Skip to main content
SpringerLink
Log in

Positive-negative selection gene targeting with the diphtheria toxin A-chain gene in mouse embryonic stem cells

  • Papers
  • Published:
Transgenic Research Aims and scope Submit manuscript

Abstract

The diphtheria toxin A-chain gene was used in a positive-negative selection gene targeting vector to alter the CD4 gene which is transcriptionally silent in mouse embryonic stem cells. Expression of the toxin gene was driven by a constitutively active enhancer, yet the targeting construct exhibited only minimal transient toxicity while enriching for targeted clones 9- to 29-fold. Germline transmissiion of the stem cell-derived genome was obtained. These data suggest the usefulness of this diphtheria toxin A-chain cassette in replacement-type positive-negative selection vectors. Its potential for novel applications, particularly in the enrichment for ‘hit-and-run’ insertion-type vectors, is discussed.

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

  • Behringer, R.R., Mathews, L.S., Palmiter, R. D. and Brinster, R.L. (1988) Dwarf mice produced by genetic ablation of growth hormone-expressing cells.Genes Devel. 2, 453–61.

    PubMed  Google Scholar 

  • Bernstein, A., and Breitman, M. (1989) Genetic ablation in transgenic mice.Mol. Biol. Med. 6, 523–30.

    PubMed  Google Scholar 

  • Breitman, M.L., Clapoff, S., Rossant, J., Tsui, L.-C., Glode, L.M., Maxwell, I.H., and Bernstein, A. (1987) Genetic ablation: targeted expression of a toxin gene causes microphthalmia in transgenic mice.Science 238, 1563–5

    PubMed  Google Scholar 

  • Breitman, M.L., Bryce, D.M., Giddens, E., Clapoff, S., Goring, D., Tsui, L.-C., Klintworth, G.K., and Bernstein, A. (1989) Analysis of lens cell fate and eye morphogenesis in transgenic mice ablated for cells of the lens lineage.Development 106, 457–63.

    PubMed  Google Scholar 

  • Bradley, A. (1987) Production and analysis of chimaeras. In Robertson, E.J. ed.,Teratocarcinomas and Embryonic Stem Cells: A Practical Approach, pp. 113–51. Oxford: IRL Press Ltd.

    Google Scholar 

  • Bruggeman, L.A., Hou-Xiang, X., Brown, K.S. and Yamada, Y. (1991) Developmental regulation for collagen II gene expression in transgenic mice.Teratology 44, 203–8.

    Article  PubMed  Google Scholar 

  • Cann, A.J., Koyanagi, Y. and Chen, I.S.Y. (1988) High efficiency transfection of primary human lymphocytes and studies of gene expression.Oncogene 3, 123–8.

    Google Scholar 

  • Capecchi, M.R. (1989) Altering the genome by homologous recombination.Science 244, 1288–92.

    PubMed  Google Scholar 

  • DeChiara, T.M., Efstratiadis, A. and Robertson, E.J. (1990) A growth-deficiency phenotype in heterozygous mice carrying an insulin-like growth factor II gene disrupted by targeting.Nature (London) 345, 78–80.

    Article  Google Scholar 

  • Evans G.A. (1989) Dissecting mouse development with toxigenics.Genes Devel. 3, 259–63.

    PubMed  Google Scholar 

  • Harrington, L., Klintworth, G.K., Secor, T.E. and Breitman, M.L. (1991) Developmental analysis of ocular morphogenesis in αA-crystallin/diphtheria toxin transgenic mice undergoing ablation of the lens.Dev. Biol. 148, 508–16.

    Article  PubMed  Google Scholar 

  • Harrison, G.S., Maxwell, F., Long, C.J., Rosen, C.A., Glode, L.M. and Maxwell, I.H. (1991) Activation of a diphtheria toxin A gene by expression of human immunodeficiency virus-1 Tat and Rev proteins in transfected cells.Hum. Gene Ther. 2, 53–60.

    PubMed  Google Scholar 

  • Hasty, P., Ramírez-Solis, R., Krumlauf, R. and Bradley, A. (1991) Introduction of a subtle mutation into theHox-2.6 locus in embryonic stem cells.Nature (London) 350, 243–6.

