Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide alters carbohydrate production in the plant
Reexamination Certificate
1999-06-22
2001-02-27
Fox, David T. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide alters carbohydrate production in the plant
C435S069100, C435S320100, C435S419000, C435S468000, C536S023200, C536S024100, C536S023600, C536S056000, C536S024500, C800S286000, C800S278000, C800S295000, C800S298000, C800S305000, C800S306000, C800S312000, C800S314000, C800S315000, C800S317100, C800S317400, C800S320000, C800S320100, C800S320200, C800S320300
Reexamination Certificate
active
06194638
ABSTRACT:
TECHNICAL FIELD
The present invention relates generally to plant molecular biology. More specifically, it relates to nucleic acids and methods for modulating their expression in plants.
BACKGROUND OF THE INVENTION
The disclosure of U.S. Ser. No. 60/090,416 is hereby incorporated by reference.
Mature plant cell walls generally contain about 30-40% hemicellulose. Hemicellulose in monocot walls consists mainly of arabinoxylan (also referred to as glucurono-arabinoxylan or pentosans), a branched polymer consisting of &bgr;-gylosyl backbone decorated with arabinosyl and glucuronosyl residues. This polymer is often referred to as arabinoxylan because of the relatively low proportion of glucuronosyl residues. Carpita, N. C., “Structure and Biogenesis of the Cell Walls of Grasses”
Annual Review of Plant Physiol. and Plant Molecular Biology
476:445-476 (1996).
Dicot cell walls, in contrast, contain xyloglucan as the main hemicellulosic polymer. Xyloglucan is also a branched polymer consisting of a linear &bgr;-glycosyl backbone which is decorated with xylosyl residues, some of which residues are substituted with galactosyl residues.
Cell walls of corn grain constitute about 6-8% of the total dry weight for both coarse and fine fiber. Whistler, R. L. et al,
Hemicelluloses In Industrial Gums: Polysaccharides and Their Derivatives,
San Diego: Academic Press, pp. 295-308 (1993). The grain cell wall contains 45-65% arabinoxylan, the remainder being mainly cellulose. Under certain circumstances it would be advantageous to reduce the concentration of arabinoxylan in the grain cell wall.
Arabinoxylans are considered to be anti-nutritional components of animal feed. This is because they can absorb large amounts of water, leading to increased viscosity and possible sequestering of other digestible feed components, such as starch and proteins, away from digestive enzymes. Arabinoxylans are also known to lower the food conversion ratio (FCR). In studies where fungal xylanase is included in cereal-derived feed, improved FCR and weight gain in broilers and pigs (Veldman and Vahl, H. A., “Xylanase in broiler diets with differences in characteristics and content of wheat”
British Poultry Science
35:537-550 (1994). The improvement observed for FCR and weight gain is higher than can be expected from arabinoxylan breakdown and digestion alone. Thus, arabinoxylan appears to reduce the digestion of other feed components. Therefore, it would be desirable to reduce the concentration of arabinoxylan in grain.
Cellulose microfibrils have the highest tensile strength of any of the other cell wall polymers. Increasing the cellulose/arabinoxylan ratio in the cell wall should lead to a harder pericarp and improved handling ability of the grain.
About 12% of the corn processed through wet-milling is fiber. Of this 12%, approximately 4% is coarse fiber (mainly pericarp-derived) and the rest is fine fiber. Whereas the coarse fiber consists of approximately 9% starch, fine fiber can have up to 30% starch which constitutes 2.5% of the total dry mass. The other major fraction of fine fiber is arabinoxylan which makes up to 21% of this fraction. It is likely that some of the starch becomes intricately associated with arabinoxylan, thereby making it difficult to extract during wet-milling. Thus, the need arises for a method to reduce the concentration of arabinoxylan in grain to improve starch extractibility.
Under other circumstances, it is desirable to increase the concentration of arabinoxylan in grain. Dietary fiber in grains from a number of plant species grown for human consumption has been shown to reduce cholesterol and low density lipoprotein levels in the plasma. Corn soluble fiber (arabinoxylan) has been shown to be superior to that of wheat in rats in lowering cholesterol levels. In the case of crops such as corn, grown for human consumption, it would therefore be desirable to increase the arabinoxylan content.
