Advertisement

Somatic Cell and Molecular Genetics

, Volume 20, Issue 6, pp 515–528 | Cite as

Intragenic regions of the murinePgk-1 locus enhance integration of transfected DNAs into genomes of embryonal carcinoma cells

  • Michael W. McBurney
  • Susan Fournier
  • Karen Jardine
  • Leslie Sutherland
Article

Abstract

Introduction of recombinant genes into mammalian cells in culture has been an important procedure in establishing the molecular mechanisms of various cellular processes. The efficiency with which plasmid borne recombinant genes are expressed following stable integration into genomes of embryonal carcinoma cells is low. Using the P19 embryonal carcinoma cells as recipients, we found that constructs carrying the promoter and intragenic regions of the murinePgk-1 gene were expressed with high efficiency. This elevated expression was associated with increased numbers of copies of the transfected plasmid DNA stably associated with the genomes of recipient cells. The elevated plasmid copy numbers may result from enhanced ligation of transfected plasmids because cotransfected plasmids were also integrated in increased numbers. The enhanced integration and expression of transfected plasmids required active transcription through an intragenic region ofPgk-1, perhaps resulting in more recombinogenic plasmid DNAs.

Keywords

Carcinoma Molecular Mechanism Mammalian Cell Cellular Process Active Transcription 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Literature Cited

