Multicellular living organisms and unmodified parts thereof and – Plant – seedling – plant seed – or plant part – per se – Higher plant – seedling – plant seed – or plant part
Reexamination Certificate
1998-02-18
2001-12-11
Nelson, Amy J. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Plant, seedling, plant seed, or plant part, per se
Higher plant, seedling, plant seed, or plant part
C435S320100, C536S024100, C800S278000
Reexamination Certificate
active
06329572
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates generally to genetic sequences which are useful in the diagnosis and treatment of fungal infections in plants which involve a susceptible interaction between a fungal pathogen and the host plant In particular, the present invention provides genetic sequences which confer, activate, or enhance expression of a gene in a plant, in response to infection of said plant by a plant fungal pathogen in a susceptible interation. The invention further provides genetic sequences such as structural genes, the expression of which is induced in response to a susceptible interaction between a plant and a fungal pathogen. The present invention further provides methods for the detection of infection by a fungal pathogen in a susceptible interaction and for the production of transgenic plants with improved resistance to said fungal pathogen. The present invention is particularly useful for developing disease resistance in crop varieties.
2. Background of the Information
Advances in plant biotechnology have dramatically altered the approaches taken to increase the economic output of productive units of agriculture. Of major significance to the agricultural and horticultural industries are the reduced productivity, due to infection by plant pathogens. Plant fungal pathogens, in particular rust fungi, represent an especially significant problem amongst broadacre crops such as legume and cereal grains. Biotechnology offers considerable scope for addressing this problem, by introducing recombinant genes into plant that either kill or disable a fungal pathogen, or restrict a fungal pathogen to a limited zone of infection, thereby preventing significant deterioration of an economically-important crop. Thus, the development of disease resistant plants by biotechnological means, is an important goal in agricultural and horticultural research.
Genetic analyses indicate that rust resistance genes of the plant genome control specific recognition of the products of rust avirulence genes. An interaction between a rust pathogen and a plant host may be classed as either “resistant” or “susceptible” depending on how the fungal infection proceeds. In a resistant interaction, infection by a fungal pathogen produces a “plant hypersensitive response” (Marineau et al., 1987; Dixon and Lamb, 1990) resulting in cell death to limit spread of the fungus. During the hypersensitive response, the expression of several infection-related genes, for example genes encoding phytoalexins, antimicrobial agents and pathogenesis-related (PR) proteins, is switched on. In contrast, a susceptible interaction involves no hypersensitive cell death and the infection alters host cell gene expression in such a way as to provide gene products that are essential for the biotrophic growth of an obligate plant pathogen. Thus, the two processes are quite distinct, involving different host cell genes and mechanisms regulating the expression of said host cell genes. This distinction is of paramount importance. For example, those host genes induced in a susceptible interaction may be essential to allow the rust to grow in the plant tissues.
Most studies have concentrated on identifying and manipulating genes encoding proteins involved in the hypersensitive response of the resistant interaction (Collinge and Slusarenko, 1987; Dixon and Lamb, 1990; Keen, 1992; van Loon, 1985; Ohashi and Ohshima, 1992). Marlini and Strittmatter (Patent Application WO 9319188) have constructed a fungus-responsive chimaeric gene, using a promoter sequence from the prp1 gene, in particular the prp1-1 gene, to direct expression of a “killer” gene in plant cells infected by a fungal pathogen. However the prp1 genetic sequence is induced in a resistant interaction only (i.e. in a pathogenesis-related fungal infection). Although genetic sequences such as the prp1 gene may provide a means of control of a pathogen in a resistant interaction, the isolation of host cell genetic sequences involved in a susceptible interaction between a plant and a fungal pathogen has not been a straightforward procedure.
In full-susceptible interactions between the flax plant
Linum usitatissimum
and the flax rust
Melampsora lini
, there is no immediate host cell death or chlorosis. Instead, host cell metabolism is directed toward the production of viable fungal spores, including for example, the translocation of photosynthates via the haustorium or lungal absorptive organ to the fungal mycelium. Although altered patterns of protein synthesis have been observed following a susceptible rust infection of flax plants (Sutton and Shaw, 1986), it has not been possible, until the present invention, to differentiate between fungal protein synthesis and modifications to plant protein synthesis. Thus, the isolation of flax genetic sequences, the expression of which is induced during a susceptible rust infection, has not been a straightforward procedure.
Bibliographic details of the publications referred to by author in this specification including information disclosed under 37 C.F.R. §1.97 and 1.98, are collected at the end of the detailed description of the invention.
SUMMARY OF THE INVENTION
In work leading up to the present invention, the inventors sought to develop plants with improved resistance to fungal pathogens which would otherwise infect the plant in a susceptible interaction. Accordingly, the inventors identified plant genetic sequences in flax and maize, the expression of which increases in response to a susceptible rust infection. The cloning of these sequences provides a means of generating transgenic plants with de novo, improved, or otherwise enhanced antifungal properties. In particular, by placing a cytotoxic gene, anti-fungal gene, antisense, ribozyme or co-supression molecule operably under control of a promoter sequence derived from a genetic sequence which is normally tanscriptionally up-regulated in response to a susceptible infection, and introducing the resulting chimeric gene into a plant, disease resistance against said fungal pathogen is conferred or otherwise facilitated in said plant The present invention also permits the screening, through genetic or immunological means, similar susceptible reaction-responsive (SRR) genetic sequences in other plants, for use in developing or enhancing the antigal properties of commercially- and economically-important species.
REFERENCES:
patent: WO91/15585 (1991-10-01), None
patent: WO 93/19188 (1993-09-01), None
patent: WO95/03690 (1995-02-01), None
patent: WO95/33818 (1995-12-01), None
Stam M, et al. “The silence of genes in transgenic plants.” Ann. Bot. 79: 3-12, 1997.*
Koziel MG, et al. “Optimizing expression of transgenes with an emphasis on post-transcriptional events.” Plant Mol. Biol. 32: 393-405, 1996.*
Smith CJS, et al. “Antisense RNA inhibition of polygalacturonase gene expression in transgenic tomatoes.” Nature 334: 724-726, Aug. 25, 1988.*
Benfey PN, et al. “The cauliflower mosaic virus 35S promoter: Combinatorial regulation of transcription in plants.” Science 250: 959-966, Nov. 1990.*
Kim Y, et al. “A 20 nucleotide element is essential for the nopaline synthase (nos) promoter activity.” Plant Mol. Biol. 24: 105-117, 1994.*
J. Cell. Biochem. 19B: 153. Keystone Symposium on Host-Fungus Pathogenic Interactions, Feb. 26, 1995.*
Castresana et al., Tissue-Specific and Pathogen-Induced Regulation of aNicotiana plumbaginifolia&bgr;-1,3-Glucanase Gene, Plant Cell, vol. 2, No. 12, pp. 1131-1143, Dec. 1990, XP-002146377.
Gough et al., “Developmental and pathogen-induced activation of an msr gene, str. 246C, from tobacco involves multiple regulatory elements”, Molecular and General Genetics (1995) 247 (3)323-327, XP-002146378.
Samac et al., “Developmental and Pathogen-Induced Activation of the Arabidopsis Acidic Chitinase Promoter”, Plant Cell, vol. 3, No. 10, 1063-1072, Oct. 1991, XP-002146376.
Roberts et al., Isolation of a flax (Linum usitatissimum) gene induced during susceptible infection by flax rust (Melampsora lini), Plant Journal (Jul. 1995)
Pryor Anthony J.
Roberts James K.
Bacon & Thomas
Commonwealth Scientific and Industrial Research Organisation
Nelson Amy J.
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