    Article  Google Scholar 

  • Kaur, S., Key, B., Stock, J., McNeich, J.D. Akeson, R. and Potter, S.S. (1989) Targeted ablation α-crystallin-synthesizing cells produces lens-deficient eyes in transgenic mice.Development 105, 613–9.

    PubMed  Google Scholar 

  • Knowles, B.B., Koncar, M., Pfizenmaier, K., Solter, D., Aden, D.P. and Trinchieri, G. (1979) Genetic control of the cytotoxic T cell response to SV40 tumor-associated specific antigen.J. Immunol. 122, 1798–806.

    PubMed  Google Scholar 

  • Korfhagen, T.R., Glasser, S.W., Wert, S.E., Bruno, M.D., Daugherty, C.C., McNeish, J.D., Stock, J.L., Potter, S.S. and Whitsett, J.A. (1990)Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice.Proc. Natl Acad. Sci. USA 87, 6122–6.

    PubMed  Google Scholar 

  • Lamb, B.T., Satyamoorthy, K., Li, L., Solter, D. and Howe, C.C. (1991) CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity.Gene Express. 1 185–96.

    Google Scholar 

  • Lee, K.-F., Li, E., Huber, L.J., Landis, S.C., Sharpe, A.H., Chao, M.V. and Jaenisch, R. (1992) Targeted mutation of the gene encoding the low affinity NGF receptor p75 leads to deficits in the peripheral sensory nervous system.Cell 69, 737–49.

    Article  PubMed  Google Scholar 

  • Le Mouellic, H., Lallemand, Y. and Brûlet, P., (1992) Homeosis in the mouse induced by a null mutation in theHox-3.1 gene.Cell 69, 251–64.

    Article  PubMed  Google Scholar 

  • Linney, E., and Donerly, S. (1983) DNA fragments from F9 PyEC mutants increase expression of heterologous genes in transfected F9 cells.Cell 35, 693–9.

    Article  PubMed  Google Scholar 

  • Luckow, B. and Schütz, G. (1987) CAT constructions with multiple unique restriction sites for the functional analysis of eukaryotic promoters and regulatory elements.Nucl. Acids Res. 15, 5490.

    PubMed  Google Scholar 

  • Mansour, S.L., Thomas, K.R. and Capecchi, M.R. (1988) Disruption of the proto-oncogeneint-2 in mouse embryo-derived stem cells: a general strategy for targeting mutations to non-selectable genes.Nature (London) 336, 348–52.

    Article  Google Scholar 

  • Maxwell, I.H., Maxwell, F., and Glode, L.M. (1986) Regulated expression of a diphtheria toxin A-chain gene transfected into human cells: possible strategy for inducing cancer cell suicide.Cancer Res. 46, 4660–4.

    PubMed  Google Scholar 

  • Maxwell, I.H., Glode, L.M. and Maxwell, F. (1991) Expression of the diphtheria toxin A-chain coding sequence under the control of promoters and enhancers from immunoglobulin genes as a means of directing toxicity to B-lymphoid cells.Cancer Res. 51, 4299–304.

    PubMed  Google Scholar 

  • McCarrick, J.W. and Andrews, P.W. (1992) Embryonal carcinoma cells and embryonic stem cells as models of neuronal development and function. In Wood, J.N. ed.,Cell Lines in Neurobiology: A Practical Approach, pp. 77–104. Oxford: Oxford University Press.

    Google Scholar 

  • McMahon, A.P. and Bradley, A. (1990) TheWnt-1 (int-1) proto-oncogene is required for development of a large region of the mouse brain.Cell 62, 1073–85.

    Article  PubMed  Google Scholar 

  • Messing, A., Behringer, R.R., Hammang, J.P., Palmiter, R.D., Brinster, R.L., and Lemke, G. (1992) Po promoter directs expression of reporter and toxin genes to Schwann cells of transgenic mice.Neuron 8, 507–20.

    Article  PubMed  Google Scholar 

  • Neville Jr, D.M. and Hudson, T.H. (1986) Transmembrane transport of diphtheria toxin, related toxins, and colicins.Ann. Rev. Biochem. 55 195–224.