  1. 1.
    Adra, C.N., Boer, P.H., and McBurney, M.W. (1987).Gene 60:65–74.Google Scholar
  2. 2.
    Boer, P.H., Potten, H., Adra, C.N., Jardine, K., Mullhofer, G., and McBurney, M.W. (1990).Biochem. Genet. 28:299–308.Google Scholar
  3. 3.
    McBurney, M.W., Sutherland, L.C., Adra, C.N., Leclair, B., Rudnicki, M.A., and Jardine, K. (1991).Nucleic Acids Res. 19:5755–5761.Google Scholar
  4. 4.
    McBurney, M.W., Fournier, S., Schmidt-Kastner, P.K., Jardine, K., and Craig, J. (1995).Somat. Cell Mol. Genet. 20:529–540.Google Scholar
  5. 5.
    Aronow, B., Lattier, D., Silbiger, R., Dusing, M., Hutton, J., Jones, G., Stock, J., McNeish, J., Potter, S., Witte, D., and Wiginton, D. (1989).Genes Dev. 3:1384–1400.Google Scholar
  6. 6.
    Aronow, B.J., Silbiger, R.N., Dusing, M.R., Stock, J.L., Yager, K.L., Potter, S.S., Hutton, J.J., and Wiginton, D.A. (1992).Mol. Cell. Biol. 12:4170–4185.Google Scholar
  7. 7.
    Ottavio, L., Chang, C.-D., Rizzo, M.-G., Travali, S., Casadevall, C., and Baserga, R. (1990).Mol. Cell. Biol. 10:303–309.Google Scholar
  8. 8.
    Sap, J., de Magistris, L., Stunnenberg, H., and Vennstrom, B. (1990).EMBO J. 9:887–896.Google Scholar
  9. 9.
    Ash, J., Ke, Y., Korb, M., and Johnson, L.F. (1993).Mol. Cell. Biol. 13:1565–1571.Google Scholar
  10. 10.
    Hurt, M.M., Bowman, T.L., and Marzluff, W.F. (1991).Mol. Cell. Biol. 11:2929–2936.Google Scholar
  11. 11.
    Lin, H., Yutzey, K.E., and Konieczny, S.F. (1991).Mol. Cell. Biol. 11:267–280.Google Scholar
  12. 12.
    Reid, L.H., Gregg, R.G., Smithies, O., and Koller, B.H. (1990).Proc. Natl. Acad. Sci. U.S.A. 87:4299–4303.Google Scholar
  13. 13.
    McBurney, M.W., and Rogers, B.J. (1982).Dev. Biol. 89:503–508.Google Scholar
  14. 14.
    Rudnicki, M.A., and McBurney, M.W. (1987). InTeratocarcinomas and Embryonic Stem Cells, A Practical Approach, (ed.) Robertson, E.J. (IRL Press, Oxford), pp. 19–49.Google Scholar
  15. 15.
    Chen, C., and Okayama, H. (1987).Mol. Cell. Biol. 7:2745–2752.Google Scholar
  16. 16.
    Southern, P.J., and Berg, P. (1982).J. Mol. Appl. Genet. 1:327–341.Google Scholar
  17. 17.
    Lupton, S.D., Brunton, L.L., Kalberg, V.A., and Overell, R.W. (1991).Mol. Cell. Biol. 11:3374–3378.Google Scholar
  18. 18.
    Vara, J.A., Portela, A., Ortin, J., and Jimenez, A. (1986).Nucleic Acids Res. 14:4617–4624.Google Scholar
  19. 19.
    Norton, P.A., and Coffin, J.M. (1985).Mol. Cell. Biol. 5:281–290.Google Scholar
  20. 20.
    Maniatis, T., Fritsch, T., and Sambrook, J. (1982).Molecular Cloning: A Laboratory Manual, (Cold Spring Harbor Laboratory, Cold Spring Harbor, New York), pp. 1–545.Google Scholar
  21. 21.
    Auffray, C., and Rougeon, F. (1980).Eur. J. Biochem. 107:303–314.Google Scholar
  22. 22.
    Adra, C.N., Ellis, N.A., and McBurney, M.W. (1988).Somat. Cell Mol. Genet. 14:69–81.Google Scholar
  23. 23.
    Pearson, B., Wolf, P.L., and Vazquez, J.A. (1963).Lab. Invest. 12:1249–1259.Google Scholar
  24. 24.
    Mori, N., Singer-Sam, J., Lee, C.Y., and Riggs, A.D. (1986).Gene 45:275–280.Google Scholar
  25. 25.
    Michaelson, A.M., Blake, C.C.F., Evans, S.T., and Orkin, S.H. (1985).Proc. Natl. Acad. Sci. U.S.A. 82:6965–6969.Google Scholar
  26. 26.
    Gorman, C.M., Merlino, G.T., Willingham, M.C., Pastan, I., and Howard, B.H. (1982).Proc. Natl. Acad. Sci. U.S.A. 79:6777–6781.Google Scholar
  27. 27.
    McBurney, M.W., and Strutt, B.J. (1980).Cell 21:357–364.Google Scholar
  28. 28.
    Leavitt, J., Gunning, P., Porreca, P., Ng, S-Y., Lin, C-S., and Kedes, L. (1984).Mol. Cell. Biol. 4:1961–1969.Google Scholar
  29. 29.
    Boer, P.H., Adra, C.N., Lau, Y.F., and McBurney, M.W. (1988).Mol. Cell. Biol. 7:107–3112.Google Scholar
  30. 30.
    Gewert, D.R., Coulombe, B., Castelino, M., Skup, D., and Williams, B.R.G. (1987).EMBO J. 6:651–657.Google Scholar
