Chemistry: molecular biology and microbiology – Process of mutation – cell fusion – or genetic modification – Introduction of a polynucleotide molecule into or...
Reexamination Certificate
2001-10-26
2004-03-23
Mehta, Ashwin (Department: 1638)
Chemistry: molecular biology and microbiology
Process of mutation, cell fusion, or genetic modification
Introduction of a polynucleotide molecule into or...
C435S320100, C800S282000, C800S298000
Reexamination Certificate
active
06709867
ABSTRACT:
BACKGROUND
Attractive lawns are considered an asset by homeowners and owners of commercial recreational establishments, such as theme parks and golf courses. Typically, such lawns require treatment with a number of chemicals, including fertilizers, pesticides, and herbicides. Unfortunately, until the lawn begins to show obvious symptoms of a nutritional deficiency or infestation with weeds or pests, there is no way to determine when application of such chemicals is appropriate or warranted. Accordingly, there is often extensive and unnecessary usage of such compounds. Excessive treatment with these chemicals is expensive and environmentally compromising.
The maintenance of attractive lawns also requires application of water at appropriate times. If watering is delayed too long, leaf browning and leaf loss can occur. In addition, plants may die from draught stress. Watering too often can also result in plant damage. Moreover, frequent watering is expensive and poses serious problems in areas with water shortage.
Attempts have been made to overcome these problems by developing more resistant grasses that require less fertilizer, herbicides and pesticides. However, there currently are no systems available which permit application of fertilizers, pesticides and water to select areas that need such treatment. Accordingly, it is desirable to have grass plants which are capable of visually communicating their need for fertilizers, chemicals or water to the observer.
SUMMARY OF THE INVENTION
In one aspect, the present invention provides transgenic grass plants which exhibit a color different from the color exhibited by the corresponding non-transgenic grass plants under conditions of stress. Examples of such conditions include, but are not limited to, lack of fertilizer, lack of water, and attack by insects or pathogens. The genome of the present transgenic grass plant comprise a transgene comprising a nucleic acid which encodes one or more regulators of anthocyanin biosynthesis, hereinafter referred to as an “anthocyanin regulatory gene”, and an inducible promoter which is responsive to a stress condition, such as for example, nutrient deprivation, water deprivation, and attack by a pathogen. The promoter, which is hereinafter referred to as a “stress” inducible promoter, is operably linked to the anthocyanin regulatory gene. Expression of the anthocyanin regulatory gene, in response to the stress condition, provides a gene product which activates expression of anthocyanin biosynthetic genes and results in the transgenic grass plant having a different color phenotype.
The present invention also relates to a construct and vector for preparing the transgenic grass. The construct and vector comprise a transgene comprising an anthocyanin regulatory gene operably linked to a stress inducible promoter.
The present invention also relates to a transgenic grass plant which constitutively exhibits a different color phenotype, particularly a red color phenotype. The genome the transgenic grass plant comprises a transgene comprising an exogenous anthocyanin regulatory gene operably linked to a constitutive promoter. Grass plants that constitutively display a colored phenotype are useful for display and decorative purposes.
The present invention also relates to methods of preparing the present transgenic grass plants and the seeds of the present transgenic grass plants.
DETAILED DESCRIPTION OF THE INVENTION
Definitions
Promoter, as used herein, refers to a recognition site on a DNA molecule that provide an expression control element for a gene and which allows the recruitment of RNA polymerase to initiate RNA synthesis (transcription) of the gene.
Regeneration, as used herein, refers to the process of growing a plant from a plant cell (e.g., plant protoplast, plant calli or plant explant).
Transformation, as used herein, refers to a process of introducing an exogenous DNA molecule (e g., a vector, a recombinant DNA molecule) into a cell, a callus, or protoplast in which that exogenous DNA is incorporated into a chromosome.
Transformed Cell, as used herein, refers to a cell whose DNA has been altered by the introduction of an exogenous DNA molecule into that cell.
Transgene, as used herein, refers to an exogenous gene which when introduced into the genome of a host cell through a process such as transformation, electroporation, particle bombardment, and the like, is expressed by the host cell and integrated into the cells genome such that the trait or traits produced by the expression of the transgene is inherited by the progeny of the transformed cell.
Transgenic Cell, as used herein, refers to any cell derived or regenerated from a transformed cell or derived from a transgenic cell. Exemplary transgenic cells include plant calli derived from a transformed plant cell and particular cells such as leaf, root, stem, e.g., somatic cells, or reproductive (germ) cells obtained from a transgenic plant.
Transgenic Plant, as used herein, refers to a plant or progeny thereof derived from a transformed plant cell or protoplast, wherein the plant DNA contains an introduced exogenous DNA molecule not originally present in a native, non-transgenic plant of the same strain.
Vector, as used herein, refers to a DNA molecule capable of replication in a host cell and/or to which another DNA segment can be operatively linked so as to bring about replication of the attached segment. A plasmid is an exemplary vector.
In one aspect, the present invention provides a nucleic acid construct comprising a transgene which comprises one or more anthocyanin regulatory genes and a stress inducible promoter which induces transcription of the anthocyanin regulatory gene in response to a stress condition. The promoter is operably linked to the anthocyanin regulatory gene or genes. The anthocyanin regulatory gene may be derived from a grass plant or from a plant other than grass. In one embodiment the, anthocyanin regulatory genes are derived from maize.
The present invention also provides a transgenic grass plant whose genome comprises a transgene comprising an exogenous anthocyanin regulatory gene and a promoter which is operably linked to the anthocyanin regulatory gene. In one embodiment, the promoter is a stress inducible promoter i.e., a promoter which induces expression of the anthocyanin regulatory gene in response to a stress condition such as for example, drought conditions, lack of fertilizer, or attack by a pathogen. In another embodiment, the promoter is a chemical inducible promoter, which induces expression of the anthocyanin regulatory gene in response to exposure of the plant to the chemical. In yet another embodiment, the promoter is a constitutive promoter which continuously drives expression of the anthocyanin regulatory gene.
Nucleic Acid Construct
A. Anthocyanin Regulatory and Biosynthetic Genes
Anthocyanins are non-toxic pigments that are responsible for many of the red and blue colors in plants. These pigments function to attract pollinating insects to plants and shield plant DNA from ultraviolet light damage. There are multiple anthocyanin genes producing pigments that cause plants to display different colors.
In corn or maize, anthocyanin biosynthesis requires expression of 20 or more genes. Some of these genes are anthocyanin biosynthetic genes; others are anthocyanin regulatory genes. An anthocyanin biosynthetic gene is a gene whose product is an enzyme that is involved in the biosynthesis of the anthocyanins. Examples of anthocyanin biosynthetic genes or loci in maize include C2, Whp, A1, CHI. A2, Bz1, and Bz2. In maize, there are at least 6 anthocyanin regulatory genes or loci, R, B, C1, P1, P and P, whose products are regulatory proteins which activate the transcription of one or more anthocyanin biosynthetic genes.
Although the genetic pathways for anthocyanin synthesis are complex, there is some understanding of regulation of anthocyanin synthesis. Two of the better understood protein factors known to activate expression of anthocyanin biosynthesis genes in maize are C1, which is a homologue of the
Calfee Halter & Griswold LLP
Mehta Ashwin
The Ohio State University Research Foundation
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