MXPA99006042A - Method for protecting plants - Google Patents

Abstract

The present invention concerns a method of protecting plants from pathogen attack through synergistic disease resistance attained by applying a conventional microbicide to immunomodulated plants. Immunomodulated plants are those in which SAR is activated and are therefore referred to as"SAR-on"plants. Immunomodulated plants may be provided in at least three different ways:by applying to plants a chemical inducer of SAR such as BTH, INA, or SA;through a selective breeding program based on constitutive expression of SAR genes and/or a disease-resistant phenotype;or by transforming plants with one or more SAR genes such as a functional form of the N/M1 gene. By concurrently applying a microbicide to an immunomodulated plant, disease resistance is unexpectedly synergistically enhanced;i.e., the level of disease resistance is greater than the expected additive levels of disease resistance.

Description

METHOD FOR THE PROTECTION OF PLANTS The present invention relates to a method for protecting a plant against the attack of pathogens, through synergistic resistance to diseases, obtained by applying a microbicide to an immunomodulated plant. Plants are constantly assaulted by a wide variety of pathogenic organisms, including viruses, bacteria, fungi, and nematodes. The crop plants are particularly vulnerable, because they are normally grown as genetically uniform monocultures; When the disease strikes, the losses can be severe. However, most plants have their own innate mechanisms of defense against pathogenic organisms. Plant breeders and pathologists have identified a natural variation for resistance to plant pathogens, and reproduce it in many crop plants. These natural disease resistance genes often provide high levels of resistance to, or immunity against, pathogens. Acquired systemic resistance (SAR) is a component of the complex system that plants use to defend against pathogens (Hunt and Ryals, Cri t Rev. in Plant Sci. 15, 583-606 (1996), incorporated herein by reference). reference in its entirety, Ryals et al., Plant Cell 8, 1809-1819 (1996), incorporated herein by reference in its entirety). See also, U.S. Patent Number 5,614,395, incorporated herein by reference in its entirety. Acquired systemic resistance is a particularly important aspect of plant-pathogen responses, because it is a systemic resistance inducible by the pathogen against a broad spectrum of infectious agents, including viruses, bacteria, and fungi. When the signal transduction pathway of acquired systemic resistance is blocked, plants become more susceptible to the pathogens that normally cause disease, and they also become susceptible to some infectious agents that would not normally cause disease (Gaffney et al., Science 261, 754-756 (1993), incorporated herein by reference in its entirety, Delaney et al., Science 266, 1247-1250 (1994) incorporated herein by reference in its entirety; Delaney et al., Proc. Nati Acad. Sci USA 92, 6602-6606 (1995), incorporated herein by reference in its entirety; Delaney, Plant Phys. 113, 5-12 (1997), incorporated herein by reference in its entirety; Bi et al., Plant. J. 8, 235-245 (1995), incorporated herein by reference in its entirety; Mauch-Mani and Slusarenko, Plant Cell 8, 235-245 (1995), incorporated herein by reference in its entirety). These observations indicate that the signal transduction pathway of acquired systemic resistance is critical to maintain the health of the plant.

Conceptually, the acquired systemic response can be divided into two phases. In the initiation phase, an infection of the pathogen is recognized, and a signal that travels through the phloem to distant tissues is released. This systemic signal is perceived by white cells, which react by expressing both genes of acquired systemic resistance and resistance to diseases. The phase of maintenance of acquired systemic resistance refers to the period of time, from weeks to the whole life of the plant, during which the plant is in an almost passive state, and resistance to diseases is maintained (Ryals et al. , nineteen ninety six) . It seems that the accumulation of salicylic acid (SA) is required for the transduction of signals of acquired systemic resistance. Plants that can not accumulate salicylic acid due to treatment with specific inhibitors, epigenetic repression of phenylalanine ammonia-lyase, or transgenic expression of salicylate hydroxylase, which specifically degrades salicylic acid, can not induce the expression of the acquired systemic resistance gene. nor resistance to diseases (Gaffney et al., 1993; Delaney et al., 1994; Mauch-Mani and Slusarenko 1996, Maher et al., Proc. Nati Acad. Sci. USA 91, 7802-7806 (1994), incorporated herein by reference in its entirety; Pallas and collaborators, Plant J. 10, 281-293 (1996), incorporated herein by reference in its entirety). Although it has been suggested that salicylic acid could serve as the systemic signal, this is currently in controversy, and to date, all that is known for sure is that if salicylic acid can not accumulate, then transduction is blocked. the signal of acquired systemic resistance (Pallas et al., 1996; Shulaev et al., Plant Cell 1, 1691-1701 (1995), incorporated herein by reference in its entirety; Vernooij et al., Plant cell 6, 959-965 ( 1994), incorporated herein by reference in its entirety). Recently, Arabidopsis has emerged as a model system for studying acquired systemic resistance (Uknes et al., Plant Cell 4, 645-656 (1992), incorporated herein by reference in its entirety; Uknes et al., Mol. Plant-Microbe Interact 6, 692-698 (1993), incorporated herein by reference in its entirety, Cameron et al., Plant J. 5, 715-725 (1994), incorporated herein by reference in its entirety; Mauch-Mani and Slusaren-ko, Mol.Plant-Microbe Interact., 1, 378-383 (1994), incorporated herein by reference in its entirety, Dempsey and Kles-sing, Bulletin de L 'Insti tuí Pasteur 93, 167 -186 (1995), incorporated herein by reference in its entirety). It has been shown that systemic resistance acquired in Arabidopsis can be activated by pathogens and chemicals, such as salicylic acid, 2,6-dichloroisonicotinic acid (INA), and S-methyl ester of benzoic acid [1, 2, 3] thiadiazole-7-carbothioic acid (BTH) (Uknes et al., 1992; Vernooij et al., Mol. Plant-Microbe Interact., 8, 228-234 (1995), incorporated herein by reference in its entirety; Lawton et al., Plant J. 10, 71-82 (1996), incorporated herein by reference in its entirety). Following treatment with 2,6-dichloroisonicotinic acid or benzo [1, 2, 3] thiadiazole-7-carbothioic S-methyl ester, or pathogenic infection, at least 3 protein genes related to pathogenesis are induced coordinately ( PR), that is, PR-1, PR-2, and PR-5, in a manner concomitant with the establishment of resistance (Uknes et al., 1992, 1993). In tobacco, the best characterized species, treatment with a pathogen or with an immunization compound, induces the expression of at least nine sets of genes (Ward et al., Plant Cell 3, 1085-1094 (1991), incorporated into the present as a reference in its entirety). Disease-resistant transgenic plants have been created by transforming plants with different acquired systemic resistance genes (U.S. Patent Number 5,614,395). A number of Arabidopsis mutants have been isolated that have modified the transduction of the acquired systemic resistance signal (Delaney, 1997). The first of these mutants are the so-called Isd mutants (lesions simulating disease), and acd2 (accelerated cell death, accelerated cell death) (Dietrich et al., Cell 11, 565-577 (1994) incorporated herein by reference in its entirety, Greenberg et al., Cell 11, 551-563 (1994), incorporated herein by reference in its entirety). All these mutants have some degree of spontaneous necrotic lesion formation in their leaves, high levels of salicylic acid, mRNA accumulation for acquired systemic resistance genes, and significantly improved disease resistance. At least seven different Isd mutants have been isolated and characterized (Dietrich et al., 1994; Weymann et al., Plant Cell 1, 2013-2022 (1995), incorporated herein by reference in its entirety). Another interesting class of mutants are the cim mutants (cpnstitutive immunity, constitutive immunity) (Lawton et al., "The molecular biology of systematic aquired resistance" in Mechanisms of Defense Responses in Plants, B. Fritig and M. Legrand, editors (Dordrecht, The Netherlands: Kluwer Academic Publishers), pages 422-432 (1993), incorporated herein by reference in its entirety). See also, International Application of TCP Number WO 94/16077, which is incorporated herein by reference in its entirety. Like the Isd and acd2 mutants, the cim mutants have an expression of the acquired systemic resistance and high salicylic acid resistance gene, but in contrast to Jsd or acd2, they do not exhibit detectable lesions on their leaves. cprl (for its acronym in English constitutive expresser of EE genes, constitutive expression of genes for resistance to pathogens) could be a type of cim mutant; however, because the presence of microscopic lesions in the cprl leaves has not been ruled out, the cprl could be a type of Isd mutant (Bowling et al., Plant Cell 6, 1845-1857 (1994), incorporated herein). as a reference in its entirety). Mutants have also been isolated that are bld in the signaling of acquired systemic resistance. ndrl (non-race specific disease resistance) is a mutant that allows the growth of both Pseudomonas syringae containing different avirulence genes, as well as the normally avirulent isolates of Peronospora parasite (Century et al, Proc. Nati, Acad. Sci. USA 92, 6597-6601 (1995, incorporated herein by reference in its entirety.) Apparently, this mutant is bld early in the signaling of acquired systemic resistance nprl (nonexpresser of PE genes, non-expressor of pathogen resistance genes) is a mutant that can not induce the expression of the signaling pathway of acquired systemic resistance following the INA treatment (Cao). and collaborators, Plant cell 6, 1583-1592 (1994), incorporated herein by reference in its entirety.) Eds mutants have been isolated (by their acronyms in English, snhanced disease = improved susceptibility to diseases), based on its ability to withstand bacterial infection following the inoculation of a low bacterial concentration (Glaze-brook et al., Genetics 143, 973-982 ( 1996), incorporated herein by reference in its entirety; Parker, Plant Cell 8, 2033-2046 (1996), incorporated herein by reference in its entirety). Certain eds mutants are phenotypically very similar to nprl, and recently, eds5 and eds53 have been shown to be allelic for nprl (Glazebrook et al., 1996). niml (non-inducible immunity, non-inducible immunity) is a mutant that supports the growth of P. parasí tica (ie causal agent of the plush mold disease) following treatment with INA (Delaney et al., 1995; International Publication Number WO 94/16077). Although the child can accumulate salicylic acid following the infection of the pathogen, it can not induce the expression of the acquired systemic resistance gene or resistance to diseases, suggesting that the mutation bl the downstream path of salicylic acid. The niml is also impaired in its ability to respond to INA or BTH, suggesting that the blge exists downstream of the action of these chemicals (Delaney and collaborators, 1995, Lawton et al., 1996). Recently, two allelic genes of Arabidopsis have been isolated and characterized, mutants that are responsible for the niml and nprl phenotypes respectively (Ryals et al., Plant Cell 9 425-439 (1997), incorporated herein by reference in its entirety).; Cao et al., Cell 88, 57-63 (1997), incorporated herein by reference in its entirety). The product of the wild type NT 1 gene is involved in the signal transduction cascade that leads to both acquired systemic resistance and resistance to gene-by-gene diseases in Arabidopsis (Ryals et al., 1997). Ryals et al., 1997, also report the isolation of 5 additional alleles of niml, which show a range of phenotypes, from weakly impaired in the expression of chemically induced PR-1 gene, and resistance to fungi to a very strong blockade. The transformation of the wild-type NPR1 gene into nprl mutants not only complemented the mutations, restoring the responsibity of the induction of acquired systemic resistance with respect to expression of the pathogen resistance gene and resistance to diseases, but also made that the transgenic plants were more resistant to infection by P. syringae in the absence of the induction of acquired systemic resistance (Cao et al., 1997). Signal transduction pathways? F-? B / l? B have been implicated in disease resistance responses in a range of organisms from Drosophila to mammals. In mammals, the transduction of NF-? B / l? B signals can be induced by a number of different stimuli, including exposure of the cells to lipopolysaccharide, tumor necrosis factor, interleukin-1 (IL-1), or infection of viruses (Baeuerle and Baltimo-re, Cell 87, 13-20 (1996); Baldwin, Annu, Rev. Immunol., 14, 649-681 (1996)). The activated pathway leads to the synthesis of a number of factors involved in inflammation and in immune responses, such as IL-2, IL-6, IL-8, and granulocyte / macrophage colony stimulating factor (deMartin et al., Gene 152, 253-255 (1995)). In studies of transgenic mice, the elimination of NF-? B / l? B signal transduction leads to a defective immune response, including an increased susceptibility to bacterial and viral pathogens (Beg and Baltimore, Science 274, 782-784 (1996), Van Antwerp et al, Science 274, 787-789 (1996), Wang et al, Science 274 784-787 (1996), Baeuerle and Baltimore (1996)). In Arabidopsis, acquired systemic resistance is functionally analogous to inflammation, in that normal resistance processes are enhanced following the activation of acquired systemic resistance, leading to better resistance to diseases (Bi et al., 1995); Cao et al., 1994; Delaney et al., 1995; Delaney et al., 1994; Gaffney et al., 1993; Mauch-Mani and Slusarenko 1996; Delaney, 1997). In addition, inactivation of the route leads to greater susceptibility to bacterial, viral, and fungal pathogens. Interestingly, it has been reported that salicylic acid blocks the activation of NF-? B in mammalian cells (Kopp and Ghosh, Science 265, 956-959 (1994)), whereas salicylic acid activates signal transduction in Arabidopsis . The bacterial infection of Drosophila activates a cascade of signal transduction that leads to the synthesis of a number of antifungal proteins, such as cercropin B, defensin, diptericin, and drosomycin (Ip et al., Cell 75, 753-763 (1993); Lemaitre et al., Cell 86, 973-983 (1996)). This induction depends on the genetic product of dorsal and dif, two homologs of NF-KB, and is represented by cactus, a homologue of I? B in the fly. Mutants that have diminished the synthesis of antifungal and antibacterial proteins have dramatically lowered their resistance to infections. Despite much research and the use of sophisticated and intense measures of crop protection, including genetic transformation of plants, there are still losses due to the disease, in billions of dollars annually. ThereforeThere is a continuing need to develop new crop protection measures based on the ever-growing understanding of the genetic basis for resistance to plant diseases. In view of the foregoing, a preferred aspect of the present invention pertains to a novel method for protecting plants from pathogen attack, through synergistic resistance to diseases obtained by the application of a microbicide to immunomodulated plants. Immunomodulated plants are those in which acquired systemic resistance is activated, which normally exhibit an expression of the acquired systemic resistance gene greater than the wild type, and are therefore referred to as "acquired-activated systemic resistance" plants. The immunomodulated plants for use in the method of the invention can be obtained in at least three different ways: by applying to the plants a chemical inductor of acquired systemic resistance, such as BTH, INA, or SA; through a selective breeding program where the plants are selected based on the constitutive expression of the acquired systemic resistance genes and / or a disease-resistant phenotype; or by the genetic design of plants by their transformation with one or more genes of acquired systemic resistance, such as a functional form of the NIM1 gene. The microbicide applied to immunomodulated plants can be a conventional microbicide, such as the metalaxyl fungicide, or, if applied to immunomodulated plants obtained through selective breeding or genetic engineering, the microbicide can be a chemical inducer of acquired systemic resistance, such as BTH, INA, or SA. Immunomodulation provides some level of resistance to diseases in a plant. Similarly, the application of a microbicide to a plant provides a certain level of resistance to diseases. The expected result of combining the immuno modulation with the application of the microbicide would be a level of control that reflects the additive levels of control provided by the individual methods to provide resistance to the diseases. However, by concurrent application of a microbicide to an immunomodulatory plant, disease resistance is unexpectedly improved synergistically, that is, the level of disease resistance is greater than the expected additive levels of disease resistance. According to the above, the present invention relates to the cultivation of immunomodulated plants, and to the application of a suitable amount of a conventional microbicide thereto. Especially preferred embodiments of the invention relate to plants genetically engineered to contain and express a functional form of the NIM1 gene, or a homologue or variant thereof. The method of the invention results in greater pathogen control than is achieved through immunomodulation or microbicide application alone. Immunomodulation provides some level of resistance to diseases in a plant. In a similar way, the application of a microbicide to a plant provides a certain level of resistance to diseases. The expected result of combining immunomodulation with the application of the microbicide would be a level of control that reflects the additive levels of control provided by the individual methods to provide resistance to diseases. However, by concurrent application of a microbicide to an immunomodulated plant, control of the pathogenic disease unexpectedly improves synergistically; that is, the level of control of the disease is greater than the expected additive levels of disease resistance. In addition to the greater resistance to diseases, another advantage of the present invention is that less microbicide is required to reach the level of disease resistance provided by the method of the invention, than is required for -Used with ordinary plants. of wild type. The result of this is, both lower economic costs of the microbicide, and less opportunity for adverse consequences for the environment resulting from the toxicity of some microbicides. In addition, the method of the invention for protecting plants by combining the effects of immuno-modulation and the application of a microbicide, results in a longer duration of antipathogenic action, and higher crop yields in total. Another advantage of this method is that, because the two combined modes of action of pathogen control are completely different from each other, the threat of developing resistance is effectively prevented. Accordingly, the present invention relates to a method for protecting a plant from pathogen attack, through synergistic resistance to diseases, which comprises the steps of: (a) providing an immunomodulated plant having a first level of resistance to diseases; and (b) applying to this immunomodulated plant at least one microbicide that confers a second level of resistance to diseases; (c) whereby, the application of the microbicide to the immunomodulated plant confers a third, synergistically improved level of resistance to diseases that is greater than the sum of the first and second levels of disease resistance. A method according to the invention is preferred, wherein the immunomodulated plant is a mutant plant of constitutive immunity (cim). In particular, a method according to the invention is preferred, wherein the cim mutant plant is selected from a population of plants according to the following steps: (a) evaluating the expression of systemic resistance genes acquired in plants not infected animals that are phenotypically normal, where the non-infected plants lack an injury imitation phenotype; and (b) selecting uninfected plants that constitutively express acquired systemic resistance genes in the absence of a viral, bacterial, or fungal infection. Also preferred is a method according to the invention, wherein the immunomodulated plant is a mutant plant that mimics injury. In particular, a method according to the invention is preferred, wherein the mutant plant that mimics injury is selected from a population of plants according to the following steps: (a) evaluating the expression of acquired systemic resistance genes in uninfected plants that have a phenotype that mimics injury; and (b) selecting uninfected plants that constitutively express acquired systemic resistance genes in the absence of a viral, bacterial, or fungal infection. Also preferred is a method according to the invention, wherein the immunomodulated plant is obtained by recombinant expression in a plant of an acquired systemic resistance gene. In particular, a method according to the invention is preferred, wherein the acquired systemic resistance gene is a functional form of a NIM1 gene. More preferred is a method according to the invention, wherein the NIM1 gene encodes a NIM1 protein involved in the signal transduction cascade that leads to the systemic resistance acquired in plants.

