Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or...
Reexamination Certificate
2000-03-09
2004-03-09
Bui, Phuong T. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
C800S288000, C800S298000, C800S295000, C800S314000, C435S419000, C435S468000, C435S320100, C536S023200, C536S024100, C536S023700
Reexamination Certificate
active
06703540
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates generally to the protection of plants against phyto-pathogens. In one aspect it provides transgenic plants which have enhanced resistance to phytopathogens. It further provides methods for protecting plants against phytopathogens.
2. Description of the Relevant Art
Living organisms possess several inherent defense mechanisms to fight microbial pathogens. One of the chronologically earliest responses to invasion is a respiratory burst that produces two reactive oxygen species that are microbicidal: superoxide and hydrogen peroxide (Mehdy, M. C. 1994
. Plant Physiol
. 105: 467-472; Wojtaszek, P. 1997
. Biochem. J
. 322: 681-692). Hydrogen peroxide and its products from reactions with endogenous metabolites possess relatively moderate antimicrobial activity (Doke et al. 1991. Active Oxygen/Oxygen Stress and Plant Metabolism, E. Pell et al. (Eds.). Am. Soc. Plant Physiol., Rockville Md.). Deadlier antimicrobial compounds such as hypohalites are generated from hydrogen peroxide in some bacterial organisms that contain haloperoxidases, for example, chloroperoxidases. Animals contain myeloperoxidase; and in the presence of myeloperoxidase, animal cells, too, can convert hydrogen peroxide to a much stronger antimicrobial compound, hypochlorous acid (HOCl), i.e.,
H
2
O
2
+HCl→H
2
O+HOCl (1)
Plants routinely become infected by fungi, viruses, and bacteria; however, vascular plants lack haloperoxidases such as myeloperoxidase and consequently lack this potent system of antimicrobial and antifungal protection. Although plants lack this endogenous haloperoxidase-based method of defense, plant fungi are sensitive to the effects of these haloperoxidases in experiments in vitro. Jacks et al. (1991
. Biochem. Biophys. Res. Commun
. 178: 1202-1204) have shown that the conversion of hydrogen peroxide to hypochlorite, catalyzed by myeloperoxidase, results in as much as a 90-fold increase in lethality against the plant fungus
Aspergillus flavus
in vitro, suggesting the possibility that plants transformed with a haloperoxidase gene could possess the advantage of an increased disease resistance to phytopathogens. However, for a transformation strategy involving the enzymes, myeloperoxidase or most haloperoxidases, to be successful, the addition of a plethora of genes coding for the multi-component synthesis of each species specific heme-containing prosthetic group is a necessary requirement (Pfeifer et al. 1992
. J. Gen. Microbiol
. 138: 1123-1131; van Pee, K-H. 1996
. Annu. Rev. Microbiol
. 50: 375-399) because these heme derivatives are not available in plants.
Accordingly, this invention provides an alternative strategy, specifically, transformation with a gene encoding a functional yet heme-free haloperoxidase. Such a strategy therefore bypasses the need for including nucleic acid sequences which encode additional components related to specific heme groups.
Presently, plant breeding and biocontrol programs, as well as the development of new antimicrobial agents and fungicides, are disease preventive methods that are being successfully practiced to protect crops. However, it is well known that rapid genetic changes take place in pathogenic organisms. Such evolutionary changes result in the resistance genes of the host quickly becoming ineffective. These facts, together with cost, time considerations, and actual field results, necessitate the constant search for novel methods and reagents for protecting agricultural crops.
SUMMARY OF THE INVENTION
We have discovered that the expression of a gene encoding a nonheme haloperoxidase in a plant provides protection to the plant against phytopathogens.
In accordance with this discovery, it is an object of the invention to provide a plant transformed with a gene which encodes a nonheme haloperoxidase and which is expressed to confer protection to the plant against phytopathogens. In particular, a plant in which a nonheme chloroperoxidase is expressed is provided.
Another object of the invention is to provide a method of transforming a plant with a gene capable of expressing a nonheme haloperoxidase sufficient to confer protection to the plant against phytopathogens. A method of transforming a plant with a gene encoding a nonheme chloroperoxidase is particularly provided.
Plant transformation vectors comprising the gene which encodes the nonheme haloperoxidase of the invention are also provided, as are plant cells transformed by these vectors, and plants and their progeny containing the genes.
