovine embryos. Hum Reprod 1996;11(3):621–6PubMedGoogle Scholar
  24. 24.
    Pergament E, Fiddler M, Cho N, Johnson D, Holmgren WJ. Sexual differentiation and preimplantation cell growth. Hum Reprod 1994;9:1730–2PubMedGoogle Scholar
  25. 25.
    Menezo YJ, Chouteau J, Torello J, Girard A, Veiga A. Birth weight and sex ratio after transfer at the blastocyst stage in humans. Fertil Steril 1999;72(2):221–4PubMedCrossRefGoogle Scholar
  26. 26.
    Tarin JJ, Bernabeu R, Baviera A, Bonada M, Cano A. Sex selection may be inadvertently performed in in-vitro fertilization-embryo transfer programmes. Hum Reprod 1995;10:2992–8PubMedCrossRefGoogle Scholar
  27. 27.
    Burgoyne P. A Y-chromosomal effect on blastocyst cell number in mice. Development 1993;117:341–5PubMedGoogle Scholar
  28. 28.
    Jurisicova A, Casper RF, MacLusky NJ, Mills GB, Librach CL. HLA-G expression during preimplantation human embryo development. Proc Natl Acad Sci USA 1996;A93:161–5CrossRefGoogle Scholar
  29. 29.
    Comiskey M, Goldstein CY, De Fazio SR, Mammolenti M, Newmark JA, Warner CM. Evidence that HLA-G is the functional homolog of Qa-2, the Ped gene product. Hum Immunol 2003;64:999–1004PubMedCrossRefGoogle Scholar
  30. 30.
    Clements CS, Kjer-Nielsen L, Kostenko L, Hoare HL, Dunstone MA, Moses E et al. Crystal structure of HLA-G: a nonclassical MHC class I molecule expressed at the fetal-maternal interface. Proc Natl Acad Sci USA 2005;102(9):3360–5PubMedCrossRefGoogle Scholar
  31. 31.
    Fuzzi B, Rizzo R, Criscuoli L, Noci I, Melchiorri L, Scarselli B et al. HLA-G expression in early embryos is a fundamental prerequisite for the obtainment of pregnancy. Eur J Immunol 2002;32:311–5PubMedCrossRefGoogle Scholar
  32. 32.
    Noci I, Fuzzi B, Rizzo R, Melchiorri L, Criscuoli L, Dabizzi S et al. Embryonic soluble HLA-G as a marker of developmental potential in embryos. Hum Reprod 2004;20(1):138–46PubMedCrossRefGoogle Scholar
  33. 33.
    Sher G, Keskintepe L, Nouriani M, Roussev R, Batzofin J. Expression of sHLA-G in supernatants of individually cultured 46-h embryos: a potentially valuable indicator of ‘embryo competency’ and IVF outcome. Reprod Biomed Online 2004;9(1):74–8PubMedCrossRefGoogle Scholar
  34. 34.
    Yie SM, Balakier H, Motamedi G, Librach CL. Secretion of human leukocyte antigen-G by human embryos is associated with a higher in vitro fertilization pregnancy rate. Fertil Steril 2005;83(1):30–6PubMedCrossRefGoogle Scholar
  35. 35.
    Sher G, Keskintepe L, Fisch JD, Acacio BA, Ahlering P, Batzofin J et al. Soluble human leukocyte antigen G expression in phase I culture media at 46 hours after fertilization predicts pregnancy and implantation from day 3 embryo transfer. Fertil Steril 2005;83(5):1410–3PubMedCrossRefGoogle Scholar
  36. 36.
    Warner CM, Comiskey M, Clisham PR, Brenner CA. Soluble HLA-G (sHLA-G)-A predictor of IVF outcome? J Asst Reprod Gen 2004;21:315–6CrossRefGoogle Scholar
  37. 37.
    Van Lierop MJ, Wijnands F, Loke YW, Emmer PM, Lukassen HG, Braat DD et al. Detection of HLA-G by a specific sandwich ELISA using monoclonal antibodies G233 and 56B. Mol Hum Reprod 2002;8(8):776–84PubMedCrossRefGoogle Scholar
  38. 38.
    Noriko S, Horotsugu H, Masanori Y, Takanori S, Motoko O, Katsuhiko H et al. Are in virto fertilized eggs able to secrete soluble HLA-G? Am J Reprod Immunol 2004;52(Suppl 1):8Google Scholar
  39. 39.
    Blaschitz A, Juch H, Volz A, Hutter H, Daxboeck C, Desoye G et al. The soluble pool of HLA-G produced by human trophoblasts does not include detectable levels of the intron 4-containing HLA-G5 and HLA-G6 isoforms. Mol Hum Reprod 2005;11(10):699–710PubMedCrossRefGoogle Scholar
  40. 40.
    Sargent I. Does ‘soluble’ HLA-G really exist? Another twist to the tail. Mol Hum Reprod 2005;11(10):695–8PubMedCrossRefGoogle Scholar
  41. 41.
    Hviid TVF. HLA-G in human reproduction: aspects of genetics, function and pregnancy complications. Hum Reprod Update 2005;Nov 9: [Epub ahead of print]Google Scholar

Copyright information

© Springer Science+Business Media, Inc. 2006

Authors and Affiliations

  • Michael J. Byrne
    • 1
  • Judith A. Newmark
    • 1
  • Carol M. Warner
    • 1
    Email author
  1. 1.Department of BiologyNortheastern UniversityBostonUSA

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