Abstract
Current research into the structure and function of plant genes involves the application of many elaborate techniques for gene cloning and analysis. The isolation of pure, intact plant mRNA is required at many stages in this process, e.g., for generation and screening of cDNA clones, for characterization and mapping of cloned genes, and for the study of gene expression. The isolation of RNA from plants, however, often presents more problems than many of the sophisticated procedures used subsequently. Not only is there the problem of RNA degradation by endogenous and exogenous nucleases (common to RNA isolation from any organism), there is also the particular problem of extracting RNA from plant material that may be rich in starch, pectins, phenolics, and various other secondary products. Many of these plant products are not removed during conventional phenol extraction procedures, such as the one described previously in this series (1). Clearly the problems encountered in obtaining clean, undegraded RNA from any particular plant tissue will vary according to the level of nuclease activity and the range of possible contaminants, and it might be expected that different solutions would be required in each case.
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Slater, R.J. (1985) The extraction of total RNA by the detergent and phenol method, in Methods in Molecular Biology Vol. 2 (Walker, J., ed.) Humana, Clifton, New Jersey.
Grierson, D., Slater, A., Spiers, J., and Tucker, G. A. (1985) The appearance of polygalacturonase mRNA in tomatoes: One of a series of changes in gene expression during development and ripening. Planta 163, 263–271.
Speirs, J., Brady, C.J., Grierson, D., and Lee, E. (1984) Changes in ribosome organisation and messenger RNA abundance in ripening tomato fruits. Aust. J. Plant Physiol 11, 225–233.
Palmiter, R.D. (1974) Magnesium precipitation of ribonucleoprotein complexes. Expedient techniques for the isolation of undegraded polysomes and messenger ribonucleic acid. Biochemistry 13, 3606–3615.
Mozer, T.J. (1980) Partial purification and characterisation of the mRNA for a-amylase from barley aleurone layers. Plant Physiol. 65, 834–837.
Slater, A., Maunders, M.J., Edwards, K., Schuch, W., and Grierson, D. (1985) Isolation and characterisation of cDNA clones for tomato polygalactouronase and other ripening-related proteins. Plant Mol. Biol. 5, 137–147.
Maunders, M.J., Slater, A., and Grierson, D. (1985) Generation and Use of cDNA Clones for Studying Gene Expression, in Molecular Biology and Biotechnology (Walker, J.M. and Gingold, E.M., eds.) Royal Society of Chemistry, London.
Slater, A. (1987) Hybrid-Release Translation, in Methods in Molecular Biology, Vol. 4 (Walker, J., ed.) Humana, Clifton, New Jersey.
Kitos, P.A., Saxon, G., and Ames, H. (1972) The isolation of polyadenylate with unreacted cellulose. Biochem. Biophys. Res. Comm. 47, 1426–1437.
Holley, R.W., Apgar, J., and Merrill, S.H. (1961) Evidence for the liberation of a nuclease from human fingers. J. Biol. Chem. 236, PC42.
Maniatis, T., Fritsch, E.F., and Sambrook, J. (1982) Molecular Cloning, A Laboratory Manual Cold Spring Harbor Laboratories, Cold Spring Harbor, New York.
Slater, R.J. (1985) Purification of Poly (A)-Containing RNA by Affinity Chromatography, Methods in Molecular Biology Vol. 2 (Walker, J., ed.) Humana, Clifton, New Jersey.
Davies, E., Larkins, B.A., and Knight, R.H. (1972) Polyribosomes from peas. An improved method for their isolation in the absence of ribonuclease inhibitors. Plant Physiol. 50, 581–584.
Gray, J.C. (1974) The inhibition of ribonuclease activity and the isolation of polysomes from leaves of the French bean, Phaseolus vulgaris. Arch. Biochem. Biophys. 163, 343–348.
Blackburn, P., Wilson, G., and Moore, S. (1977) Ribonuclease inhibitor from human placenta. J. Biol. Chem. 252, 5904–5910.
Smith, C.J.S., Slater, A., and Grierson, D. (1986) Rapid appearance of an mRNA correlated with ethylene synthesis encoding a protein of molecular weight 35,000. Planta 168, 94–100.
Loomis, W.D. (1974) Overcoming problems of phenolics and quinones in the isolation of plant enzymes and organelles. Meth. Enzymol. 31, 528–544.
McGookin, R. (1985) RNA Extraction by the Guanidine Thiocyanate Procedure, in Methods in Molecular Biology, Vol. 2 (Walker, J., ed.) Humana, Clifton, New Jersey.
Nelson, C.E. and Ryan, C. A. (1980) In vitro synthesis of pre-proteins of vacuolar compartmented proteinase inhibitors that accumulate in leaves of wounded tomato plants. Proc. Natl. Acad. Sci. USA 77, 1975–1979.
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© 1988 The Humana Press Inc.
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Slater, A. (1988). Extraction of RNA from Plants. In: Walker, J.M. (eds) New Nucleic Acid Techniques. Methods in Molecular Biology, vol 4. Humana Press. https://doi.org/10.1385/0-89603-127-6:437
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DOI: https://doi.org/10.1385/0-89603-127-6:437
Publisher Name: Humana Press
Print ISBN: 978-0-89603-127-2
Online ISBN: 978-1-59259-491-7
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