Especially preferred is a method according to the invention, wherein the NIM1 protein comprises the amino acid sequence stipulated in SEQ ID N0.-2. Especially preferred is a method according to the invention, wherein the NIM1 gene is hybridized under the following conditions, up to the coding sequence stipulated in SEQ ID NO: 1: hybridization in 1 percent bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in 6X SSC for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. Especially preferred is a method according to the invention, wherein the NIM1 gene comprises the coding sequence stipulated in SEQ ID NO: 1, and all the DNA molecules that hybridize thereto, use moderate astringent conditions. In particular, a method according to the invention is preferred, wherein the acquired systemic resistance gene encodes an altered form of a NIM1 protein that acts as a negative-dominant regulator of the signal transduction pathway of acquired systemic resistance. More preferred is a method according to the invention, wherein the altered form of the NIM1 protein has alanines in place of serines at amino acid positions corresponding to positions 55 and 59 of SEQ ID NO: 2.

Especially preferred is a method according to the invention, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 8. A method according to the invention is particularly preferred, wherein the molecule of DNA comprises the nucleotide sequence shown in SEQ ID NO: 7, and all DNA molecules that hybridize thereto use moderate astringent conditions. Especially preferred is a method according to the invention, wherein the DNA molecule is hybridized under the following conditions, up to the nucleotide sequence stipulated in SEQ ID NO: 7: hybridization in 1 percent bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt, - 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. More preferred is a method according to the invention, wherein the altered form of the NIM1 protein has an N-terminal truncation of amino acids corresponding approximately to amino acid positions 1-125 of SEQ ID NO: 2. Especially preferred is a method according to the invention, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID.

NO: 10. A method according to the invention is particularly preferred, wherein the DNA molecule comprises the sequence of. nucleotides shown in SEQ ID NO: 9, and all DNA molecules that hybridize thereto use moderate astringent conditions. Especially preferred is a method according to the invention, wherein the DNA molecule is hybridized under the following conditions to the nucleotide sequence stipulated in SEQ ID NO: 9: hybridization in 1 percent bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. Especially preferred is a method according to the invention, wherein the altered form of the NIM1 protein has a C-terminal truncation of amino acids corresponding approximately to amino acid positions 522-593 of SEQ ID NO: 2. Especially preferred is a method according to the invention, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 12. A method according to the invention is especially preferred., wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 11, and all DNA molecules that hybridize thereto use moderate astringent conditions. Especially preferred is a method according to the invention, wherein the DNA molecule is hybridized under the following conditions to the nucleotide sequence stipulated in SEQ ID NO: 11: hybridization in 1% bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. More preferred is a method according to the invention, wherein the altered form of the NIM1 protein has an N-terminal truncation of amino acids corresponding to approximately amino acid positions 1-125 of SEQ ID NO: 2, and a truncation C-terminal amino acids corresponding to approximately amino acid positions 522-593 of SEQ ID NO: 2. A method according to the invention is particularly preferred, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 14 A method in accordance with the invention is particularly preferred, wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 13, and all DNA molecules that hybridize with the same using moderate astringent conditions. Especially preferred is a method according to the invention, wherein the DNA molecule is hybridized under the following conditions to the nucleotide sequence stipulated in SEQ ID NO: 13: hybridization in 1 percent bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. More preferred is a method according to the invention, wherein the altered form of the NIM1 protein consists essentially of ankyrin motifs corresponding to approximately amino acid positions 103-362 of SEQ ID NO: 2. Especially preferred is a method according to the invention, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 16. A method according to the invention is especially preferred, wherein the DNA comprises the nucleotide sequence shown in SEQ ID NO: 15, and all the molecules of DNA that hybridize with it using moderate astringent conditions. Especially preferred is a method according to the invention, wherein the DNA molecule is hybridized under the following conditions to the nucleotide sequence stipulated in SEQ ID NO: 15: hybridization in 1 percent bovine serum albumin; NaP04 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; 1 mM EDTA; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (three times) SSC3X for 15 minutes (once) at 55 ° C. Examples of target crops for the indication areas disclosed herein include, without limitation, the following plant species: cereals (corn, wheat, barley, rye, oats, rice, sorghum, and related crops), * beet (sugar beet and forage beet); grapefruit, hard fruit and soft fruit (apples, pears, plums, peaches, almonds, cherries, strawberries, raspberries, and blackberries); leguminous plants (beans, lentils, peas, soybeans), * oil plants (rapeseed, mustard, poppy, olives, sunflowers, coconut, castor oil plants, cocoa seeds, peanuts); cucumber plants (courgettes, cucumber, melons), * fiber plants (cotton, flax, hemp, jute); Citric fruit (oranges, lemons, grapefruit, tangerines); vegetables (spinach, lettuce, asparagus, cabbages, carrots, onions, tomatoes, potatoes, paprika); lauraceae (avocados, cinnamon, camphor); or plants such as tobacco, nuts, coffee, sugarcane, tea, vines, hops, bananas, and natural rubber plants, as well as ornamental plants (flowers, shrubs, broadleaved trees and evergreen trees, such as conifers). This list does not represent a limitation. The method of the present invention can be used to confer resistance to a broad set of plant pathogens, including, but not limited to, the following: viruses or viroids, such as tobacco or cucumber mosaic virus, ring spot or necrosis virus, pelargonium leaf roll virus, mottled red clover virus, tomato shrub wilt virus, and similar viruses; fungi Ascomycetes, such as genera Venturia, Podosphaera, Erysiphe, Monolinia, Mycosphaerella, and Uncinula; Basidiomycete fungi, such as from the genera Hemi -leia, Rhizoctoria, and Puccinia; imperfect fungi, such as the genera Botrytis, Helminthosporium, Rhynchosporium, Fusarium (ie, F. monoli form), Septoria, Cercospora, Al ternarla, Pyricularia, and Pseudocercosporella (ie, P. herpotrichoi -des); Oomycete fungi, such as from the genera Phytophthora (ie P. parasí tica), Peronospora (ie P. tabacina), Bremia, Pythium, and Plasmopara, - as well as other fungi, such as Scleropthora macrospora, Sclerophthora rayissiae, Sclerospora graminicola, Peronosclerospora sorghi, Peronosclerospora phiippinensis, Peronosclerospora sacchari and Peronosclerospora maydis, Physopella zeae, Cercospora zeae-maydis, Colletotrichum graminicola, Gibberella zeae, Exserohilum turcicum Kabatiellu zeae, and Bipolaris maydis; bacteria, such as Pseudomonas syringae, Pseudomonas tabaci, and Erwinia stewartir; insects, such as aphids, for example Myzus persicae; and lepidoptera, such as Heliothus spp; and nematodes such as Meloidogyne incognita.

Obtaining Immunomodulated Plants The three following general routes for obtaining immunomodulatory plants are related because they all fit in the model of the transduction pathway of the acquired systemic resistance signal stipulated in Ryals et al. (1996). By activating the transduction pathway of the acquired systemic resistance signal to achieve disease resistance, the same set of acquired systemic resistance genes is "activated", and disease resistance results, regardless of which of the three routes described. later take it. The differences between these three routes belong only to the point at which the acquired systemic resistance pathway is activated; The final result is the same among the three routes. Accordingly, the analyzes and results observed with respect to the immunomodulated plants obtained through a route can be extrapolated and applied to the immunomodulated plants obtained through a different route.

