Abstract
The availability of transgenic plant material has provided an important new tool for unraveling the complex mechanisms by which auxins regulate plant development (1–4). The accurate estimation of indole-3-acetic acid (IAA) and IAA-conjugates remains a vital part of the analysis if we are to understand auxin action. Several methods are available utilizing immunoassays, GC-MS, high-performance liquid chromatography (HPLC), and fluorescence. One reliable method for IAA analysis in transgenic material is based on the method of Blakesley et al. (5), which uses a combination of HPLC and fluorimetric detection of the derivatized IAA (see Fig. 1). The straightforward method described has been used successfully for the analysis of IAA and indole-3-acetamide (IAM) in Solanum species and sugar beet hairy roots (1,6) and should be suitable for most plant tissues, although additional purification steps may be required, as explained in the Notes section. This protocol is intended as an introduction to the methodology and the reader is referred to refs. 7 and 8 and Chapter 23 in this volume, which cover the analysis of other metabolites and the methods used.
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References
Gartland, K. M. A., McInnes, E., Hall, J. F., Mulligan, B. J., Morgan, A. J., Elliott, M. C., and Davey, A. R. (1991) Effects of Ri plasmid rol gene expression on the IAA content of transformed roots of Solanum dulcamara L. Plant Growth Regul. 10, 235–241.
Stiller, J., Svoboda, S., Nemcova, B., and Machackova, I. (1992) Effects of agrobacterial oncogenes in kidney vetch (Anthyllis vulneraria L). Plant Cell Rep. 11, 363–367.
Nilsson, O., Crozler, A., Schmulling, T., Sandberg, G., and Olsson, O. (1993) Indole-3-acetic acid homeostasis in transgenic tobacco plants expressing the Agrobacterium rhizogenes rolB gene. Plant J. 3, 681–689.
Van Onckelen, H., Rudelsheim, P., Inze, D., Follin, A., Messens, E., Horemans, S., et al (1985) Tobacco plants transformed with the Agrobacterium T-DNA gene 1 contain high amounts of indole-3-acetamide. FEBS Lett. 181, 373–376.
Blakesley, D., Hall, J. F., Weston, G. D., and Elliott, M. C. (1983) Simultaneous analysis of indole-3-acetic acid and detection of 4-chloroindole-3-acetic acid and 5-hydroxyindole-3-acetic acid in plant tissues by high-performance liquid chromatography of their 2-methylindolo-α-pyrone derivatives. J. Chromatogr. 258, 155–164.
Brown, S. J., Gartland, K. M. A., Slater, A., Hall, J. F., and Elliott, M. C. (1990) Plant growth regulator manipulations in sugar beet, in Progress in Plant Cellular and Molecular Biology (Nijkamp, H. J. J., Van Der Plas, L. W. H., and Van Aartrijk, J., eds.), Kluwer, Dordrecht, The Netherlands, pp. 486–492.
Morgan, P. W. and Durham, J. I. (1983) Strategies for extracting, purifying and assaying auxins from plant tissues. Bot. Gaz. 144, 20–31.
Sandberg, G., Crozier, A., and Ernsten, A. (1987) Indole-3-acetic acid and related compounds, in Principles and Practice of Plant Hormone Analysis, vol. 2 (Rivier, L. and Crozier, A., eds.), Biological Techniques Series, Academic, London, pp. 169–284.
Blakesley, D., Allsopp, A. J. A., Hall, J. F., Weston, G. D., and Elliott, M. C. (1984) Use of reversed-phase ion-pair high performance liquid chromatography for the removal of compounds inhibitory to the formation of the 2-methyl indolo-α-pyrone derivative of indole-3-acetic acid. J. Chromatogr. 294, 480–484.
Crosby, D. G., Boyd, J. B., and Johnson, H. E. (1960) Indole-3-alkanamides. J. Organic Chem. 25, 1826, 1827.
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© 1995 Humana Press Inc., Totowa, NJ
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Hall, J.F., Brown, S.J., Gartland, K.M.A. (1995). IAA Analysis in Transgenic Plants. In: Gartland, K.M.A., Davey, M.R. (eds) Agrobacterium Protocols. Methods in Molecular Biology™, vol 44. Springer, Totowa, NJ. https://doi.org/10.1385/0-89603-302-3:237
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DOI: https://doi.org/10.1385/0-89603-302-3:237
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