  31. 31.
    Ng, S-Y., Gunning, P., Eddy, R., Ponte, P., Leavitt, J., Shows, T., and Kedes, L. (1985).Mol. Cell. Biol. 5:2720–2732.Google Scholar
  32. 32.
    Loyter, A., Seangos, G., and Ruddle, F.H. (1982).Proc. Natl. Acad. Sci. U.S.A. 79:422–426.Google Scholar
  33. 33.
    Wigler, M., Sweet, R., Sim, G.K., Wold, B., Pellicer, A., Lacy, E., Maniatis, T., Silverstein, S., and Axei, R. (1977).Cell 16:777–785.Google Scholar
  34. 34.
    Dröge, P. (1993).Proc. Natl. Acad. Sci. U.S.A. 90:2759–2763.Google Scholar
  35. 35.
    Kohwi, Y., and Panchenko, Y. (1993).Genes Dev. 7:1766–1778.Google Scholar
  36. 36.
    Thomas, B.J., and Rothstein, R. (1989).Cell 56:619–630.Google Scholar
  37. 37.
    Nickoloff, J.A. (1992).Mol. Cell. Biol. 12:5311–5318.Google Scholar
  38. 38.
    Heinzel, S.S., Krysan, P.J., Tran, C.T. and Calos, M.P. (1991).Mol. Cell. Biol. 11:2263–2272.Google Scholar
  39. 39.
    Krysan, P.J., and Calos, M.P. (1991).Mol. Cell. Biol. 11:1464–1472.Google Scholar
  40. 40.
    McWhinney, C., and Leffak, M. (1990).Nucleic Acids Res. 18:1233–1242.Google Scholar
  41. 41.
    Landry, S., and Zannis-Hadjopoulos, M. (1991).Biochem. Biophys. Acta 1088:234–244.Google Scholar
  42. 42.
    Krysan, P.J., Smith, J.G., and Calos, M.P. (1993).Mol. Cell. Biol. 13:2688–2696.Google Scholar
  43. 43.
    McArthur, J.G., and Stanners, C.P. (1991).J. Biol. Chem. 266:6000–6005.Google Scholar
  44. 44.
    Lue, N.F., and Kornberg, R.D. (1993).Proc. Natl. Acad. Sci. U.S.A. 90:8018–8022.Google Scholar
  45. 45.
    DePamphilis, M.L. (1993).Annu. Rev. Biochem. 62:29–63.Google Scholar
  46. 46.
    Hoeijmakers, J.H.J., Odijk, H., and Westerveld, A. (1987).Exp. Cell Res. 169:111–119.Google Scholar
  47. 47.
    Monroe, T.J., Muhlmann-Diaz, M.C., Kovach, M.J., Carlson, J.O., Bedford, J.S., and Beaty, B.J. (1992).Proc. Natl. Acad. Sci. U.S.A. 89:5725–5729.Google Scholar
  48. 48.
    Grosveld, F., Blom van Assendelft, G., Greaves, D.R., and Kollias, G. (1987).Cell 51:975–985.Google Scholar
  49. 49.
    Stief, A., Winter, D.M., Strätling, W.H., and Sippel, A.E. (1989).Nature 341:343–345.Google Scholar
  50. 50.
    Palmiter, R.D., Sandgren, E.P., Koeller, D.M., and Brinster, R.L. (1993).Mol. Cell. Biol. 13:5266–5275.Google Scholar
  51. 51.
    Okamoto, K., Okazawa, H., Okuda, A., Sakai, M., Muramatsu, M., and Hamada, H. (1990).Cell 60:461–472.Google Scholar
  52. 52.
    Schöler, H.R., Ruppert, S., Suzuki, N., Chowdhury, K., and Gruss, P. (1990).Nature 344:435–439.Google Scholar
  53. 53.
    Xin, J.-H., Cowie, A., Lachance, P., and Hassell, J.A. (1992).Genes Dev. 6:481–496.Google Scholar
  54. 54.
    Slack, R.S., Hamel, P.A., Bladon, T.S., Gill, R.M., and McBurney, M.W. (1993).Oncogene 8:1585–1591.Google Scholar
  55. 55.
    Linzer, D.I.H., and Levine, A.J. (1979).Cell 17:43–52.Google Scholar
  56. 56.
    Livingstone, L.R., White, A., Sprouse, J., Livanos, E., Jacks, T., and Tlsty, T.D. (1992).Cell 70:923–935.Google Scholar
  57. 57.
    Yin, Y., Tainsky, M.A., Bischoff, F.Z., Strong, L.C., and Wahl, G.M. (1992).Cell 70:937–948.Google Scholar
  58. 58.
    McBurney, M.W., Staines, W.A., Boekelheide, K., Parry, D., Jardine, K., and Pickavance, L. (1994).Dev. Dyn. 200:278–293.Google Scholar
  59. 59.
    Pari, G., Jardine, K., and McBurney, M.W. (1991).Mol. Cell. Biol. 11:4796–4803.Google Scholar
  60. 60.
    Pratt, M.A.C., Langston, A.W., Gudas, L.J., and McBurney, M.W. (1993).Differentiation 53:105–113.Google Scholar

Copyright information

© Plenum Publishing Corporation 1994

Authors and Affiliations

  • Michael W. McBurney
    • 1
  • Susan Fournier
    • 1
  • Karen Jardine
    • 1
  • Leslie Sutherland
    • 1
  1. 1.Department of MedicineUniversity of OttawaOttawaCanada

Personalised recommendations