A. Application of a Chemical Inductor of Acquired Systemic Resistance A first route to obtain immunomodulated plants involves applying to a plant a chemical capable of inducing acquired systemic resistance. Particularly potent chemical inducers of acquired systemic resistance are benzothiadiazoles, such as S-methyl ester of ben-zo [1,2,3] thiadiazole-7-carbothioic acid (BTH). Benzothiadiazole derivatives that can be further used as regulators are described in U.S. Patent Nos. 5,523,311 and 5,614,395, which are incorporated herein by reference. The acquired systemic resistance induced by BTH, which provides protection in the field against a broad spectrum of diseases in a variety of crops, is described in detail in Freidrich et al., Plant Journal 10 (1), 61-70 (1996); Lawton et al., Plant Journal 10 (1), 71-82 (1996); and Gorlach et al., Plant Cell 8, 629-643 (1996), each of which is incorporated herein by reference. Other chemical inducers of acquired systemic resistance that can be used to obtain an immunomodulated plant for use in the method of the invention include isonicotinic acid compounds, such as 2,6-dichloroisonicotinic acid (INA), and its lower alkyl esters, as well as the salicylic acid (SA) compounds. Examples of suitable INA and SA compounds are described in U.S. Patent Number 5,614,395.

B. Reproduction of Mutant Plants of Constitutive Immunity (CIM). A second route to obtain immunomodulated plants is through a selective breeding program based on the constitutive expression of acquired systemic resistance genes and / or a disease-resistant phenotype. There are considerable data showing a close correlation between the expression of acquired systemic resistance genes and the acquired systemic resistance itself (Ward et al. (1991); Uknes et al. (1992); Uknes et al. (1993); Lawton et al. 1993) and Alexander and collaborators (1993) PNAS USA 90, 7327-7331, incorporated herein by reference). In Arabidopsis, examples of well-characterized acquired systemic resistance genes are PR-1, PR-2, and PR-5, with PR-1 expressed at the highest level with the lowest background. To identify and select plants that constitutively express acquired systemic resistance genes, Northerm analysis is performed to detect the expression of acquired systemic resistance genes. Acquired systemic resistance DNA sequences known in cross-hybridization experiments can be used, as described in Uknes et al. (1992). Methods for hybridization and cloning of nucleic acid sequences are well known in the art. (See, for example, Molecular Cloning, A Laboratory Manual, 2nd edition, volumes 1-3, Sambrook et al. (editors), Cold Spring Harbor Laboratory Press (1989) and references cited therein). At least two classes of transduction mutants of the acquired systemic resistance signal, which constitutively express the acquired systemic resistance genes, have been isolated. One class has been designated as "Jsd" mutants (Isd = lesion simulating disease), which are also referred to as "Class I cim" mutants. See International Publication Number WO 94/16077. Isd (cim Class I) mutants form spontaneous lesions on leaves, accumulated high concentrations of salicylic acid, high levels of mRNA of PR-1, PR-2, and PR-5 and are resistant to fungal and bacterial pathogens (Dietrich et al., 1994; Weymann et al., 1995). A second class has been designated as "cim" mutants (cim = constitutive immunity, constitutive immunity), which are also referred to as "Class II cim mutants". See International Publication Number WO 94/16077. The cim mutants have all the characteristics of the Isd mutants, with the exception of spontaneous lesions. That is, the cim mutants are visibly phenotypically normal. Once the plants constitutively expressing the acquired systemic resistance genes are selected, they can be used in breeding programs to incorporate the constitutive expression of the genes of acquired systemic resistance and pathogen resistance in plant lines. The offspring for additional crosses are selected based on the expression of the genes of acquired systemic resistance and disease resistance, as well as on other characteristics important for production and quality according to methods well known to the experts in culture. the technique of plant reproduction. For example, because the Isd mutants exhibit lesion formation and necrosis, the cim mutants with their normal phenotypes are preferable for use in these breeding programs and in the method of the present invention, although if desired, the mutants Jsd.

C. Transformation of Plants with Acquired Systemic Resistance Genes A third route to obtain immunomodulated plants is by transforming the plants with an acquired systemic resistance gene, preferably a functional form of the NIM1 gene. 1. Recombinant Expression of the Wild-type NIM1 Gene The recombinant overexpression of the wild-type form of NJ 1 (SEQ ID? 0: 1) gives rise to transgenic plants with a disease-resistant phenotype. See U.S. Patent Application Pending Serial Number 08 / 880,179, incorporated herein by reference. Increased levels of active NIM1 protein produce the same disease resistance effect as chemical induction with chemical inducers, such as BTH, INA, and SA. Preferably, the expression of the NIM1 gene is at a level that is at least two times higher than the level of expression of the NIM1 gene in wild type plants, and more preferably is at least 10 times above the level of expression of the wild type. The next section, entitled "Recombinant DNA Technology" stipulates protocols that can be used to recombinantly express the wild type NIM1 gene in transgenic plants at higher levels than the wild type. Alternatively, plants with the wild type NPRl gene can be transformed to produce disease resistant plants, as described in Cao et al. (1997). 2. Recombinant Expression of an Altered Form of the NIM1 Gene Immunomodulatory plants can also be created for use in the method of the present invention, by the recombinant expression of an altered form of the NIM1 gene, whereby the alteration of the NIM1 gene exploits both the recognition that the route of systemic resistance acquired in plants shows functional parallels with the regulation scheme of NF-? B / l? B in mammals and in flies, as well as the discovery that the NIM1 gene product is a structural homolog of the factor of transduction of mammalian signal I? B, subclass a. See the pending TCP Request "METHODS OF USING THE NJM1 GENE TO CONFER DISEASE RESISTANCE IN PLANTS" incorporated herein by reference. The sequence of the NIM1 gene (SEQ ID NO: 1) was used in BLAST searches, and matches were identified based on the homology of a rather highly conserved domain in the genetic sequence of NIM1, with the ankyrin domains found in a number of proteins, such as spectrins, ankyrins, in NF-KB and I? B (Michaely and Bennett, Trends Cell Biol, 2, 127-129 (1992)). Visual paired inspections were performed between the NIM1 protein (SEQ ID NO: 2), and 70 known ankyrin-containing proteins, and striking similarities were found with members of the I? B transcription regulator class (Baeuerle and Baltimore 1996; nineteen ninety six) . As shown in Figure 1, the NIM1 protein (SEQ ID NO: 2) shares significant homology with the IβB proteins of mouse, rat, and pig (SEQ ID NOs: 3, 4, and 5, respectively). NIM1 contains several important structural domains of I? Ba throughout the entire length of the protein, including ankyrin domains (indicated by the lower diagonal labeling in Figure 1), 2 amino-terminal serines (amino acids 55 and 59 NIM1) , a pair of power plants (amino acids 99 and 100 in-NIMl), and an acidic C-terminus. Above all, NIM1 and I? Ba share identity in 30 percent of the waste, and conservative replacements in 50 percent of the waste. Therefore, there is homology between I? B and NIM1 across all proteins, with an overall similarity of 80 percent. One way in which the IKBOI protein works in signal transduction is by binding to the cytosolic transcription factor NF-KB, and preventing it from entering the nucleus, and altering the transcription of the target genes (Baeuerle and Baltimore, 1996; Baldwin, 1996). The NF-KB target genes regulate (activate or inhibit) several cellular processes, including antiviral, antimicrobial, and cell death responses (Baeuerle and Baltimore, 1996). When the signal transduction pathway is activated, I? Ba is phosphorylated on two serine residues (amino acids 32 and 36 of mouse I? Ba). This programs the ubiquitination in a double lysine (amino acids 21 and 22 of mouse I? Ba). Following ubiquitination, the NF-? B / L? B complex is directed through the proteasome, where the I? Ba degrades, and NF-? B is released to the core. The phosphorylated serine residues important in the I? Ba function are conserved in NIM1 within a large contiguous block of sequence conserved from amino acids 35 to 84 (Figure 1). In contrast to I? Ba, where double lysine is located at about 15 amino acids in front of the N-terminus of the protein, at NIM1, a double lysine is located at about 40 amino acids in front of the C-terminus. In addition, there is a high degree of homology between NIM1 and I? Ba in the carboxyl terminal region rich in serine / threonine, which has been shown to be important in the rate of basal change (Sun et al., Mol. Cell. Biol. 16, 1058-1065 (1996)). According to the present invention, based on structural homology analysis and on the presence of elements known to be important for the function of I? B, NIM1 is expected to function as I? B, having analogous effects on regulation genetics of plants. It is predicted that plants containing the wild-type NTM1 gene, when treated with chemical inducers, have more product of the NIM1 gene (I? B homolog), or less phosphorylation of the NIM1 gene product (I? B homologue). ). According to this model, the result is that the homologue? F-? B of the plant is kept outside the nucleus, and expression responses of the acquired systemic resistance and resistance gene are allowed to occur. In neither the mutant plants, a non-functional NIM1 gene product is present. Accordingly, according to this model, the homologue F-KB is free to go to the nucleus and repress the resistance and expression of the acquired systemic resistance gene. Consistent with this idea, animal cells treated with salicylic acid show a greater stability / abundance of I? B, and a reduction of active? F-KB in the nucleus (Kopp and Ghosh, 1994). It is known that I? B mutations act as super-repressors or negative-dominants (Britta-Mareen Traenckner et al., EMBO 14: 2876-2883 (1995); Brown et al., Science 267: 1485-1488 (1996); Brockman et al., Molecular and Cellular Biology 15: 2809-2818 (1995); Wang et al., Science 274: 784-787 (1996)). These mutant forms of I? B bind to NF-KB, but are not phosphorylated or ubiquitinated, and therefore, do not degrade. NF-KB remains linked to I? B, and can not move towards the core. In view of the above, altered forms of NJM1 can be created that act as negative-dominant regulators of the transduction pathway of the acquired systemic resistance signal. Plants transformed with these negative-dominant forms of NIM1 have the opposite phenotype to that of niml mutant plants, in which plants transformed with altered forms of NIM1 exhibit the constitutive expression of the acquired systemic resistance gene, and consequently, a CIM phenotype; that is, the transgenic plants are immunomodulatory. Because the position of the NIM1 gene is maintained in the transduction pathway of the acquired systemic resistance signal, it is expected that a number of alterations to the gene, beyond those specifically disclosed herein, will result in the expression constitutive of acquired systemic resistance genes, and consequently, a CIM phenotype. The following section entitled "Recombinant AD Technology" stipulates protocols that can be used to express recombinantly the altered forms of the NIM1 gene in transgenic plants, at higher levels than in the wild type. Later, several altered forms of the NIM1 gene are described that act as negative-dominant regulators of the transduction pathway of the acquired systemic resistance signal. to. Changes in Serine Residues 55 and 59 to Alanine Residues: Phosphorylation of the serine residues in human I? B is required for the degradation activated by stimulation of I? B, thus activating NF-B. The mutagenesis of the serine residues (S32 and S36) in I? Ba human to alanine residues, inhibits stimulus-induced phosphorylation, thus blocking the degradation mediated by the I? B proteasome (Traenckner et al., 1995; Brown et al., 1996; Brockman et al., 1995; and collaborators, 1996). This altered form of I? B can function as a negative-dominant form by retaining NF-? B in the cytoplasm, thereby blocking downstream signaling events. Based on the comparison of the amino acid sequence between NIM1 and I? B shown in Figure 1, serines 55 (S55) and 59 (S59) in NIM1 (SEQ ID NO: 2) are homologous to S32 and S36 in I? Human Ba To construct negative-dominant forms of NIM1, the serines at amino acid positions 55 and 59 are mutated to alanine residues. Accordingly, in a preferred embodiment, the NIM1 gene is altered, such that the encoded product has alanines in place of serines at the amino acid positions corresponding to positions 55 and 59 of the amino acid sequence of NIM1 of Arabidopsis (SEQ. ID NO: 2). b. N-terminal suppression: The deletion of amino acids 1-36 (Brockman et al., 1995; Sun et al., 1996), or 1-72 (Sun et al., 1996) of human I? Ba, which includes lysine residues K21 and K22 of ubiquitination, as well as the phosphorylation sites S32 and S36, result in a negative I-Ba phenotype in transfected human cell cultures. An N-terminal deletion of the first 125 amino acids of the NIM1 gene product will remove 8 lysine residues that could serve as ubiquitination sites, as well as the putative phosphorylation sites in S55 and S59 discussed above. Therefore, in a preferred embodiment, the NIM1 gene is altered, such that the encoded product is missing approximately in the first 125 amino acids, compared to the amino acid sequence of NIM1 of native Arabidopsis (SEQ ID NO: 2). c. C-terminal Suppression: The deletion of amino acids 261-317 from human I? Ba may result in better intrinsic stability, by blocking the constitutive phosphorylation of the serine and trionine residues at the C-terminus. this altered form of I? B functions as a negative-dominant form. A region rich in serine and threonine is present at amino acids 522-593 of the C-terminus of NIM1. Accordingly, in a preferred embodiment, the NIM1 gene is altered, so that the encoded product is missing in approximately its C-terminal portion including amino acids 522-593, compared to the native amino acid sequence of NIM1 of Arabidopsis (SEQ ID. NO: 2). d. Chimera and Ankyrin Domains of N-terminal Suppression / C-erminal Altered forms of the NIM1 gene product may also occur as a result of deletions or chimeras from the C-terminal and N-terminal segments. In still another embodiment of the present invention, constructs comprising the ankyrin domains from the NIM1 gene are provided. 3. Recombinant Expression of Other Acquired Systemic Resistance Genes. Immunomodulated plants can also be created for use in the method of the present invention, by recombinant expression of different acquired systemic resistance genes, such as those described in Ward et al., (1991). See, for example, United States Patent Number 5,617,395, which describes plants resistant to diseases created by overexpression of one or more PR protein genes. Although it refers to the recombinant expression of NIM1 gene forms in particular, the following section entitled "Recombinant DNA Technology" stipulates protocols that can also be used to recombinantly express other genes of acquired systemic resistance, such as protein genes. PR in transgenic plants, at levels higher than the wild type.