    Article  PubMed  Google Scholar 

  • Palmiter, R.D., Behringer, R.R., Quaife, C.J., Maxwell, F., Maxwell, I.H. and Brinster, R.L. (1987) Cell lineage ablation in transgenic mice by cell-specific expression of a toxin gene.Cell 50, 435–43. (and ErrataCell 62: 608.)

    Article  PubMed  Google Scholar 

  • Rahemtulla, A., Fung-Leung, W.P., Schilham, M.W., Kündig, T.M., Sambhara, S.R., Narendran, A., Arabian, A., Wakeham, A., Paige, C.J., Zinkernagel, R.M., Miller, R.G. and Mak, T.W. (1991) Normal development and function of CD8+ cells but markedly decreased helper cell activity in mice lackingCD4.Nature (London) 353, 180–4.

    Article  Google Scholar 

  • Reid, L.H., Shesely, E.G., Kim, H.-S., and Smithies, O. (1991) Cotransformation and gene targeting in mouse embryonic stem cells.Mol. Cell. Biol. 11, 2769–77.

    PubMed  Google Scholar 

  • Robertson, E.J. (1987) Embryo-derived stem cell lines. In Robertson, E.J. ed.Teratocarcinomas and Embryonic Stem Cells: A Practical Approach, pp. 71–112. Oxford: IRL Press Ltd.

    Google Scholar 

  • Robertson, E., Bradley, A., Kuehn, M. and Evans, M. (1986) Germ-line transmission of genes introduced into cultured pluripotential cells by a retroviral vector.Nature (London) 323, 445–8.

    Article  Google Scholar 

  • Teshigawara, K. and Katsura, Y. (1992) A simple and efficient mammalian gene expression system using an EBV-based vector transfected by electroporation in G2/M phase.Nucl. Acids Res. 20, 2607.

    PubMed  Google Scholar 

  • Tybulewicz, V.L.J., Crawford, C.E., Jackson, P.K., Bronson, R.T. and Mulligan, R.C. (1991) Neonatal lethality and lymphopenia in mice with a homozygous disruption of the c-abl proto-oncogene.Cell 65, 1153–63.

    Article  PubMed  Google Scholar 

  • Valancius, V. and Smithies, O. (1991) Testing an ‘in-out’ targeting procedure for making subtle genomic modifications in mouse embryonic stem cells.Mol. Cell. Biol. 11, 1402–8.

    PubMed  Google Scholar 

  • Yamaizumi, M., Mekada, E., Uchida, T. and Okada, Y. (1978) One molecule of diphtheria toxin fragment A introduced into a cell can kill the cell.Cell 15, 245–50.

    Article  PubMed  Google Scholar 

  • Yagi, T., Ikawa, Y., Yoshida, K., Shigetani, Y., Takeda, N., Mabuchi, I., Yamamoto, T. and Aizawa, S. (1990) Homologous recombination at c-fyn locus of mouse embryonic stem cells with use of diphtheria toxin A-fragment gene in negative selection.Proc. Natl Acad. Sci. USA 87, 9918–22.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. The Wistar Institute of Anatomy and Biology, 19104, Philadelphia, PA, USA

    James W. McCarrick III & Barbara B. Knowles

  2. Division of Immunology and Rheumatology, Department of Medicine, Stanford University School of Medicine, 94305, Stanford, CA, USA

    Jane R. Parnes & Rho H. Seong

  3. Department of Developmental Biology, Max-Planck Institut für Immunobiologie, Stübeweg 51, P.O. Box 1169, D-7800, Freiburg-Zähringen, Germany

    Davor Solter

Authors
  1. James W. McCarrick III
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jane R. Parnes
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rho H. Seong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Davor Solter
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Barbara B. Knowles
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

McCarrick, J.W., Parnes, J.R., Seong, R.H. et al. Positive-negative selection gene targeting with the diphtheria toxin A-chain gene in mouse embryonic stem cells. Transgenic Research 2, 183–190 (1993). https://doi.org/10.1007/BF01977348

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF01977348

Keywords

Search

Navigation