DNA sequences coding for a cinnamoyl CoA reductase and their use

Multicellular living organisms and unmodified parts thereof and – Plant – seedling – plant seed – or plant part – per se – Higher plant – seedling – plant seed – or plant part

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800295, 800278, 4353201, 435419, 435468, 536 241, 536 236, C12N 1500, C12N 1529, C12N 1582, A01H 400

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06015943&

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BRIEF SUMMARY
FIELD OF THE INVENTION

A subject of the present invention is the use of DNA sequences coding for a cinnamoyl CoA reductase (CCR) in plants, or any fragment of these sequences, or also any sequence derived from the latter, or their complementary sequences, within the scope of the implementation of processes for the regulation of the lignin level in plants.


BACKGROUND OF THE INVENTION

Lignin is a complex heterogeneous aromatic polymer which waterproofs and reinforces the walls of certain plant cells.
Lignin is formed by the polymerization of free radicals derived from monolignols such as paracoumarylic, coniferylic and sinapylic alcohols (Higuchi, 1985, in Biosynthesis and degradation of wood components (T. Higuchi, ed), Academic Press, Orlando, Fla. pp. 141-160).
Lignins have a large variation in their relative monolignol levels, as a function of the species and the different tissues within the same plant.
This variation is probably due to and controlled by the different activities and specificities of substrates, of enzymes necessary for the biosynthesis of lignin monomers (Higuchi, 1985, mentioned above).
Beyond its role in the structure and development of plants, lignin represents a major component of the terrestrial biomass and assumes a great economic and ecological significance (Brown, 1985, J. Appl. Biochem. 7, 371-387; Whetten and Sederoff, 1991, Forest Ecology and Management, 43, 301-316).
With regard to the exploitation of the biomass, first and foremost it should be noted that lignin is a factor limiting the digestibility and nutritional yield of fodder crops. In fact, it is clearly demonstrated that the digestibility of fodder crops by ruminants, is inversely proportional to the lignin level of these plants, the nature of the lignins also being a determining factor in this phenomenon (Buxton and Roussel, 1988, Crop. Sci., 28, 553-558; Jung and Vogel, 1986, J. Anin. Sci., 62, 1703-1712).
Among the principle fodder crops in which it would be useful to reduce the level of lignins, there can be mentioned: alfalfa, fescue, maize, fodder used for silage . . . .
It should also be noted that high lignin levels are in part responsible for the limited quality of sunflower cakes intended for cattle food and the reduction in the viability of certain seeds in the horticultural domain.
It can also be emphasized that the intense lignification which occurs during the storage of plant organs after harvest, rapidly renders crops such as asparagus, yams, carrots, etc. unfit for consumption.
Moreover, it should also be noted that more than 50 million tons of lignins are extracted from ligneous material each year in the production of paper pulp in the paper industry. This extraction operation which is necessary to obtain cellulose is costly in terms of energy and secondarily polluting due to the chemical compounds employed in the extraction and which find their way into the environment (Dean & Eriksson, 1992, Holzforschung, 46, 135-147; Whetten and Sederoff, 1991, mentioned above).
To reduce the proportions of lignins (which depending on the species represent 20 to 30% of the dry material) to a few percent (2 to 5%) would represent an increase in yield, a substantial saving (chemical products) and would contribute to the improvement of the environment (reduction in pollution). Given the scale on which ligneous material is used, these effects would have extremely significant repercussions. In this case, the species concerned could be the poplar, the eucalyptus, Acacia mangium, the Casuarina genus and all the angiosperms and gymnosperms used in the production of paper pulp.
In the two fields considered, it is clear that the reduction of lignin levels must be moderate in order for the plant (or tree) to retain its characteristics of rigidity and its normal architecture, as the lignins which reinforce the cell walls play an important role in maintaining the upright habit of plants.
Natural variations in the levels of lignins observed in nature for the same species (difference which can range up to 6-8% of the dry ma

REFERENCES:
J.Cell.Biochem. Suppl., vol. 17A, 1993, p. 26, M.M. Campbell, et al. "Hydroxycinnamoyl-coA reductase from Eucalyptus, Molecular analysis of a key control point of lignification", No. A305.
Theoretical and Applied Genetics, vol. 87, No. 8, Mar. 1994, pp. 1006-1015, T.R. Carron, et al., "Genetic Modification of Condensed Tannin Biosynthesis in Lotus Corniculatus.1. Heterologus Antisense Dihydroflavonol Reductase Down-Regulates Tannin Accumulation in `Hairy Root`Cultures".
Bull. Liaison--Groupe Polyphenols, vol. 16 (Pt.2) 1992 pp. 295-300, M.P. Robbins et al. "Manipulation of Condensed Tannin Biosynthesis in Forage Legumes".
EMBL Accession No. T13840, Rel. 38, Mar. 7, 1994, 2005 Arabidopsis Thaliana CDNA Clone 41H9T7.
New Phytologist 129(2), 1995, pp. 203-236, A.M. Boudet, et al., "Tansley review No. 80; Biochemistry and molecular biology of lignification".
Biological Abstracts, vol. 78, 1984, Philadelphia, PA., Abst. No. 14461, F. SARNI, et al., Purification and properties of cinnamoyl-coenzyme A reductase (EC 1.2.1.44) and cinamyl alcohol dehydrogenase (EC 1.1.1.1) from poplar stems (populus euamericana).
Plant Physiology (1994), vol.106, pp. 625-632, Goffner, et al., "Purification and characterization of cinnamoyl-coenzyme A:NADP oxido-reductase in Ecalyptus gunnii".
Plant Molecular Biology, vol. 13, 1989, pp. 491-502, M. Beld, et al. "Flavonoid Synthesis in Petunia Hybrida: Partial Characterization of Dihydroflavonol-4-Reductase Genes".
French Research Report dated Dec. 16, 1994.
Sarni et al. Biological Abstracts vol. 78, 1984, abstract No. 14461, 1984.

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