Recobinante DNA technology The wild-type form or the altered form of the gene NIM1, which confers resistance to plant diseases, improving the expression of the acquired systemic resistance gene, can be incorporated into plant cells using conventional recombinant DNA technology. In general, this involves inserting the DNA molecule encoding the selected form of NIM1 described above, into. an expression system with which the DNA molecule is heterologous (i.e., which is not normally present), using conventional cloning procedures known in the art. The vector contains the necessary elements for the transcription and translation of the protein coding sequences inserted. A large number of vector systems known in the art can be used, such as plasmids, bacteriophage viruses, and other modified viruses. Suitable vectors include, but are not limited to, viral vectors, such as the lambda? Gtll,? GtlO and Charon 4 vector systems; plasmid vectors, such as pBI121, pBR322, pACYC177, pACYC184, the pAR series, pKK223-3, pUC8, pUC9, pUC18, pUC19, pLG339, pRK290, pKC37, pKClOl, pCDNAII; and other similar systems. The components of the expression system can also be modified to increase expression. For example, truncated sequences, nucleotide substitutions or other modifications may be employed. The expression systems described herein can be used to transform virtually any cell of a crop plant under suitable conditions. The transformed cells can be regenerated in whole plants, such that the selected form of the NJM1 gene activates the systemic resistance acquired in the transgenic plants.

A. Construction of Expression Cassettes in Plants First genetic sequences are assembled, intended for their expression in transgenic plants, in expression cassettes behind a suitable promoter that can be expressed in plants. The expression cassettes may also comprise any additional sequences required or selected for the expression of the transgene. These sequences include, but are not restricted to, transcription terminators, foreign sequences to improve expression, such as introns, vital sequences, and sequences intended to direct the gene product toward specific cell organelles and compartments. These expression cassettes can then be easily transferred to the transformation vectors in plants described below. The following is a description of different components of typical expression cassettes. 1. Promoters The selection of promoter - used in the expression cassettes, will determine the spatial and temporal expression pattern of the transgene in the transgenic plant. The selected promoters will express the transgenes in specific cell types (such as leaf epidermal cells, mesophyll cells, cells of the root cortex), or in specific tissues or organs (roots, leaves, or flowers, for example) , and the selection will reflect the desired location of accumulation of the genetic product. In an alternative way, the selected promoter can direct the expression of the gene under different induction conditions. The promoters vary in their strength, that is, their ability to promote transcription. Depending on the host cell system used, any of a number of suitable promoters can be used. The following are non-limiting examples of promoters that can be used in expression cassettes. to. Constitutive Expression, CaMV 35S Promoter: The construction of plasmid pCGN1761 is described in Published Patent Application Number EP-0,392,225 (Example 23), which is incorporated herein by reference. pCGN1761 contains the 35S promoter of "double" CaMV, and the tml transcription terminator with a unique EcoRI site between the promoter and the terminator, and has a base structure of type pUC. A derivative of pCGN1761 is constructed, which has a modified polylinker that includes the Notl and Xhol sites in addition to the existing EcoRI site. This derivative is designated pCGN1761ENX pCGN1761ENX is useful for the cloning of 7DNA sequences or genetic sequences (including open reading frame microbial sequences) inside its polylinker, for the purpose of its expression under the control of the 35S promoter in transgenic plants. All the cassette of promoter 35S-tml gene-terminator sequence of this construction can be excised by the sites HidlII, Sphl, Salí, and Xbal 5 'for the promoter, and by the sites Xbal, BamHl, and Bgll 3' for the terminator , to be transferred to transformation vectors, such as those described below. Additionally, the double 35S promoter fragment can be removed by 5 'cleavage with HindIII, SphI, SalI, Xbal, 6 PstI, and 3' cleavage with any of the polylinker restriction sites (EcoRI, Notl, or Xhol), to the replacement with another promoter. If desired, modifications can be made around the cloning sites by introducing sequences that can improve translation. This is particularly useful when overexpression is desired. For example, pCGN1761ENX can be modified by optimization of the translation initiation site, as described in Example 37 of U.S. Patent No. 5,639,949 incorporated herein by reference. b. Expression Under a Chemically Regulatory / Pathogenic Promoter: The double 35S promoter of pCGN1761ENX can be replaced with any other promoter of choice that results in adequately high expression levels. By way of example, one of the chemically-regulatable promoters described in U.S. Patent No. 5,614,395 can replace the double 35S promoter. The promoter of choice is preferably cleaved from its source by restriction enzymes, but alternatively it can be amplified by the Polymerase Chain Reaction, using primers carrying appropriate terminal restriction sites. If the amplification is undertaken with the Polymerase Chain Reaction, then the promoter must be sequenced again to verify the amplification errors after the cloning of the amplified promoter in the target vector. The chemically / pathogenically regulatable tobacco PR-promoter is dissociated from plasmid pCIB1004 (for construction, see Example 21 of European Patent Number EP-0,332, 104, which is incorporated herein by reference), and transferred to the plasmid pCGN1761ENX (Uknes et al., 1992). pCIB1004 is dissociated with Ncol and the 3 'pendant resulting from the linearized fragment is made blunt by its treatment with T4 DNA polymerase. The fragment is then dissociated with HindIII, and the fragment containing the resulting PR-la promoter is gel purified and cloned into pCGN1761ENX, from which the double 35S promoter has been removed. This is done by dissociation with XhoI, and blunting with T4 polymerase, followed by dissociation with HindIII, and isolation of the fragment containing the terminator of the larger vector, in which the promoter fragment pCIB1004 is cloned. This generates a derivative of pCGN1761ENX with the PR-la promoter and the tml terminator, and an intervening polylinker with unique EcoRI and NotI sites. The selected coding sequence can be inserted into this vector, and subsequently the fusion products (i.e., promoter-gene-terminator) can be transferred to any selected transformation vector, including those described below. Various chemical regulators can be employed to induce the expression of the selected coding sequence in transformed plants according to the present invention, including the benzothiadiazole, isonicotinic acid, and salicylic acid compounds disclosed in US Pat. United States of America Numbers 5,523,311 and 5,614,395. c. Constitutive Expression, the Actin Promoter: It is known that several isoforms of actin are expressed in most cell types, and consequently, the actin promoter is a good choice for a constitutive promoter. In particular, the promoter has been cloned and characterized from the rice ActI gene (McElroy et al., Plant Cell 2 .: 163-171 (1990)). It was found that a 1.3 kb fragment of the promoter contains all the regulatory elements required for expression in rice protoplasts. In addition, numerous expression vectors based on the ActI promoter have been specifically constructed for use in monocots.

(McElroy et al., Mol. Gen. Genet, 231: 150-160 (1991)).

These incorporate the flanking sequence 5 'of Adhl, Actl-intron 1, and Adhl-intron 1 (from the corn alcohol dehydrogenase gene), and the sequence from the 35S promoter of CaMV. The vectors that showed the highest expression were 35S fusions and the ActI intron, or the 5 'flanking sequence of ActJ and the ActI intron. The optimization of the sequences around start ATG (of the GUS reporter gene) also improved the expression. The expression cassettes of the promoter described by McElroy et al. (Gen. Genet 22: 150-160). (1991), can be easily modified for gene expression, and are particularly suitable for use in monocotyledonous hosts. For example, fragments containing the promoter of the McElroy constructs are removed, and used to replace the double 35S promoter in pCGN1761ENX, which is then available for the insertion of specific genetic sequences. The fusion genes thus constructed can then be transferred to appropriate transformation vectors. In a separate report, it has also been found that the ActI promoter of rice with its first intron directs high expression in cultured barley cells (Chibbar et al., Plant Cell Rep. 12: 506-509 (1993)). d. Constitutive Expression, the Ubiquitin Promoter: Ubiquitin is another genetic product that is known to accumulate in many cell types, and its promoter has been cloned from several species for use in transgenic plants (eg, sunflower - Binet and collaborators, Plant Science 79: 87-94 (1991), and corn - Christensen et al., Plant Molec Biol. 12: 619-632 (1989)). The corn ubiquitin promoter has been developed in transgenic monocotyledonous systems, and its sequence and vectors for monocot transformation have been constructed, and are disclosed in European Patent Publication Number EP-0,342,926 (to Lubrizol), which it is incorporated herein by reference. Taylor et al. (Plant Cell Rep. 12: 491-495 (1993)) describe a vector (pAHC25) comprising the maize ubiquitin promoter and the first intron, and its high activity in cell suspensions of numerous monocotyledons when introduced by means of bombing microprojectiles. The ubiquitin promoter is suitable for gene expression in transgenic plants, especially monocotyledons. Suitable vectors are derivatives of pAHC25, or any of the transformation vectors described in this application, modified by the introduction of the ubiquitin promoter and / or appropriate introns sequences. and. Root Specific Expression: Another pattern of gene expression is the expression of the root. A suitable root promoter is described by de Framond (FEBS 290: 103-106 (1991)), and also in Published Patent Application Number EP-0,452, 269, which is incorporated herein by reference. This promoter is transferred to a suitable vector, such as pCGN1761ENX, for the insertion of a selected gene, and the subsequent transfer of the entire promoter-gene-terminator cassette to a transformation vector of interest. f. Wound-Induced Promoters: Wound-inducible promoters may also be suitable for gene expression. Numerous of these promoters have been described (eg, Xu et al., Plant Molec. Biol. 22: 573-588 (1993), Logemann et al., Plant Cell 1: 151-158 (1989), Rohrmeier and Lehle, Plant Molec, Biol. 22: 783-792 (1993), Firek et al., Plant Molec.

