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
1999-02-25
2002-04-23
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
C800S278000, C800S286000, C800S287000, C800S320100, C435S101000, C435S468000
Reexamination Certificate
active
06376749
ABSTRACT:
BACKGROUND OF THE INVENTION
Characteristics and Commercial Utility of Starch
Starch is a mixture of two polysaccharides, amylose and amylopectin. Amylose is an unbranched chain of up to several thousand &agr;-D-glucopyranose units linked by &agr;-1,4 glycosidic bonds. Amylopectin is a highly branched molecule made up of up to 50,000 &agr;-D-glucopyranose residues linked by &agr;-1,4 and &agr;-1,6 glycosidic bonds. Approximately 5% of the glycosidic linkages in amylopectin are &agr;-1,6 bonds, which leads to the branched structure of the polymer.
Amylose and amylopectin molecules are organized into granules that are stored in plastids. The starch granules produced by most plants are 15-30% amylose and 70-85% amylopectin. The ratio of amylose to amylopectin and the degree of branching of amylopectin affects the physical and functional properties of the starch. Functional properties, such as viscosity and stability of a gelatinized starch determine the usefulness and hence the value of starches in food and industrial applications. Where a specific functional property is needed, starches obtained from various crops such as corn, rice, or potatoes may meet the functionality requirements. If a starch does not meet a required functional property, if for example it must have stable viscosity under high temperatures and acidic conditions, the functionality can sometimes be achieved by chemically modifying the starch. Various types and degrees of chemical modification are used in the starch industry, and the labeling and use of chemically modified starches must meet government regulations.
Within the starch bearing organs of plants, the proportion of amylose to amylopectin and the degree of branching of amylopectin are under genetic control. For example, plants homozygous recessive for the waxy (wx) gene lack a granule-bound starch synthase enzyme and produce nearly 100% amylopectin. Plants homozygous recessive for the amylose extender (ae) gene can produce starch granules that are up to 90% amylose (see U.S. Pat. No. 5,300,145). The dull gene has been shown to influence the levels of activity of a starch synthase and a starch branching enzyme.
Most cereal crops are handled as commodities, and many of the industrial and animal feed requirements for these crops can be met by common varieties which are widely grown and produced in volume. However, there exists at present a growing market for crops with special end-use properties which are not met by grain of standard composition. Most commonly, specialty corn is differentiated from “normal” corn, also known as field corn, by altered endosperm properties, such as an overall change in the ratio of amylose to amylopectin as in waxy or high amylose corn, an increased accumulation of sugars as in sweet corn, or an alteration in the degree of endosperm hardness as in food grade corn or popcorn; Glover, D. V. and E. T. Mertz, (1987) in Corn: Nutritional Quality of Cereal Grains; Genetic and Agronomic Improvement, R. A. Olson and K. J. Frey, eds. American Society of Agronomy, Madison Wisconsin, pp. 183-336. Rooney, L. W. and S. O. Serna-Saldivar, (1987) Food Uses of Whole Corn and Dry-milled Fractions, in Corn: Chemistry and Technology, S. A. Watson and P. E. Ramstead, eds. American Association of Cereal Chemists, Inc., St. Paul, Minn., pp. 399-429. The current invention offers the buyers of specialty corn a source of starch having properties distinct from waxy starch and offers farmers the opportunity to grow a higher value-added crop than normal or waxy corn.
Purified starch is obtained from plants by a milling process. Corn starch is extracted from kernels through the use of a wet milling process. Wet milling is a multi-step process involving steeping and grinding of the kernels and separation of the starch, protein, oil and fiber fractions. A review of the corn wet milling process is given by S. R. Eckhoff in the Proceedings of the Fourth Corn Utilization Conference, Jun. 24-26, 1992, St. Louis, Mo., printed by the National Corn Growers Association, CIBA-GEIGY Seed Division and the United States Department of Agriculture. Starch is used in numerous food and industrial applications and is the major source of carbohydrates in the human diet. Typically, starch is mixed with water and cooked to form a thickened gel. Three important properties of a starch are the temperature at which it cooks, the viscosity the gel reaches, and the stability of the gel viscosity over time. The physical properties of unmodified starch during heating and cooling limit its usefulness in many applications. As a result, considerable effort and cost is needed to chemically modify starch in order to overcome these limitations of starch and to expand the usefulness of starch in industrial applications.
Some limitations of unmodified starches and properties of modified starches are given in Modified Starches: Properties and Uses, O. B. Wurzburg, ed., (1986) CRC Press Inc., Boca Raton, Fla. Unmodified starches have very limited use in food products because the granules swell and rupture easily, thus forming weak bodied, undesirable gels. Chemical modifications are used to stabilize starch granules thereby making the starch suitable for thousands of food and industrial applications including baby foods, powdered coffee creamer, surgical dusting powders, paper and yarn sizings and adhesives. Common chemical modifications include cross linking in which chemical bonds are introduced to act as stabilizing bridges between starch molecules, and substitution in which substituent groups such as hydroxyethyl, hydroxypropyl or acetyl groups are introduced into starch molecules.
The use of chemically modified starches in the United States is regulated by the Food and Drug Administration (FDA). “Food starch-modified” starches may be used in food but must meet specified treatment limits, and “industrial starch-modified” starches may be used in items such as containers that come in contact with food and must also meet specified treatment requirements; Code of Federal Regulations, Title 21, Chapter 1, Part 172, Food Additives Permitted in Food for Human Consumption, Section 172, 892, Food Starch-Modified, U. S. Government Printing Office, Washington, D. C. 1981; (a) Part 178, Indirect Food Additives, Sect. 178.3520, Industrial Starch-Modified. These regulations limit the degree of chemical modification by defining the maximum amount of chemical reagent that can be used in the modification steps. The levels of by-products in starch resulting from the modification process are also regulated. For example, propylene chlorohydrin residues in hydroxypropyl starch are of special concern; Tuschhoff, J. V., (1986) Hydroxypropylated Starches, In Modified Starches: Properties and Uses, O. B. Wurzburg, ed., CRC Press, Boca Raton, Fla. pp. 89-96.
Alteration of Starch Fine Structure Through Molecular Genetic Manipulation of Starch-Bearing Plants
Differences in the degree of starch branching or polymerization are known to result in a change in the physiochemical properties of starch. It has been suggested that starches, tailor-made for specific applications, may be generated by alteration of the branch chain distribution of the amylopectin molecule, the relative proportion of amylose to amylopectin or the degree of polymerization of amylose. However, achieving phenotypic alteration of starch composition has been problematic; while key enzymes in starch biosynthesis have been identified, their exact roles remain uncertain. Thus, correlation of activities of particular enzymes with particular molecular characteristics of starch structure and, in turn, with starch function in food and industrial products has been difficult. Although desirable functional properties that an ideal starch might need can be envisioned, there is only a vague understanding of what the molecular structure of the starch should be to achieve this and little understanding of how particular starch biosynthetic enzymes specifically affect those parameters. For example, the role of individual enzymes in determining the branching pattern
Broglie Karen E.
Hubbard Natalie L.
Klein Theodore M.
Lightner Jonathan E.
E. I. du Pont de Nemours and Company
Fox David T.
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