It is an additional object of the invention to provide a host cell containing the DNA of the invention, wherein said host cell is a bacterial cell, in particular, an
Agrobacterium tumefaciens
cell.
Still further, it is an object of the invention to engineer the DNA molecules of the invention to allow in the plant for constitutive expression, expression in response to an inducer, expression in particular tissues, and expression at particular stages of development.
REFERENCES:
patent: 4956282 (1990-09-01), Goodman et al.
patent: 5773696 (1998-06-01), Liang et al.
Cleveland, T.E., et al., “Identification of Traits for Enhancing Crop Resistance to Preharvest Aflatoxin Contamination”, USDA CRIS Report Project No. 6435-42000-009-00D for Fiscal Year 1995, Published Mar., 1995.
Cleveland, T.E., et al., “Aflatoxin Control Through Addition Enhancement of Antifungal Genes in Corn and Cotton”, USDA CRIS Report Project No. 6435-42000-012-00D for Fiscal Year 1997, Published Mar., 1997.
Jacks, T.J., et al., “Potential of Animal Myeloperoxidase to Protect Plants from Pathogens”,Biochemical and Biophysical Research Communications, vol. 178(3), pp. 1202-1204, Aug. 15, 1991.
Jacks, T.J., et al., “Superoxide, Hydrogen Peroxide, and the Respiratory Burst of Fungally Infected Plant Cells”,Molecular and Cellular Biochemistry, vol. 158, pp. 77-79, 1996.
Jacks, T.J., et al., “Effects of Chloroperoxidase and Hydrogen Peroxide on the Viabilities ofAspergillus flavusConidiospores”,Molecular and Cellular Biochemistry, vol. 195, pp. 169-172, 1999.
Pfeifer, O., et al., “Molecular Cloning and Sequencing of a Non-haem Bromoperoxidase Gene fromStreptomyces aureofaciensATCC 10762”,J. General Microbiology, vol. 138, pp. 1123-1131, 1992.
Picard, M., et al., “Metal-Free Bacterial Haloperoxidases as Unusual Hydrolases: H2O2by the Formation of Peracetic Acid”,Angew. Chem. Int. Ed. Engl., vol. 36(11), pp. 1196-1199, 1997.
Mehdy, M.C., et al., “The Role of Activated Oxygen Species in Plant Disease Resistance”,Physiologia Plantarum, vol. 98, pp. 365-374, 1996.
Wiesner, W., et al., “Purification and Properties of Bromoperoxidase fromPseudomonas pyrrocinia”, J. Biol. Chem., vol. 366, pp. 1085-1091, Dec. 1985.
Wiesner, W., et al., “Purification and Characterization of a Novel Bacterial Non-heme Chloroperoxidase fromPseudomonas pyrrocinia”, J. Biological Chemistry, vol. 263(27), pp. 13725-13732, Sep. 25, 1988.
Wojtaszek, Przemyslaw, “Oxidative Burst: An Early Plant Response to Pathogen Infection”,J. Biochem., vol. 322, pp. 681-692, 1997.
Wolfframm, C., et al., “Chloroperoxidase-encoding Gene fromPseudomonas pyrrocinia: Sequence, Expression in Heterologous Hosts, and Purification of the Enzyme”,Gene, vol. 130, pp. 131-135, 1993.
Wolfframm, C., et al., “Cloning and High-level Expression of a Chloroperoxidase Gene fromPseudomonas pyrrociniainEscherichia coli”, FEBS Letters, vol. 238(2), pp. 325-328, Oct. 1988.
Van Pee, K-H., “Biosynthesis of Halogenated Metabolites by Bacteria”,Annu. Rev. Microbiol., vol. 50, pp. 375-399, 1996.
Cary Jeffrey W.
Cleveland, III Thomas E.
Jacks Thomas J.
Rajasekaran Kanniah
Van Pee Karl-Heinz
Bui Phuong T.
Fado John D.
Ibrahim Medina A.
Rabin Evelyn M.
The United States of America as represented by the Secretary of
LandOfFree
Transformation of plants with a chloroperoxidase gene to... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Transformation of plants with a chloroperoxidase gene to..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Transformation of plants with a chloroperoxidase gene to... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3275002