Biol. 2? 129-142 (1993), Warner et al. Plant J. 2 .: 191-201 (1993)), and all are suitable for use with the present invention. Logemann et al. Describe the 5 'upstream sequences of the wunl gene of dicotyledonous potato. Xu et al. Show that a promoter inducible by wound from the dicotyledonous potato (pin2), is active in the monocotyledonous rice. In addition, Rohrmeier and Lehle describe the cloning of maize WipI cDNA that is induced by wound, and that can be used to isolate the known promoter using conventional techniques. In a similar manner, Firek et al. And Warner et al. Have described a wound-induced gene from the monocot Asparagus officinalis, which is expressed at sites of injury and invasion of local pathogens. Using cloning techniques well known in the art, these promoters can be transferred to suitable vectors, can be fused with the genes belonging to this invention, and can be used to express these genes at the wound sites of the plant. g. Preferred Expression by Sap: Patent Application Number WO 93/07278, which is incorporated herein by reference, describes the isolation of the maize trpA gene, which is preferentially expressed in sap cells. The genetic sequence and promoter are presented, which extends to -1726 base pairs from the start of transcription. Using conventional molecular biological techniques, this promoter, or parts thereof, can be transferred to a vector, such as pCGN1761, where it can replace the 35S promoter, and can be used to drive the expression of a foreign gene in a preferred manner. the SAP. In fact, fragments containing the preferred promoter by the sap, or parts thereof, can be transferred to any vector, and can be modified to have utility in transgenic plants. h. Specific Leaf Expression: A corn gene that encodes phosphoenol carboxylase (PEPC), has been described by Hudspeth and Gruía (Plant Molec Biol 12: 579-589 (1989)). Using conventional molecular biological techniques, the promoter can be used for this gene, in order to direct the expression of any gene in a leaf-specific manner in transgenic plants. 2. Transcription Terminators There are a variety of transcription terminators available for use in expression cassettes. These are responsible for the termination of transcription beyond the transgene and its correct polyadenylation. Suitable transcription terminators are those that are known to work in plants, and include the CaMV 35S terminator, the tml terminator, the nopaline synthase terminator, and the rbcS E9 pea terminator. These can be used in both monocots and dicots. 3. Sequences for Improving or Regulating Expression It has been found that numerous sequences improve gene expression from within the transcription unit, and these sequences can be used in conjunction with the genes of this invention, to increase their expression in transgenic plants. It has been shown that different sequences of introns improve expression, particularly in monocotyledonous cells. For example, it has been found that corn Adhl gene introns, significantly improve the expression of the wild-type gene under its known promoter, when introduced into corn cells. Intron 1 was found to be particularly effective, and improved expression in fusion constructs with the chloramphenicol acetyltransferase gene (Callis et al., Genes Develop 1: 1183-1200 (1987)). In the same experimental system, the intron of the corn bronzel gene had a similar effect to improve expression. Intron sequences have been routinely incorporated into transformation vectors in plants, usually within the non-translated leader. It is also known that a number of untranslated leader sequences derived from viruses improve expression, which are particularly effective in dicotyledonous cells. Specifically, it has been shown that the leader sequences from Tobacco Mosaic Virus (TMV, the "Sequence W), Corn Chlorotic Speck Virus (MCMV), and Alfalfa Mosaic Virus (AMV), are effective in improving the expression (for example, Gallie et al., Nucí Acids Res. lü .: 8693-8711 (1987); Skuzeski et al., Plant Molec. Bíol 15 .: 65-79 (1990) ) . 4. Direction of the Genetic Product Into the Cell It is known that in plants there are different mechanisms to direct the genetic products, and the sequences that control the functioning of these mechanisms have been characterized in some detail. For example, the direction of the gene products towards the chloroplast is controlled by a signal sequence that is at the amino-terminal end of different proteins, which dissociates during the chloroplast import, to produce the mature protein (for example, Comai et al., J. Biol. Chem. 2 £ 2: 15104-15109 (1988)). These signal sequences can be fused with heterologous gene products to effect the importation of the heterologous products into the chloroplast (van den Broeck, et al., Nature 3L3 .: 358-363 (1985)). The DNA encoding the appropriate signal sequences can be isolated from the 5 'end of the cDNAs encoding the RUBISCO protein, the CAB protein, the EPSP synthase enzyme, the GS2 protein, and many other proteins known to be they locate in the chloroplast. See also the section entitled "Expression with Chloroplast Direction" in Example 37 of the United States Patent Number 5,639,949. Other genetic products are located in other organelles, such as the mitochondria and the peroxisome (for example, Unger et al., Plant Molec. Biol. 13: 411-418 (1989)). The cDNAs encoding these products can also be manipulated to direct the heterologous gene products towards these organelles. Examples of these sequences are the ATPases encoded by the nucleus, and the specific isoforms of aspartate aminotransferase for the mitochondria. The direction of cellular protein bodies has been described by Rogers et al. (Proc. Nati, Acad. Sci. USA £ 2: 6512-6516 (1985)). In addition, sequences responsible for the direction of the gene products towards other cell compartments have been characterized. The amino-terminal sequences are responsible for the direction towards the endoplasmic reticulum, the apoplast, and the extracellular secretion from the aleurone cells (Koehier and Ho, Plant Cell 2: 769-783 (1990)). In addition, the amino-terminal sequences, in conjunction with the carboxy-terminal sequences, are responsible for the vacuolar direction of the gene products (Shinshi et al., Plant Molec, Biol. A.: 357-368 (1990)). By fusing the appropriate targeting sequences described above, to the transgenic sequences of interest, it is possible to direct the transgenic product to any organelle or cell compartment. For chloroplast targeting, for example, the chloroplast signal sequence from the RUBISCO gene, the CAB gene, the EPSP synthase gene, or the GS2 gene, is fused within the framework with the amino-terminal ATG of the transgene. The selected signal sequence must include the known dissociation site, and the constructed fusion must take into account any amino acids after the dissociation site that are required for dissociation. In some cases, this requirement can be met by adding a small number of amino acids between the dissociation site and ATG of the transgene, or alternatively, the replacement of some amino acids within the transgenic sequence. Mergers constructed to be imported into the chloroplast can be tested to determine the efficiency of chloroplast recovery by translating in vitro transcribed constructions, followed by recovery of the chloroplast in vi tro, using the techniques described by Bartlett et al., In: Edelmann et al. (Editores) Methods in Chloroplast Molecular Biology, Elsevier, pages 1081-1091 (1982), and Wasmann et al., Mol. Gen Genet 205: 446-453 (1986). These construction techniques are well known in the art, and are equally applicable to mitochondria and peroxisomes. The mechanisms described above for cell targeting can be used, not only in conjunction with their known promoters, but also in conjunction with heterologous promoters, to effect a specific cell targeting goal, under the transcription regulation of a promoter having a standard of expression different from that of the promoter from which the directional signal is derived.

B. Plant Transformation Vectors Constructions Experts from ordinary experience in the plant transformation technique know of numerous transformation vectors available for plant transformation, and the relevant genes for this invention can be used in conjunction with any of those vectors The selection of the vector will depend on the preferred transformation technique and the target species for the transformation. For certain white species, different antibiotic or herbicide selection markers may be preferred. The selection markers routinely used in the transformation include the nptll gene, which confers resistance to kanamycin and related antibiotics (Messing and Vierra, Gene 12: 259-268 (1982); Bevan et al., Nature 304: 184-187 81983)), the bar gene, which confers resistance to the herbicide phosphinothricin (White et al., Nucí Acids REs 18: 1062 (1990), Spencer et al., Theor. Appl. Genet 79 : 625-631 (1990)), the hph gene, which confers resistance to the antibiotic hygromycin (Blochinger and Diggelmann, Mol Cell Biol 4: 2929-2931), and the dhfr gene, which confers resistance to methotrexate (Bourouis et al. collaborators, EMBO J. 2 (7): 1099-1104 (1983)), and the EPSPS gene, which confers resistance to glyphosate (Patents of the United States of North America Nos. 4,940,935 and 5,188,642). 1. Vectors Suitable for Transformation with Agrobacterium There are many vectors available for transformation using Agrobacterium tumefaciens. These normally carry at least one T-DNA limit sequence, and include vectors such as pBIN19 (Bevan, Nucí Acids Res. (1984)) and pXYZ. Below, the construction of two typical vectors suitable for transformation with Agrobacterium is described. to. pCIB200 and pCIB2001: The binary vectors pCIB200 and pCIB2001 are used for the construction of recombinant vectors, for use with Agrobacterium, and are constructed in the following manner. PTJS75kan is created by digestion with NarI of pTJS75 (Schmidhauser and Helinski, J. Bacteriol 164: 446-455 (1985)) allowing cleavage of the tetracycline resistance gene, followed by the insertion of an Accl fragment from pUC4K, which carries an NPTII (Messing and Vierra, Gene 19_ 259-268 (1982); Bevan and collaborators, Nature 304: 184-187 (1983): McBride et al., Plant Molecular Biology 14: 266-276 (1990 )). Xhol linkers are ligated to the EcoRV fragment of PCIB7, which contains the boundaries of left and right T-DNA, a chimeric us / nptll gene selectable in plants, and the pUC polylinker (Rothstein and collaborators, Gene 53: 153 -161 (1987)), and the fragment digested with XhoI is cloned into pTJS75kan digested with SalI to create pCIB200 (see also European Patent Number EP-0, 332, 104, Example 19). pCIB200 contains the following unique polylinker restriction sites: EcoRI, SstI, Kpnl, BglII, Xbal, and SalI. pCIB2001 is a derivative of pCIB200 created by inserting additional restriction sites into the polylinker. The unique restriction sites in the polylinker of pCIB2001 are EcoRI, SstI, Kpnl, BglII, XbaI, SalI, MIuI, Bel I, Avrll, Apal, Hpal, and Stul. pCIB2001, in addition to containing these unique restriction sites, also has selection of kanamycin in plants and bacteria, left and right T-DNA boundaries for Agrobacterium-mediated transformation, the function of trfA derived from RK2 for mobilization between E coli and other hosts, and the OriT and OriV functions also from RK2. The polylinker pCIB2001 is suitable for the cloning of expression cassettes in plants containing their own regulatory signals. b. pCIBlO and Hygromycin Selection Derivatives of the Same: The binary vector pCIBlO contains a gene that encodes kanamycin resistance for selection in plants, and the right and left border sequences of T-DNA, and incorporates sequences from the wide-ranging plasmid of pRK252 hosts, which allow it to replicate in both E. coli and Agrobacterium. Its construction is described by Rothstein et al. (Gene 52.: 153-161 (1987)). Different pCIBlO derivatives are constructed which incorporate the gene for hygromycin B phosphotransferase described by Gritz et al. (Gene 2-5 .: 179-188 (1983)). These derivatives make it possible to select cells from transgenic plants on hygromycin alone (pCIB743), or hygromycin and kanamycin (pCIB715, pCIB717). 2. Vectors Suitable for Transformation without Agroba. cteri um Transformation without the use of Agrobacterium tumefaciens circumvents the requirement for T-DNA sequences in the selected transformation vector, and consequently, vectors lacking this sequence can be used in addition to vectors such as those described above. -te, which contain T-DNA sequences. Transformation techniques that do not rely on Agrobacterium include transformation by means of particle bombardment, protoplast recovery (e.g., PEG and electroporation), and microinjection. The choice of vector depends largely on the preferred selection for the species being transformed. Below, the construction of typical vectors suitable for transformation without Agrobacterium is described. to. pCIB3064: pCIB3064 is a vector derived from pUC suitable to direct the direct gene transfer technique in combination with selection by the coarse herbicide (or phosphinothricin). Plasmid pCIB246 comprises the CaMV 35S promoter in fusion operable with the GUS gene of E. coli, and the CaMV 35S transcription terminator, and is described in Published PCT Application No. WO 93/07278. The 35S promoter of this vector contains two 5 'sequences of ATG from the start site. These sites are mutated using conventional techniques of the Polymerase Chain Reaction, in such a way that the ATGs are removed, and the SspI and PvuII restriction sites are generated. The new restriction sites are 96 and 37 base pairs from the single Salí site, and 101 to 42 base pairs from the actual start site. The resulting derivative of pCIB246 is designated pCIB3025. The GUS gene of pCIB3025 is then cleaved by digestion with Sali and SacI, the ends blunted, and ligated again to generate the plasmid pCIB3060. Plasmid pJIT82 is obtained from John Innes Center, Norwich, and the 400 base pair Smal fragment containing the bar gene from Streptomyces viridochromogenes is excised and inserted into the Hpal site of pCIB3060 (Thompson et al., EMBO J £ : 2519-2523 (1987)). This generates pCIB3064, which comprises the bar gene under the control of the 35S promoter of CaMV, and the terminator for the selection of the herbicide, a gene for resistance to ampicillin (for selection in E. coli), and a polylinker with the unique sites Sphl, PstI, HindIII, and BamHl. This vector is suitable for the cloning of expression cassettes in plants that contain their own regulatory signals. b. pSOG19 and pSOG35: pSOG35 is a transformation vector that uses the dihydrofolate reductase of the E. coli gene (DFR) as a selectable marker that confers resistance to methotrexate. The Polymerase Chain Reaction is used to amplify the 35S promoter (-800 base pairs), intron 6 from the Adhl maize gene (-550 base pairs), and 18 base pairs of the leader sequence. translated from GUS from pSOGlO. The 250 base pair fragment encoding the type II dihydrofolate reductase gene from E. coli is also amplified by the Polymerase Chain Reaction, and these two fragments of the Polymerase Chain Ration are assembled with a Sacl-PstJ fragment from pB1221 (Clontech), which comprises the base structure of the pUC19 vector, and the synthase terminator of the polymerase chain. nopaline The assembly of these fragments generates pS0G19, which contains the 35S promoter in fusion with the intron 6 sequence, the GUS leader, the DHFR gene, and the nopaline synthase terminator. The replacement of the GUS leader in pS0G19 with the leader sequence from Corn Chlorotic Speck Virus (MCMV) generates the pSOG35 vector. pS0G19 and pSOG35 carry the pUC gene for ampicillin resistance, and have the HindIII, Sphl, PstJ, and EcoRI sites available for the cloning of foreign substances.

