Production of trehalose in plants

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

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4353201, 435419, 435468, 536 232, 536 237, 800284, 800317, 8003172, A01H 500, A01H 510, C12N 514, C12N 1531, C12N 1552, C12N 1582

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059258047

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BRIEF SUMMARY
FIELD OF THE INVENTION

This invention relates to the modification of plant carbohydrate metabolism using recombinant DNA techniques, recombinant DNA for use therein, as well as plants and parts of plants having a modified genetic constitution. Said plants may be used to extract specific carbohydrate compounds, or alternatively, they may be processed as food, feed, or ingredients thereof, having improved properties due to the presence of said carbohydrate compounds, e.g. during processing.


STATE OF THE ART

Trehalose is a general name given to D-glucosyl D-glucosides which comprise disaccharides based on two .alpha.-,.alpha.,.beta.- and .beta.,.beta.-linked glucose molecules. Trehalose, and especially .alpha.-trehalose 1-(O-a-D-glucopyranosyl)-1'-O-.alpha.-D-glucopyranose) is a widespread naturally occurring disaccharide.
The chemical synthesis of trehalose is difficult (protecting groups required) and inefficient. Current natural sources of trehalose are mushrooms and the yeast Saccharomyces cerevisiae, that can accumulate over 10% of dry weight as trehalose. However production is hampered by high trehalase activity causing rapid metabolization of trehalose. Elbein A. D. (1974, Adv. Carbohydrate Chem. and Biochem. 30, 227-256) gives a review of the occurrence and metabolism of the disaccharide trehalose, particularly .alpha.,.alpha.-trehalose, in living organisms. In plants, the presence of trehalose has been reported in some lower plant species, as well as in a number of higher plant species belonging to the spermatophyta; Echinops persious, Carex brunescens; Fagus silvaticus. However, these results have never been firmly established by other authors (e.g. Kendall et al., 1990, Phytochemistry 29, No. 8, 2525-2528). For instance, Kendall et al, supra, referring to the occurrence of trehalose in spermatophytes, stated that the presence thereof has only been firmly documented for caraway seed (Carum caryi). A report of the presence of trehalose in sunflower by Cegla et al., (1977, J. Am. Oil Chem. Soc. 54, 150 et seq.) was questioned by Kandler et al., (in: The Biochemistry of Plants Vol. 3 Carbohydrates: Structure and Function; Preiss, J., ed., p. 228. Academic Press) according to Kendall et al, 1990, supra. Reports of trehalose in beech (Fagus sylvaticus) and cabbage could not be verified by other authors (Kendall et al., 1990, supra, and references therein).
In spite of the apparent rarity of trehalose in higher plants, the presence of trehalose degrading activities was reported for a significant number of the investigated plant families. Stable high trehalase activity was found in three wheat lines, jack pine, and Selaginella lepidophylla. Stable, low trehalase activity was found in alfalfa, black Mexican sweet corn and white spruce. Labile, moderate activities were found in two different suspensions of canola, but these could probably not be ascribed to specific trehalase activity. Barley, brome grass, soybean and black spruce were reported to contain no trehalase activity at all (Kendall, 1990, supra).
In organisms capable of its production, trehalose is believed to be biosynthesized as the 6-phosphate, whereas the storage form is the free sugar. It is therefore believed, that organisms that produce and/or store trehalose contain a phosphatase capable of cleaving trehalose 6-phosphate (Elbein, 1974, supra). Little is known about the presence of specific trehalose phosphate phosphatases in higher plants.
International patent application WO93/17093 A1, published on Sep. 2, 1993, describes the production of trehalose in yeast transgenic for yeast genes coding for trehalose phosphate synthase. It was suggested that trehalose may be produced in higher plants as well, using these yeast genes, but there is no actual disclosure of trehalose production in a plant. It is to be noted that WO93/17093 A1 was published prior to the filing date but subsequent to the priority date of the instant application.


SUMMARY OF THE INVENTION

The present invention provides for a method for the production of trehalose in a

REFERENCES:
patent: 5422254 (1995-06-01), Londesborough et al.
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