Dual-Expression Vectors for Efficient Protein Expression in Both E. coli and Mammalian Cells

  • Rebecca L. Mullinax
  • David T. Wong
  • Heidi A. Davis
  • Kerstein A. Padgett
  • Joseph A. Sorge
Part of the Methods in Molecular Biology™ book series (MIMB, volume 205)

Abstract

In the near future, the nucleotide sequence of the genomes from many different organisms will be available. The next and more challenging step will be to characterize the biological role of each gene and the way in which the encoded protein functions in the cell. Dual-expression vectors for expression of proteins encoded by these genes in mammalian and bacterial cells can be used for this characterization. Typically, eukaryotic genes are expressed in mammalian cells to characterize biological functions and in bacterial cells to facilitate isolation of the protein. This generally requires the use of more than one vector. In contrast, use of a dual-expression vector eliminates the need to subclone from one vector system to another by combining the essential features of both eukaryotic and prokaryotic vectors in a single vector.

Keywords

Nickel Agar Codon Electrophoresis Polypropylene 

References

  1. 1.
    Mullinax, R. L., Davis, H. A., Wong, D. T., et al. (2000) Sequence-validated and expression-tested human cDNA in a dual expression vector. Strategies 13, 41–43.Google Scholar
  2. 2.
    Davis, H. A., Wong, D. T., Padgett, K. A., Sorge, J. A., and Mullinax, R. L. (2000) High-level dual mammalian and bacterial protein expression vector. Strategies 13, 136–137.Google Scholar
  3. 3.
    Shine, J. and Dalgarno, L. (1974) The 3′-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc. Natl. Acad. Sci. USA 71, 1342–1346.PubMedCrossRefGoogle Scholar
  4. 4.
    Kozak, M. (1986) Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell 44, 283–292.PubMedCrossRefGoogle Scholar
  5. 5.
    Padgett, K. A. and Sorge, J. A. (1996) Creating seamless junctions independent of restriction sites in PCR cloning. Gene 168, 31–35.PubMedCrossRefGoogle Scholar
  6. 6.
    Evan, G. I., Lewis, G. K., Ramsay, G., and Bishop, J. M. (1985) Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol. Cell Biol. 5, 3610–3616.PubMedGoogle Scholar
  7. 7.
    Hochuli, E., Dobeli, H., and Schacher, A. (1987) New metal chelate adsorbent selective for proteins and peptides containing neighbouring histidine residues. J. Chromatogr. 411, 177–184.PubMedCrossRefGoogle Scholar
  8. 8.
    Felts, F., Rogers, B., Chen, K., Ji, H., Sorge, J., and Vaillancourt, P. (2000) Recombinant Renilla reniformis GFP displays low toxicity. Strategies 13, 85–87.Google Scholar
  9. 9.
    Wynne, K., Wong, D. T., Nioko, V., et al. (2000) Sequence-validated and protein expression-tested human cDNA clones now available. Strategies 13, 133–134.CrossRefGoogle Scholar
  10. 10.
    Sambrook, J., Fritsch, E. F., and Maniatis, T., eds. (1989), Molecular Cloning a Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.Google Scholar
  11. 11.
    Wigler, M., Silverstein, S., Lee, L. S., Pellicer, A., Cheng, Y. and Axel, R. (1977) Transfer of purified herpes virus thymidine kinase gene to cultured mouse cells. Cell 11, 223–232.PubMedCrossRefGoogle Scholar
  12. 12.
    McCutchan, J. H. and Pagano, J. S. (1968) Enchancement of the infectivity of simian virus 40 deoxyribonucleic acid with diethylaminoethyl-dextran. J. Natl. Cancer Inst. 41, 351–357.PubMedGoogle Scholar

Copyright information

© Humana Press Inc. 2003

Authors and Affiliations

  • Rebecca L. Mullinax
    • 1
  • David T. Wong
    • 2
  • Heidi A. Davis
    • 3
  • Kerstein A. Padgett
    • 4
  • Joseph A. Sorge
    • 5
  1. 1.StratageneLa Jlla
  2. 2.GenVaultCarlsbad
  3. 3.The Center for Reproduction of Endangered SpeciesSan Diego
  4. 4.Division of Insect BiologyUniversity of CaliforniaBerkeley
  5. 5.StratageneLa Jolla

Personalised recommendations