C. Transformation Once the coding sequence of interest in an expression system has been cloned, it is transformed into a plant cell. Methods for the transformation and regeneration of plants are well known in the art. For example, Ti plasmid vectors have been used for the delivery of foreign DNA, as well as direct DNA recovery, liposomes, electroporation, microinjection, and microprojectiles. In addition, bacteria from the genus Agrobacterium can be used to transform plant cells. Below are descriptions of representative techniques for the transformation of both dicotyledonous and monocotyledonous plants. 1. Transformation of Dicotyledonous Transformation techniques for dicotyledons are well known in the art, and include techniques based on Agrobacterium, and techniques that do not require Agrobacterium. The techniques without Agrobacterium involve the recovery of the exogenous genetic material directly by the protoplasts or the cells. This can be done by means of PEG-mediated recovery or electroporation, mediated delivery by particle bombardment, or microinjection. Examples of these techniques are described by Paszkowski et al., EMBO J. 2.: 2717-2722 81984), Potrykus et al., Mol. Gen. Genet. 199: 169-177 (1985), Reich et al., Biotechnology 4 .: 1001-1004 (1986) and Klein et al., Nature 327: 70-73 (1987). In each case, the transformed cells are regenerated to whole plants, using conventional techniques known in the art. The transformation mediated by Agrobacterium is a preferred technique for the transformation of dicotyledons, due to its high transformation efficiency, and its wide utility with many different species. Transformation with Agrobacterium usually involves the transfer of the binary vector carrying the foreign DNA of interest (eg, pCIB200 or pCIB2001) to an appropriate Agrobacterium strain, which may depend on the complement of vir genes carried by the Agrobacterium host strain, either on a co-resident Ti plasmid, or chromosomally (e.g., strain CIB542 for pCIB200 and pCIB2001 (Uknes et al., Plant Cell 5: 159-169 (1993).) The transfer of the recombinant binary vector to Agrobacterium is performed by a triparental coupling method, using E. coli carrying the recombinant binary vector, an auxiliary E. coli strain carrying a plasmid, such as pRK2013, and which can mobilize the recombinant binary vector towards the white Agrobacterium strain In an alternative way, the recombinant binary vector can be transferred to Agrobacterium by DNA transformation (Hófgen and Willmitzer, Nucí. ds. Res., 16 .: 9877 (1988)). Transformation of the white plant species by recombinant Agrobacterium usually involves co-cultivation of Agrobacterium with plant explants, and follows protocols well known in the art. The transformed tissue is regenerated on a selectable medium that carries the antibiotic or herbicide resistance marker present between the T-DNA boundaries of the binary plasmid. Another approach to transforming plant cells with a gene involves propelling inert or biologically active particles into the tissues and cells of the plant. This technique is disclosed in U.S. Patent Nos. 4,945,050, 5,036,006, and 5,100,792, all to Sanford et al. In general, this method involved propelling inert or biologically active particles to the cells under conditions effective to penetrate the outer surface of the cell, and provide incorporation thereinto. When inert particles are used, the vector can be introduced into the cell by coating the particles with the vector containing the desired gene. In an alternative way, the target cell can be surrounded by the vector, such that the vector is brought into the cell by means of the particle. Biologically active particles (eg, dried yeast cells, dried bacteria, or a bacteriophage, each containing DNA to be introduced), can also be propelled into the tissue of the plant cell. 2. Monocotyledon Transformation The transformation of most monocotyledonous species has also become a routine. Preferred techniques include direct gene transfer into the protoplasts using techniques with PEG or electroporation, and bombardment of particles into the callus tissue. Transformations can be undertaken with a single species of DNA, or with multiple species of DNA (i.e., cotransformation), and both techniques are suitable for use with this invention. The co-transformation can have the advantage of avoiding a complete construction of the vector, and of generating transgenic plants with unlinked sites for the gene of interest and the selectable marker, making possible the removal of the selectable marker in the following generations, if this is considered desirable. Nevertheless, a drawback of the use of co-transformation, is the frequency less than 100 percent with which separate species of DNA are integrated into the genome (Schocher et al., biotechnology 4: 1093-1096 (1986)). Patent Applications Numbers EP 0,292,435, EP 0,392,225, and WO 93/07278, describe techniques for the preparation of callus and protoplasts from an elite inbred line of maize, transformation of protoplasts using PEG or electrophoresis, and regeneration of corn plants from transformed protoplasts. Gordon-Kamm et al. (Plant Cell 2: 603-618 (1990)) and Fromm et al. (Biotechnology 8: 833-839 (1990)) have published techniques for the transformation of the corn line derived from A188, using bombardment of particles. In addition, International Publication Number WO 93/07278, and Koziel et al. (Biotechnology 11: 194-200 (1993)) describe techniques for the transformation of elite inbred corn lines by particle bombardment. This technique uses immature maize embryos 1.5 to 2.5 millimeters long, separated from a corn cob 14 to 15 days after pollination, and a PDS-100OO Biolistics device for bombardment. Rice transformation can also be undertaken through direct gene transfer techniques, using protoplasts or particle bombardment. Transformation mediated by the protoplast has been described for the Japanese types and the Indica types (Zhang et al., Plant Cell Rep. 7: 379-384 (1988); Shimamoto et al., Nature 338: 274-277 (1989); Datta; and collaborators, Biotechnology 2.: 736-740 (1990)). Both types can also be transformed in a routine manner using particle bombardment (Christou et al., Biotechnology 2 .: 957-962 (1991)). Patent Application Number EP 0,332,581 describes techniques for the generation, transformation, and regeneration of Pooideae protoplasts. These techniques allow the transformation of Dactylis and wheat. In addition, wheat transformation has been described by Vasil et al. (Biotechnology 1Q_: 667-674 (1992)) using particle bombardment of long-term regenerable C-type callus cells, and also by Vasil et al. (Biotechnology 11: 1553-1558 (1993)) and Weeks et al. (Plant Physiol 102: 1077-1084 (1993)), using bombardment with immature embryo particles and callus derived from immature embryos. However, a preferred technique for wheat transformation involves the transformation of wheat by bombardment with immature embryo particles, and includes either a high pass in sucrose, or high in maltose, before the delivery of the gene. Before bombardment, any number of embryos (0.75 to 1 millimeter in length) are coated on an MS medium with 3 percent sucrose (Murashiga and Skoog, Physiologia Plantarum 15 .: 473-497 81962), and 3 milligrams / liter of 2, 4-D for the induction of somatic embryos, which is allowed to proceed in the dark. On the day chosen for the bombardment, embryos are removed from the induction medium, and placed on the osmotic (i.e., induction medium with added sucrose or maltose at the desired concentration, usually 15 percent). The embryos are allowed to plasmolize for 2 to 3 hours, and then they are bombarded. It is typical 20 embryos per white plate, although it is not critical. A plasmid carrying the appropriate gene (such as pCIB3064 or PSG35) is precipitated onto gold particles in micron size using conventional procedures. Each embryo plate is fired with the DuPont Biolis-ticsR helium device, using a burst pressure of approximately 70 kg / cm2, using a standard 80 mesh. After the bombardment, the embryos are placed back in the dark to recover for approximately 24 hours (still on the osmotic). After 24 hours, the osmotic embryos are removed, and placed back on the induction medium, where they remain for approximately 1 month before regeneration. Approximately 1 month later, the embryo explants with embryogenic callus in development are transferred to the regeneration medium (MS + 1 milligram / liter of NAA, 5 milligrams / liter of GA), which also contains the appropriate selection agent (10). milligrams / liter of coarse in the case of pCIB3064, and 2 milligrams / liter of methotrexate in the case of pSOG35).

After about 1 month, the developed shoots are transferred to larger sterile containers known as "GA7s", which contain MS at a medium concentration, 2 percent sucrose, and the same concentration of selection agent. More recently, the transformation of monocotyledons using Agrobacterium has been described. See Publication International Number WO 94/00977 and U.S. Patent Number 5,591,616, both of which are incorporated herein by reference.

Reproduction Immunomodulated plants obtained by transformation with an acquired systemic resistance gene, such as a form of the NIM1 gene, can be any of a variety of plant species, including those of monocotyledons and dicots; however, the immunomodulated plants used in the method of the invention are preferably selected from the list of agronomically important target crops stipulated above. The expression of the selected form of NIM1 gene, in combination with other characteristics important for production and quality, can be incorporated into plant lines through reproduction. The approach and techniques of reproduction are known in the subject matter. See, for example, Welsh J. R., Fundamentals of Plant Genetics and Breeding, John Wiley and Sons, NY (1981); Crop Breeding, Wood D. R. (ed.) American Society of Agronomy Madison, Wisconsin (1983); May O., The Theory of Plant Breeding, second edition, Clarendon Press, Oxford (1987); Singh, D.P., Breeding for Resistance to Diseases and Insect Pests, Springer-Verlag, NY (1986); and Wricke and Weber, Quantitatíve Genetics and Selection Plant Breeding, Walter de Gruyter and Co. , Berlin (1986). The genetic properties designed in the seeds and transgenic plants described above, are transmitted by sexual reproduction or vegetative growth, and therefore, can be maintained and propagated in descending plants. In general, this maintenance and propagation makes use of known agricultural methods developed to suit specific purposes such as tillage, sowing, or harvesting. Specialized processes, such as hydroponics or greenhouse technologies, can also be applied. As the growing crop is vulnerable to attack and damage caused by insects or infections, as well as competition for herbal plants, measures are taken to control weeds, plant diseases, insects, nematodes, and other adverse conditions. to improve performance. These include mechanical measures, such as tillage of the soil or removal of infected herbs and plants, as well as the application of agrochemicals, such as herbicides, fungicides, gametocides, nematocides, growth regulators, ripening agents, and insecticides. Furthermore, the suitable genetic properties of the transgenic plants and seeds according to the invention can be used in the reproduction of plants, which has as its objective the development of plants with better properties, such as tolerance to pests, to the herbicides, or to the tension, better nutritional value, greater yield, or better structure that causes less loss of lodging or dislocation. The different reproduction steps are characterized by a well defined human intervention, such as selection of the lines to be crossed, direction of the pollination of the parental lines, or selection of the appropriate descendant plants. Depending on the desired properties, different reproduction measures are taken. Relevant techniques are well known in the art, and include, but are not limited to, hybridization, inbred reproduction, backcrossed reproduction, multiple line reproduction, variety blending, interspecific hybridization, aneuploid techniques, and so on. Hybridization techniques also include the sterilization of plants to produce sterile male or female plants, by mechanical, chemical, or biochemical means. The cross-pollination of a sterile male plant with pollen from a different line, ensures that the genome of the male sterile but fertile female plant uniformly obtains the properties of both parental lines. Therefore, seeds and transgenic plants according to the invention, can be used for the reproduction of better lines of plants that, for example, increase the effectiveness of conventional methods, such as treatment with herbicide or pesticide, or allow to eliminate these methods due to its modified genetic properties. Alternatively, new crops with better stress tolerance can be obtained, which, due to their optimized genetic "equipment", produce a harvested product of better quality than the products that could not tolerate comparable adverse development conditions. In the production of seeds, the quality of the germination and the uniformity of the seeds are essential characteristics of the product, whereas the quality of the germination and the uniformity of the seeds harvested and sold by the farmer, are not important. Since it is difficult to keep a crop free of other crop and herbal seeds, to control seed diseases, and to produce seeds with good germination, the seed producers, who have experience in the cultivation, conditioning,. and marketing of pure seeds, have developed very extensive and well-defined seed production practices. Therefore, it is common practice for the farmer to buy certified seeds that meet specific quality standards, instead of using the seeds harvested from his own crop. The propagation material to be used as seeds is customarily treated with a protective coating comprising herbicides, insecticides, fungicides, bactericides, nematocides, molluscicides, or mixtures thereof. The customary protective coatings comprise compounds such as captan, carboxin, thiram (TMTDR), metalaxyl (Apron), and pirimiphos-methyl (Actellic). If desired, these compounds are formulated together with other vehicles, surfactants, or application-promoting auxiliaries employed by custom in the art of formulation, to provide protection against damage caused by bacterial, fungal, or animal pests. The protective coatings can be applied by impregnating the propagation material with a liquid formulation, or by coating it with a combined wet or dry formulation. Other methods of application are also possible, such as the treatment directed towards the buds or towards the fruit. It is a further aspect of the present invention to provide new agricultural methods, such as the methods exemplified above, which are characterized by the use of transgenic plants, transgenic plant material, or transgenic seeds in accordance with the present invention. The seeds may be provided in a bag, container, or container comprised of a suitable packaging material, the bag or container being capable of being closed to contain seeds. The bag, container, or container can be designed either for storage of the seeds in the short term or in the long term, or both. Examples of a suitable packaging material include paper, such as paper delays, rigid or flexible plastic, or other polymeric material, glass or metal. Desirably, the bag, container, or container is comprised of a plurality of layers of packaging materials, of the same type or of a different type. In one embodiment, the bag, container, or container is provided to exclude or limit water and moisture from contact with the seeds. In one example, the bag, container, or container is sealed, for example sealed by heat, to prevent water or moisture from entering. In another embodiment, water absorbing materials are placed between or adjacent to the layers of the packaging material. In yet another embodiment, the bag, container, or container, or the packaging material of which it is comprised, is treated to limit, suppress, or prevent disease, contamination, or other adverse effects of storage or transport of seeds. An example of this treatment is sterilization, for example by chemical methods, or by exposure to radiation. The present invention comprises a commercial bag comprising seeds of a transgenic plant comprising a form of a NIM1 gene, or a NIM1 protein that is expressed in that plant transformed into higher levels than in a wild type plant, together with a vehicle suitable, along with label instructions for use, to confer resistance to broad-spectrum diseases to plants.

Application of a Microbicide to Immunomodulated Plants As described herein, the method of the invention for protecting plants involves two steps: first, activating the path of acquired systemic resistance, to provide an immunomodulated plant, and second, applying a microbicide to these immunomodulated plants, to obtain synergistically enhanced disease resistance.

