Abstract
Gene targeting can be used for genetic studies of human cell lines and has significant potential for somatic cell gene therapy. These applications are however restricted by the low frequency of homologous recombination in higher eukaryotes compared to the relatively efficient nonhomologous integration of transfected DNA into the genome. As part of our attempts to overcome this problem, we compared two widely used transfection methods for their efficiency in gene targeting. To our surprise we found that, for conditions that render similar frequencies of nonhomologous integrants, lipofection is much less efficient than electroporation in generating targeted clones. This suggests that nonhomologous and homologous recombination have different requirements for DNA delivery in human cells.
This is a preview of subscription content, log in via an institution to check access.
LITERATURE CITED
Capecchi, M.R. (1989). Science 244:1288–1292.
Koller, B.H., and Smithies, O. (1992). Annu. Rev. Immunol. 10:705–730.
Yáñez, R.J., and Porter, A.C.G. (1998). Gene Ther. 5:149–159.
Porter, A.C.G. (1996). In N.R. Lemoine, and D.N. Cooper, editors, Gene therapy. BIOS Scientific Publishers, Oxford, Great Britain, pp. 169–190.
Itzhaki, J.E., Gilbert, C.S., and Porter, A.C.G. (1997). Nat. Genet. 15:258–265.
Brown, J.P., Wei, W., and Sedivy, J.M. (1997). Science 277:831–834.
Smithies, O., Gregg, R.G., Boggs, S.S., Koralewski, M.A., and Kucherlapati, R.S. (1985). Nature 317: 230–234.
Lin, F.L., Sperle, K., and Sternberg, N. (1985). Proc. Natl. Acad. Sci. U.S.A. 82:1391–1395.
Song, K.Y., Schwartz, F., Maeda, N., Smithies, O., and Kucherlapati, R. (1987). Proc. Natl. Acad. Sci. U.S.A. 84:6820–6824.
Thomas, K.R., Folger, K.R., and Capecchi, M.R. (1986). Cell 44:419–428.
Boggs, S.S., Gregg, R.G., Borenstein, N., and Smithies, O. (1986). Exp. Hematol. 14:988–994.
Nairn, R.S., Adair, G.M., Porter, T., Pennington, S.L., Smith, D.G., Wilson, J.H., and Seidman, M.M. (1993). Somat. Cell Mol. Genet. 19:363–375.
Waldman, B.C., O'Quinn, J.R., and Waldman, A.S. (1996). Biochim. Biophys. Acta. 1308:241–250.
Ellis, J., and Bernstein, A. (1989). Mol. Cell Biol. 9:1621–1627.
Wang, Q., and Taylor, M.W. (1993). Mol. Cell. Biol. 13:918–927.
Mitani, K., Wakamiya, M., Hasty, P., Graham, F.L., Bradley, A., and Caskey, C.T. (1995). Somat. Cell Mol. Genet. 21:221–231.
Russell, D.W., and Hirata, R.K. (1998). Nat. Genet. 18:325–330.
Shirasawa, S., Furuse, M., Yokoyama, N., and Sasazuki, T. (1993). Science 260:85–88.
Waldman, T., Kinzler, K.W., and Vogelstein, B. (1995). Cancer Res. 55:5187–5190.
Yáñez, R.J., and Porter, A.C.G. (1999). Gene Ther. 6:1282–1290.
Kinosita, K., Jr., and Tsong, T.Y. (1977). Biochim. Biophys. Acta. 471:227–242.
Klenchin, V.A., Sukharev, S.I., Serov, S.M., Chernomordik, L.V., and Chizmadzhev Yu, A. (1991). Biophys. J. 60:804–811.
Zabner, J., Fasbender, A.J., Moninger, T., Poellinger, K.A., and Welsh, M.J. (1995). J. Biol. Chem. 270: 18997–19007.
Coonrod, A., Li, F.Q., and Horwitz, M. (1997). Gene Ther. 4:1313–1321.
Dean, D.A. (1997). Exp. Cell Res. 230:293–302.
Loyter, A., Scangos, G., Juricek, D., Keene, D., and Ruddle, F.H. (1982). Exp. Cell Res. 139:223–234.
Fasbender, A., Marshall, J., Moninger, T.O., Grunst, T., Cheng, S., and Welsh, M.J. (1997). Gene Ther. 4:716–725.
Kowalczykowski, S.C., and Eggleston, A.K. (1994). Annu. Rev. Biochem. 63:991–1043.
Chu, G. (1997). J. Biol. Chem. 272:24097–24100.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Yáñez, R.J., Porter, A.C. Influence of DNA Delivery Method on Gene Targeting Frequencies in Human Cells. Somat Cell Mol Genet 25, 27–31 (1999). https://doi.org/10.1023/B:SCAM.0000007137.28557.73
Issue Date:
DOI: https://doi.org/10.1023/B:SCAM.0000007137.28557.73