There are markets for isolated arabinoxylan preparations for various industrial use, i.e., as thickeners, emulsifiers or stabilizers in food, cosmetics, and pharmaceuticals. Whistler, supra. Increasing the concentration of arabinoxylan in corn grain targeted for wet-milling would make the process of arabinoxylan isolation more economical.
Finally, the modulation of hemicellulose content can be utilized to control plant growth. For example, plant growth is determined by concerted synthesis of cell wall polymers. It is expected that increased synthesis of one of the cell wall polymers, such as hemicellulose, will cause an increase in the synthesis of the rest of the polymers as well. It is expected that increased production of arabinoxylan or xyloglucan in vegetative tissue will accelerate plant growth. In contrast, it is expected that decreased production of arabinoxylan or xyloglucan will inhibit the synthesis of other cell wall polymers. In this regard, reduced production of arabinoxylan will slow plant growth. Additionally, tissue-specific control of hemicellulose productivity is used to modify plant organ growth and development. Early flowering, larger fruit size, or stronger stalk or stem quality is achieved by operably linking a tissue specific promoter to an Rgp gene the expression of which increases hemicellulose biosynthesis.
In view of the foregoing, it would be desirable to modulate the arabinoxylan and xyloglucan concentration in crop plants.
SUMMARY OF THE INVENTION
Generally, it is the object of the present invention to provide nucleic acids and proteins relating to hemicellulose production. It is an object of the present invention to provide: 1) antigenic fragments of the proteins of the present invention; 2) transgenic plants comprising the nucleic acids of the present invention; 3) methods for modulating, in a transgenic plant, the expression of the nucleic acids of the present invention.
It is another object of the present invention to provide methods for decreasing the concentration of arabinoxylan in the grain of a crop plant.
It is yet another object of the present invention to provide methods for producing livestock feed grain with a higher feed conversion ratio.
It is yet another object of the present invention to provide methods for improving feed grain handling quality.
It is yet another object of the present invention to provide methods for producing grain with improved starch extractibility.
It is an object of the present invention to provide methods for increasing the concentration of arabinoxylan in the grain of a crop plant.
It is yet another object of the present invention to provide methods for producing grain for human comsumption with a reduced caloric content.
It is yet another object of the present invention to provide methods for producing grain for human consumption with an increased soluble fiber content.
It is yet another object of the present invention to provide plants with improved stalk or stem quality.
It is yet another object of the present invention to provide plants with altered growth rates to affect flowering.
Therefore, in one aspect, the present invention relates to an isolated nucleic acid comprising a member selected from the group consisting of:
(a) a polynucleotide that encodes a polypeptide of SEQ ID NOS 2 or 4;
(b) a polynucleotide amplified from a plant tissue nucleic acid library using the primers of SEQ ID NOS 5 and 6, or, 7 and 8;
(c) a polynucleotide comprising at least 20 contiguous bases of SEQ ID NOS 1 and 3;
(d) a polynucleotide encoding plant RGP polypeptides;
(e) a polynucleotide having at least 60% sequence identity to SEQ ID NOS 1 or 3:, wherein the % sequence identity is based on the entire coding sequence and is determined by BLAST 2.0 under default parameters;
(f) a polynucleotide comprising at least 25 nucleotides in length which hybridizes under stringent conditions to a polynucleotide having the sequence set forth in SEQ ID NOS 1 and 3, wherein the conditions include a wash with 0.1X SSC at 60° C.;
(g) a polynucleotide comprising the sequence set forth in SEQ ID NOS 1 and 3; and
(h) a polynucleotide complementary to a pol
Dhugga Kanwarpal S.
Fallis Patricia Lynne
Nichols Scott E.
Fox David T.
Ibrahim Medina A
Pioneer Hi-Bred International , Inc.
Pioneer Hi-Bred International Inc.
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