A. Conventional Microbicides According to the method of the present invention, any commercial or conventional microbicide can be applied to immunomodulated plants, through any of the three routes described above. Examples of suitable microbicides include, but are not limited to, the following fungicides: 4- [3- (4-chlorophenyl) -3- (3,4-dimethoxyphenyl) acryloyl] morpholine ("dimetomorph"), (reference: C. Tomlin (editor): The Pesticide Manual, 10th edition, Farnham, United Kingdom, 1994, pages 351-352); 5-methyl-l, 2,4-triazole [3,4-b] [1,3] benzothiazole ("tricicla-zol"), (reference: C. Tomlin (ed.): The Pesticide Manual, 10th edition, Farnham , United Kingdom, 1994, pages 1017-1018); 3-Allyloxy-1,2-benzothiazole 1,1-dioxide ("probonazole"), (reference: C. Tomlin (editor): The Pesticide Manual, 10th edition, Farnham, United Kingdom, 1994, pages 831-832); ai- [2- (4-chlorophenyl) ethyl] - - - (1,1-dimethylethyl) -1H-1,2,4-triazole-1-ethanol, ("tebuconazole"), (reference: European Patent Number EP-A-40, 345); l - [[3- (2-chlorophenyl) -2- - (4-fluorophenyl) oxirane-2-yl] methyl] -1H-1,2,4-triazole, ("epoxiconazole"), (reference: European Patent Number EP-A-196, 038); μ- (4-chlorophenyl) -μ- (1-cyclopropylethyl) -1H-1, 2,4-triazole-1-ethanol, ("ciproconazole"), (reference: Patent of the United States of America Number US-4, 664, 696); 5- (4-chlorobenzyl) -2,2-dimethyl-1- (1H-1,2,4-triazol-1-ylmethyl) -cyclopentanol, ("metconazole"), (reference: European Patent EP-A-267,778); ether2- (2,4-dichlorophenyl) -3- (1H-1,2,4-triazol-1-yl) -propyl-1,2,2,2-tetrafluoroethyl ("tetraconazole"), (reference: European Patent Number EP-A-234, 242); (E) -2-. { 2- [6- (2-cyanophen-xi) pyrimidin-4-yloxy] phenyl} -3-methoxyacrylate ("ICI A 5504", "azoxyestrobin"), (reference: European Patent Number EP-A-382, 375); (E) -2-methoxy-imino-2- [a- (o-tolyloxy) -o-tolyl] methyl acetate, ("BAS 490 F", "cresoxim-methyl"), (reference: Patent European Number EP-A-400, 417), * acetamide 2- (2-phenoxyphenyl) - (E) -2-methoxy-imino-N-methyl, (reference: European Patent Number EP-A-398, 692); acetamide [2- (2, 5-dimethylphenoxymethyl) -phenyl] - (E) -2-methoxy-imino-N-methyl, (reference: European Patent Number EP-A-398, 692); (1R, 3S / 1S, 3R) -2, 2-dichloro- -N- [(R) -1- (4-chlorophenyl) ethyl] -l-ethyl-3-methylcyclopropanecarboxamide, ("KTU 3616") ( reference: European Patent Number EP-A-341, 475); Manganese polymer complex of ethylenebis (dithiocarbamate) -zinc ("mancozeb"), (reference: United States Patent of North American Number US 2,974,156); 1- [2- (2,4-dichlorophenyl) -4-propyl-1,3-dioxolan-2-methylmethyl] -1 H-1, 2, 4-triazole, ("propicone-zol"), ( Reference: British Patent Number GB-1522657); l-. { 2- [2-chloro-4- (4-chlorophenoxy) phenyl] -4-methyl-l, 3-dioxolan-2-ylmethyl) -1 H-1,2,4-triazole ("difenoconazole") "), (reference: Patent British Number GB-209860); 1- [2- (2,4-dichlorophenyl) pentyl-1H-1,2,4-triazole ("penconazole"), (reference: British Patent Number GB-1589852); cis-4- [3- (4-tertiary butyl-phenyl) -2-methylpropyl] -2,6-dimethylmorpholine, ("phenpropimorf"), (German Patent No. DE 2752135); 1- [3- (4-tertiary butyl-phenyl) -2-methylpropyl] -piperidine, ("fenpropidine"), (reference: German Patent Number DE 2752135); 4-cyclopropyl-6-methyl-N-phenyl-2-pyrimidinamine ("ciprodinil"), (reference: European Patent Number EP-A-310, 550); (RS) -N- (2,6-dimethylphenyl-N- (methoxyacetyl) -alanine ("metalaxyl") methyl ester, (reference: British Patent Number GB-1500581); methyl ester of (R) -N- (2,6-dimethylphenyl-N- (methoxyacetyl) -alanine ("R-metalaxyl"), (reference: British Patent Number GB-1500581); 1,2,5,6 -tetrahydro-4H-pyrrolo [3, 2, 1-ij] quinolin-4-one ("pyroquinone"), (reference: British Patent Number GB-1394373); ethyl acid phosphonate ("fosetyl"), (reference : C. Tomlin (editor): The Pesticide Manual 10th edition, Farnhan, United Kingdom, 1994, pages 530-532), and copper hydroxide (reference: C. Tomlin (editor): The Pesticide Manual 10th edition, Farnhan, United Kingdom, 1994, pages 229-230). The selected microbicide is preferably applied to the immunomodulated plants to be protected, in the form of a composition with other vehicles, surfactants, or other application-promoting auxiliaries customarily employed in the technology of the formulation. Suitable carriers and auxiliaries can be solid or liquid, and are the substances ordinarily employed in the technology of the formulation, for example, natural or regenerated mineral substances, solvents, dispersants, wetting agents, viscosifiers, thickeners, binders, or fertilizers. A preferred method for applying a microbicidal composition is the application to the parts of the plants above the ground, especially to the leaves (foliar application). The frequency and concentration of application depend on the biological and climatic life conditions of the pathogen. However, the microbicide can also penetrate the plant through the roots through the soil, or through water (systemic action), if the place of the plant is impregnated with a liquid formulation (for example, in the rice cultivation), or if the microbicide is introduced into a solid form in the soil, for example, in the form of granules (application to the soil). In order to treat the seeds, the microbicide can also be applied to the seeds (coating), either by impregnation of the tubers or grains with a liquid formulation of the microbicide, or by coating them with a wet or dry formulation already combined. In addition, in special cases, other methods of application to the plants are possible, for example, the treatment directed towards the buds or towards the bunches of fruit. The microbicide can be used in an unmodified form, or preferably, together with auxiliaries conventionally employed in the technology of the formulation, and therefore, it is formulated in a known manner, for example, in emulsifiable concentrates, coatable pastes, solutions directly sprayable or dilutable, dilute emulsions, wettable powders, soluble powders, dry powders, granules, or by encapsulation, for example, in polymeric substances. As with the nature of the compositions, the methods of application, such as spraying, atomizing, dusting, scattering, coating, or watering, are selected in accordance with the intended objectives and the prevailing circumstances. Convenient application concentrations of the microbicide are usually from 50 grams to 2 kilograms of active ingredient per hectare, preferably from 100 to 1,000 grams of active ingredient per hectare, especially from 150 grams to 700 grams of active ingredient per hectare. In the case of seed treatment, the application concentrations are from 0.5 grams to 1,000 grams, preferably from 5 grams to 100 grams of active ingredient per 100 kilograms of seeds. The formulations are prepared in a known manner, for example by homogeneous mixing and / or grinding the microbicide with extenders, for example solvents, solid carriers, and where appropriate, surface-active compounds (surfactants). Suitable solvents are: aromatic hydrocarbons, preferably fractions containing from 8 to 12 carbon atoms, for example mixtures of substituted xylenes or naphthalenes, phthalates, such as dibutyl phthalate or dioctyl phthalate, aliphatic hydrocarbons, such as cyclohexane or paraffins, alcohols and glycols and their ethers and esters, such as ethanol, ethylene glycol, ethylene glycol monomethyl or monoethyl ether, ketones, such as cyclohexanone, strongly polar solvents, such as 2-pyrrolidone N-methyl, dimethyl sulfoxide , or dimethyl formamide, as well as vegetable oils, or epoxidized vegetable oils, such as coconut oil or epoxidized soybean oil; or water. The solid carriers used, for example, for dry powders and dispersible powders, are normally natural mineral fillers, such as calcite, talc, kaolin, montmorillonite, or attapulgite. In order to improve physical properties, it is also possible to add highly dispersed silicic acid or highly dispersed absorbent polymers. Suitable granulated adsorbent carriers are porous types, for example pumice, broken septum, sepiolite, or bentonite, and suitable non-sorbent carriers are, for example, calcite or sand. In addition, a large number of previously granulated materials of an inorganic or organic nature can be used, for example especially dolomite or pulverized plant residues. Depending on the nature of the microbicide, suitable surface active compounds are nonionic, cationic, and / or anionic surfactants having good emulsifying, dispersing, and wetting properties. It will also be understood that the term "surfactants" comprises mixtures of surfactants. Particularly suitable promoter auxiliaries are also natural or synthetic phospholipids of the cephalin and lecithin series, for example phosphatidylethanolamine, phosphatidyl serine, phosphatidyl glycerol, and lysolecithin. The agrochemical compositions generally comprise from 0.1 to 99 percent, preferably from 0.1 to 95 percent active microbicidal ingredient, from 99.9 to 1 percent, preferably from 99.9 to 5 percent of a solid or liquid auxiliary, and from 0 to 25 percent, preferably 0.1 to 25 percent of a surfactant. Although the commercial products of preference will be formulated as concentrates, the end user will usually employ diluted formulations.

B. Plant Activating Microbicides If applied to immunomodulated plants obtained through the second or third route described above (selective breeding or genetic engineering), the microbicide may alternatively be a chemical inducer of acquired systemic resistance (plant activating microbicide), such as a benzothiadiazole compound, an isonicotinic acid compound, or a salicylic acid compound, which are described in U.S. Patent Nos. 5,523,311 and 5,614,395. Accordingly, two methods of immunomodulation are used concurrently. By applying plant-activating microbicides to the immunomodulated plants obtained through a selective breeding route or a genetic engineering route, an "extra immunomodulation" results, and a synergistically improved disease resistance is achieved. As described below, transgenic immunomodulated plants that over-express NIM1 responded much faster at much lower doses of BTH, as shown by the expression of the PR-1 gene and P. parasitic resistance, than the plants of wild type. See Example 35 and the Northern blots of Figure 3. Synergistically enhanced disease resistance in NIM1 overexpressors can be achieved with only one application of 10 μM BTH, a concentration normally insufficient for any efficacy. The normally phytotoxic or otherwise undesirable concentrations of chemical inducers of acquired systemic resistance can be avoided, taking advantage of this synergism. In addition, one can take advantage of the alteration of the time course of the activation of acquired systemic resistance that occurs when chemical products that induce acquired systemic resistance are applied to immunomodulated plants, such as NIM1 overexpressors. In addition, economic grains can be realized as a result of the lower amount of acquired systemic resistance inducing chemicals required to provide a given level of protection to the plants.

C. Conventional Microbicides in Conjunction with Plant Activating Microbicides. For even greater disease resistance, both a conventional microbicide and a plant-activating microbicide can be applied to immunomodulated plants obtained through either a selective breeding route or a genetic engineering route. This results in an even higher level of synergistic resistance to diseases, compared to the level of disease resistance obtained through immunomodulation alone, through immunomodulation plus only one type of microbicide, or through the simultaneous application of both types of microbicides (conventional and activator of plants). See, for example, Table 35 of Example 19.

Disease Resistance Evaluation The evaluation of resistance to diseases is carried out by methods known in the field. See Uknes et al., (1993), Molecular Plant Microbe Interactions 6: 680-685; Gorlach et al., (1996), Plant Cell 8 .: 629-643; Alexander et al., Proc. Nati Acad. Sci. USA 90 .: 7327-7331 (1993). For example, several representative tests of disease resistance are described below.

A. Phytophthora parasitic Resistance Test (Black stem) Tests are carried out to determine resistance to Phytophthora parasitica, the causative organism of the black stem, in 6-week-old plants grown as described in Alexander et al., Proc. Nati Acad. Sci. USA 90: 7327-7331 (1993). The plants are watered, left to drain well, and then inoculated by applying 10 milliliters of a sporangia suspension (300 sporangia / milliliter) to the soil. The inoculated plants are kept in a greenhouse maintained at 23-25 ° C of diurnal temperature, and 20-22 ° C of nocturnal temperature. The wilt index used for the trial is as follows: 0 = no symptoms; 1 = without symptoms; 1 = some sign of wilting, with reduced turgor; 2 = clear symptoms of wilting, but there is no putrefaction or atrophy, * '3 = clear symptoms of wilting with atrophy, but there is no apparent rot of the stem; 4 = severe wilting, with visible rotting of the stem and some damage to the root system; 5 = as for 4, but the plants are close to death or dead, and with a severe reduction of the root system. All trials are scored blind on plants arranged in a randomized design.

