Abstract
Molecular and cellular processes related to the senescence syndrome are determined by programs of differential gene expression. Subtractive cDNA hybridization is a powerful approach to identify and isolate differentially expressed genes in various systems. A highly effective method, termed suppression subtractive hybridization (SSH), has been applied for the generation of subtracted cDNA library of senescing leaves. The method consists of two main stages, the normalization step that equalizes the abundance of cDNAs within the target population and the subtraction step that eliminates the common sequences between the target and the driver populations. The successful generation of library containing high numbered of rare and abundant cDNA clones in senescing plant cells proves the applicability of this method for global identification of differentially expressed genes during cellular senescence.
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References
Buchanan-Wollaston, V. (1994) Isolation of cDNA clones for genes that are expressed during leaf senescence in Brassica napus. Identification of a gene encoding a senescence-specific metallothionein-like protein. Plant Physiol. 105, 839–846.
Drake, R, John, I., Fanell, A., Cooper, W., Schuch, W., and Grierson, D. (1996) Isolation and analysis of cDNAs encoding tomato cysteine proteases expressed during leaf senescence. Plant Mol. Biol. 30, 755–767.
Park, J. H., Oh, S. A., Kim, Y. H., Woo, H. R., and Nam, H. G. (1998) Differential expression of senescence-associated mRNAs during leaf senescence induced by different senescence-inducing factors in Arabidopsis. Plant Mol. Biol. 37, 445–454.
Smart, C. M, Hosken, S. E., Thomas, H., Greaves, J. A., Blair, B. G., and Shcuch, W. (1995) The timing of maize leaf senescence and characterization of senescence-related cDNAs. Physiologia Plantarum 93, 673–682
Buchanan-Wollaston, V. and Ainsworth, C. (1997) Leaf senescence in Brassica napus: cloning of senescence related genes by subtractive hybridisation. Plant Mol. Biol. 33, 821–834.
Fujiki, Y., Yoshikawa, Y., Sato, T., et al. (2001) Dark-inducible genes from Arabidopsis thaliana are associated with leaf senescence and repressed by sugars. Physiol. Plant 111, 345–352.
Hajouj, T., Michelis, R., and Gepstein, S. (2000) Cloning and characterization of a receptor-like protein kinase gene associated with senescence. Plant Physiol. 124, 1305–1314.
Kleber-Janke, T. and Krupinska, K. (1997) Isolation of cDNA clones for genes showing enhanced expression in bailey leaves during dark-induced senescence as well as during senescence under field conditions. Planta 203, 332–340.
Yoshida, S., Ito, M., Nishida, I., and Watanabe, A. (2001) Isolation and RNA gel blot analysis of genes that could serve as potential molecular markers for leaf senescence in Arabidopsis maliana. Plant Cell Physiol. 42, 170–178.
Page, T., Griffiths, G., and Buchanan-Wollaston, V. (2001) Molecular and biochemical characterization of postharvest senescence in broccoli. Plant Physiol. 125, 718–727.
Guterman, A., Hajouj, T., and Gepstein, S. (2003) Senescence-associated mRNAs that may participate in signal transduction and protein trafficking. Physiol. Plant 118, 439–446.
Hinderhofer, K. and Zentgraf, U. (2001) Identification of a transcription factor specifically expressed at the onset of leaf senescence. Planta 213, 469–473.
Gepstein, S., Sabehi, G., Carp, M. J., et al. (2003) Large-scale identification of leaf senescence-associated genes. Plant J. 36, 629–642.
Diatchenko, L., Lau, Y. F., Campbell, A. P., et al. (1996) Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries. Proc. Natl. Acad. Sci. USA 93, 6025–6030.
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© 2007 Humana Press Inc., Totowa, NJ
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Sternberg, M.B., Gepstein, S. (2007). Subtractive Hybridization Techniques to Study Cellular Senescence. In: Tollefsbol, T.O. (eds) Biological Aging. Methods in Molecular Biology™, vol 371. Humana Press. https://doi.org/10.1007/978-1-59745-361-5_21
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DOI: https://doi.org/10.1007/978-1-59745-361-5_21
Publisher Name: Humana Press
Print ISBN: 978-1-58829-658-0
Online ISBN: 978-1-59745-361-5
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