Transgenic Plants: Methods and Protocols pp 203-213

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Histochemical and Fluorometric Assays for uidA (GUS) Gene Detection

  • Magdalena Cervera


Transgenic plant production has been intimately connected to the β-glucuronidase (uidA or GUS) gene used as a reporter marker gene. The enzyme stability and the high sensitivity and amenability of the GUS assay to qualitative (histochemical assay) and to quantitative (fluorometric or spectrophotometric assay) detection are some of the reasons that explain the extensive use of uidA gene in plant genetic transformation. Methods for uidA gene detection have been thoroughly described in the literature. The aim of this chapter is to describe the basic protocols needed for GUS detection in a plant genetic transformation laboratory.

Key Words

Fluorometric GUS detection β-glucuronidase GUS histochemical GUS detection reporter marker genes uidA gene 


  1. 1.
    Novel, G. and Novel, M. (1973) Mutants d’Escherichia coli affectés pour leur croissance sur methyl β-glucuronide: localisation du gene de structure de la β-glucuronidase (uidA). Mol. Gen. Genet. 120, 319–335.PubMedGoogle Scholar
  2. 2.
    Jefferson, R. A., Kavanagh, T. A., and Bevan, M. W. (1987) GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. Embo J. 6, 3901–3907.PubMedGoogle Scholar
  3. 3.
    Jefferson, R. A., Bevan, M., and Kavanagh, T. (1987) The use of the Escherichia coli beta-glucuronidase as a gene fusion marker for studies of gene expression in higher plants. Biochem. Soc. Trans. 15, 17–18.PubMedGoogle Scholar
  4. 4.
    Hu, C.-Y., Chee, P. P., Chesney, R. H., Zhou, J. H., and Miller, P. D. (1990) Intrinsic GUS-like activities in seed plants. Plant Cell Rep. 9, 1–5.CrossRefGoogle Scholar
  5. 5.
    Kosugi, S., Ohashi, Y., Nakajima, K., and Arai, Y. (1990) An improved assay for β-glucuronidase in transformed cells: methanol almost completely suppresses a putative endogenous β-glucuronidase activity. Plant Sci. 70, 133–140.CrossRefGoogle Scholar
  6. 6.
    Mascarenhas, J. P. and Hamilton, D. A. (1992) Artifacts in the localization of GUS activity in anthers of petunia transformed with a CaMV 35S-GUS construct. Plant J. 2, 405–408.CrossRefGoogle Scholar
  7. 7.
    Muhitch, M. J. (1998) Characterization of pedicel β-glucuronidase activity in developing maize (Zea mays) kernels. Physiol. Plant. 104, 423–430.CrossRefGoogle Scholar
  8. 8.
    Jefferson, R. A., Burgess, S. M., and Hirsh, D. (1986) β-Glucuronidase from Escherichia coli as a gene-fusion marker. Proc. Natl. Acad. Sci. USA 83, 8447–8451PubMedCrossRefGoogle Scholar
  9. 9.
    Thomasset, B., Ménard, M., Boetti, H., Denmat, L. A., Inzé, D., and Thomas, D. (1996) β-Glucuronidase activity in transgenic and non-transgenic tobacco cells: specific elimination of plant inhibitors and minimization of endogenous GUS background. Plant Sci. 113, 209–219.CrossRefGoogle Scholar
  10. 10.
    Serres, R., McCown, B., and Zeldin, E. (1997) Detectable β-glucuronidase activity in transgenic cranberry is affected by endogenous inhibitors and plant development. Plant Cell Rep. 16, 641–646.Google Scholar
  11. 11.
    Tör, M., Mantell, S. H., and Ainsworth, C. (1992) Endophytic bacteria expressing β-glucuronidase cause false positives in transformation of Dioscorea species. Plant Cell Rep. 11, 452–456.CrossRefGoogle Scholar
  12. 12.
    Vancanneyt, G., Schmidt, R., O’Connor-Sanchez, A., Willmitzer, L., and Rocha-Sosa, M. (1990) Construction of an intron-containing marker gene: splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacteriummediated plant transformation. Mol. Gen. Genet. 220, 245–250.PubMedCrossRefGoogle Scholar
  13. 13.
    Kirchner, G., Kinslow, C. J., Bloom, G. C., and Taylor, D. W. (1993) Non-lethal assay system of β-glucuronidase activity in transgenic tobacco roots. Plant Mol. Biol. Rep. 11, 320–325.CrossRefGoogle Scholar
  14. 14.
    Martin, T., Wöner, R.-V., Hummel, S., Willmitzer, L., and Frommer, W. B. (1992) The GUS reporter system as a tool to study plant gene expression, in GUS Protocols: Using the GUS Gene as a Reporter of Gene Expression (Gallagher, S. R., ed.), Academic Press, San Diego, CA, pp. 23–43.Google Scholar
  15. 15.
    Stewart, C. N., Jr. (2001) The utility of green fluorescent protein in transgenic plants. Plant Cell Rep. 20, 376–382.PubMedCrossRefGoogle Scholar
  16. 16.
    Gallagher, S. R., ed. (1992) GUS Protocols: Using the GUS Gene as a Reporter of Gene Expression, Academic Press, San Diego, CA.Google Scholar
  17. 17. Stomp, A.-M. (1992) Histochemical localization of β-glucuronidase, in GUS Protocols: Using the GUS Gene as a Reporter of Gene Expression (Gallagher, S. R., ed.), Academic Press, San Diego, CA, pp. 103–113.Google Scholar
  18. 18.
    Wilkinson, J. E., Twell, D., and Lindsey, K. (1994) Methanol does not specifically inhibit endogenous β-glucuronidase (GUS) activity. Plant Sci. 97, 61–67.CrossRefGoogle Scholar
  19. 19.
    Naleway, J. J. (1992) Histochemical, spectrophotometric, and fluorometric GUS substrates, in GUS Protocols: Using the GUS Gene as a Reporter of Gene Expression (Gallagher, S. R., ed.), Academic Press, San Diego, CA, pp. 61–76.Google Scholar
  20. 20.
    Guivarc’h, A., Caissard, J. C., Azmi, A., Elmayan, T., Chriqui, D., and Tepfer, M. (1996) In situ detection of expression of the gus reporter gene in transgenic plants: ten years of blue genes. Transgen. Res. 5, 281–288.CrossRefGoogle Scholar

Copyright information

© Humana Press Inc. 2005

Authors and Affiliations

  • Magdalena Cervera
    • 1
  1. 1.Department of Plant Protection and BiotechnologyInstituto Valenciano de Investigaciones AgrariasValenciaSpain

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