B. Pseudomonas syringae Resistance Test Pseudomonas syringae, variety tabaci, strain # 551, is injected into the two lower leaves of several plants from 6 to 7 weeks of age, in a concentration of 10 or 3 x 10 per milliliter in H20. Six individual plants are evaluated at each point of time. Plants infected with Pseudomonas tabaci are evaluated on a disease severity scale of 5 points, 5 = 100 percent dead tissue, 0 = no symptoms. A T-test (LSD) is conducted on the evaluations for each day, and the groupings are indicated after the average disease assessment value. The values followed by the same letter on that day of the evaluation are not significantly different in a statistical way.

C. Cercospora nicotianae Resistance Test A suspension of Cercospora nicotianae spores (ATCC # 18366) (100,000-150,000 spores per milliliter) is sprayed to the imminent spill on the surface of the leaves. The plants are kept at 100 percent humidity for 5 days. Afterwards, the plants are nebulized with water 5 to 10 times a day. Six individual plants are evaluated at each point of time. The Cercospora nicotianae is evaluated on a percentage of the leaf area that shows the basis of the symptoms of the disease. A T-test (LSD) is conducted on the evaluations for each day, and the groupings are indicated after the value of the Average disease assessment. The values followed by the same letter on that day of the evaluation are not significantly different in a statistical way.

D. Parasitic Peronospora Resistance Test Tests are carried out to determine resistance to parasitic Peronospora in plants, as described in Uknes et al. (1993). The plants are inoculated with a combative isolate of P. parasí tica, spraying with a suspension of conidia (approximately 5 x 10 spores per milliliter). The inoculated plants are incubated under humid conditions at 17 ° C in a culture chamber, with a cycle of 14 hours of day / 10 hours of night. The plants are examined 3-14 days, preferably 7-12 days after inoculation, to determine the presence of conidiophores. In addition, several plants of each treatment are randomly selected, and stained with lactophenol-trypan blue (Keogh et al., Trans Br. Mycol. Soc. 74: 329-333 (1980)) for microscopic examination.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a sequence alignment of the sequence of the NIM1 protein with mouse I ?, Bc, rat, and pig. The vertical bars (I) above the sequences indicate the identity of amino acids between the sequences of NIM1 and I? B (matrix rating equal to 1.5); the double points (:) above the sequences indicate a similarity rating > 0.5; single points (.) above the sequences indicate a similarity rating < 0.5, but > 0.0; and a rating < 0.0 indicates that there is no similarity and there are no indications above the sequences (See the Examples). The locations of the ankyrin domains of mammalian I? Ba were identified according to de Martin et al., Gene 152, 253-255 (1995). The points inside a sequence indicate gaps between the NIM1 and I? B proteins. The five ankyrin repeats in IKBOÍ are indicated by dotted lines below the sequence. The amino acids are numbered in relation to the NIM1 protein with holes introduced where appropriate. Signs of plus (+) are placed on top of the sequences every 10 amino acids.

Figure 2 is a comparison of amino acid sequences of the NIM1 protein regions (the numbers correspond to the positions of the amino acids in SEQ ID NO: 2), and the EST rice protein products (SEQ ID NOs: 17-24). Figure 3 presents the results of the Northern analysis showing the time course of expression of the PR-1 gene in the wild type, and lines overexpressing NIM1, followed by treatment with water or BTH. RNA was prepared from the treated plants, and analyzed as described in the Examples. "Ws" is the Ws ecotype of wild type Arabidopsis thaliana. "3A", "5B", "6E", and "7C" are the lines of plants that overexpress individual NIM1 produced according to Example 21. "0 BTH" is the water treatment; "10 BTH" is the treatment with BTH 10 μM; "100 BTH" is the 100 μM BTH treatment. "0" is the zero day of the control samples "1", "3", and "5" are the samples on days 1, 3, and 5.

BRIEF DESCRIPTION OF THE SEQUENCES IN THE LIST OF SEQUENCES SEQ ID NO: l is a genomic sequence of 5655 base pairs comprising the coding region of the NIM1 gene of Arabidopsis thaliana wild type. SEQ ID NO: 2: is the amino acid sequence of the wild-type Arabidopsis thaliana NIM1 protein, encoded by the coding region of SEQ ID NO: 1.

SEQ ID NO: 3 is the amino acid sequence of mouse iKBa from Figure 1. SEQ ID NO: is the amino acid sequence of rat iKBa from Figure 1. SEQ ID NO: 5 is the sequence of amino acids of pig Iβ Ba from Figure 1. SEQ ID NO: 6 is the cDNA sequence of the NJM2 gene of Arabidopsis thaliana. SEQ ID? Os: 7 and 8 are the coding sequence of the AD? and the encoded amino acid sequence, respectively, of a negative-dominant form of the? IM1 protein, which has alanine residues in place of serine residues at amino acid positions 55 and 59. SEQ ID? Os: 9 and 10 are the coding sequence of the AD ?, and the encoded amino acid sequence, respectively, of a negative-dominant form of the? IM1 protein, which has a? -terminal deletion. SEQ ID: Os: 11 and 12 are the coding sequence of the AD ?, and the encoded amino acid sequence, respectively, of a negative-dominant form of the? IM1 protein, which has a C-terminal deletion. SEQ ID: Os: 13 and 14 are the coding sequence of the AD ?, and the encoded .amino acid sequence, respectively, of an altered form of the NIM1 gene, which has deletions of both? -terminal and C-terminal amino acids.

SEQ ID NOs: 15 and 16 are the coding sequence of the DNA, and the encoded amino acid sequence, respectively, of the ankyrin domain of NIM1. SEQ ID NO: 17 is the amino acid sequence of Rice-1, 33-155, of Figure 2. SEQ ID NO: 18 is the amino acid sequence of Rice-1, 215-328 of Figure 2. SEQ ID NO : 19 is the amino acid sequence of Rice-2, 33-155 of Figure 2. SEQ ID NO: 20 is the amino acid sequence of Rice-2, 208-288 of Figure 2. SEQ ID NO: 21 is the amino acid sequence of Rice-3, 33-155 of Figure 2. SEQ ID NO: 22 is the amino acid sequence of Rice-3, 208-288 of Figure 2. SEQ ID NO: 23 is the amino acid sequence of Rice-4, 33-155 of Figure 2. SEQ ID NO: 24 is the amino acid sequence of Rice-4, 215-271 of Figure 2. SEQ ID NOs: 25 to 32 are oligonucleotide primers.

DEFINITIONS The following definitions will assist in the understanding of the present invention: Plant Cell: the structural and physiological unit of plants, consisting of a protoplast and the cell wall. The term "plant cell" refers to any cell that is part of, or is derived from, a plant. Some examples of cells include differentiated cells that are part of a living plant; Differentiated cells in culture; undifferentiated cells in culture; cells of undifferentiated tissue, such as callus or tumors; Differentiated cells of seeds, embryos, propagules, and pollen. Plant tissue: a group of plant cells organized into a structural and functional unit. Any tissue of a plant in the plant or in culture is included. This term includes, but is not limited to, whole plants, plant organs, plant seeds, tissue culture, and any groups of plant cells organized into structural and / or functional units. The use of this term, in conjunction with, or in the absence of, any specific type of plant tissue, as mentioned above or otherwise encompassed by this definition, is not intended to be exclusive of any other type of plant tissue. Protoplast: a plant cell without a cell wall. Descendant Plant: a sexually or asexually derived future generation plant, which includes, but is not limited to, progeny plants. Transgenic plant: a plant that has recombinant DNA stably incorporated in its genome. Recombinant DNA: any DNA molecule formed by joining segments of DNA from different sources, and produced using recombinant DNA technology. Recombinant DNA technology: technology that produces Recombinant DNA in vi tro and transfers the recombinant DNA to the cells, where it can be expressed or propagated (See Concise Dictionary of Biomedicine and Molecular Biology, Ed. Juo, CRC Press, Boca Raton (1996)), for example, the transfer of DNA to a protoplast or cell in different forms, including, for example: (1) naked DNA in circular, linear, or super-straight, (2) DNA contained in nucleosomes or chromosomes or nuclei or parts thereof, ( 3) DNA complexed or associated with other molecules, (4) DNA encased in liposomes, spheroplasts, cells or protoplasts, or (5) DNA transferred from other organisms different from the host organism (eg, Agrobacterium tumefiaciens). These and other different methods for introducing recombinant DNA into cells are known in the art, and can be used to produce the transgenic cells or the transgenic plants of the present invention. Recombinant DNA technology also includes the methods of homologous recombination described in Treco et al., International Publication Number WO 94/12650, and Treco et al., International Publication Number WO 95/31560, which can be applied to increasing peroxidase activity in a monocot. Specifically, regulatory regions (e.g., promoters) can be introduced into the plant genome to increase the expression of endogenous peroxidase. Also included as recombinant DNA technology is the insertion of a peroxidase coding sequence that lacks selected expression signals in a monocotyledon, and assay of the transgenic monocotyledonous plant to determine the increased expression of peroxidase due to the endogenous control sequences in the monocot. This would result in an increase in the number of copies of peroxidase coding sequences inside the plant. The initial insertion of the recombinant DNA into the genome of the R ° plant is not defined as being done by traditional plant breeding methods, but rather by technical methods as described herein. Following the initial insertion, transgenic descendants can be propagated using essentially traditional breeding methods. Chimeric Gene: A DNA molecule that contains at least two heterologous parts, for example the parts derived from previously existing DNA sequences that are not associated in their previously existing states, these sequences having been generated preferably using recombinant DNA technology. Expression Catete: A DNA molecule comprising a promoter and a terminator between which a coding sequence can be inserted. Coding Sequence: A DNA molecule that, when transcribed and translated, results in the formation of a polypeptide or a protein. Gene: A separate chromosomal region comprising a regulatory DNA sequence responsible for the control of expression, i.e., transcription and translation, and a coding sequence that is transcribed and translated to a different polypeptide or protein. acd: mutant plant of accelerated cell death (accelerated = .ell death). AFLP: Amplified Fragment Length Polymorphism. avrRpt2: avirulence gene Rpt2, isolated from Pseudomonas syringae. BAC: Bacterial Artificial Chromosome (Chromosome Artificial Bacterial) BTH: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. CIM: Phenotype of Constitutive Immunity (Constitutive Mmunity) (acquired systemic resistance is activated in a constitutive way). cim: Constitutive immunity mutant plant cM: centimorgans cprl: Constitutive expressor of gene mutant plant EE. Col-O: Arabidopsis, ecotype Columbia ECs: Enzyme combinations Emwa: Peronospora parasitic isolate compatible in the Ws-0 ecotype of Arabidopsis. EMS: ethyl methane sulfonate. INA: acid 2, 6-dichloroisonicotinic. Ler: Arabidopsis, ecotype Landsberg erect Isd: lesion-simulating disease mutant plant nahG: Pseudomonas putida with salicylate hydroxylase that converts salicylic acid to catechol. NahG: Arabidopsis line transformed with the nahG ndr gene. : non-race-specific disease resistance mutant plant nim: mutant plant of non-inducible immunity (non-inducible immunity) NIM1: The wild-type gene, involved in the transduction cascade of the acquired systemic resistance signal. NIM1: Protein encoded by the wild-type NIM1 gene. niml: mutant allele of NIM1, which confers susceptibility to diseases to the plant; it also refers to the mutant plants of Arabidopsis thaliana, which have the niml mutant allele of NI 2? oco: Peronospora parasitic isolate compatible in the Arabidopsis Col-O ecotype ORF: open reading frame PCs: PR primer combinations: Related to pathogenesis SA: salicylic acid SAR: Acquired Systemic Resistance SAR-on: Immunomodulated plants where acquired systemic resistance is activated, which normally exhibits an expression of the acquired systemic resistance gene greater than that of the wild type, and which have a phenotype resistant to diseases. SSLP: Simple Sequence Length Polymorphism. UDS: Universal Phenotype Susceptible to Wela Diseases: Peronospora isolated parasitic compatible in ecotype einingen of Arabidopsis. s-O: Arabidopsis ecotype Issilewskija WT: Wild type YAC: Yeast Artificial Chromosome EXAMPLES The invention is illustrated in more detail by the following detailed procedures, preparations, and examples. The examples are for illustration only, and should not be construed to limit the scope of the present invention. The standard recombinant DNA, and the molecular cloning techniques used herein, are well known in the art, and are described by Sambrook et al., Molecular Cloninq, editors, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989). , and by TJ Silhaby, M.L. Berman, and L.W. Enquist, Experiments with Gene Fusions, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1984) and by Ausubel, F.M. and collaborators, Current Protocols in Molecular Bioloay, published by Greene Publishing Assoc. and Wiley-Interscience (1987). I. Effects of Synergistic Resistance to Diseases Achieved through the Coordinated Application to the Plants of a Chemical Inductor of Acquired Systemic Resistance with a Conventional Microbicide In this set of examples, acquired systemic resistance in plants was induced by the application of a chemical inductor. of acquired systemic resistance, such as a benzothiadiazole. In addition, conventional microbicides were applied to the plants. Then the plants were subjected to disease pressure from different pathogens. The combination of both methods to combat pathogens (chemical + microbicide) produced a resistance to diseases greater than the additive, that is, synergistic. This was determined as the synergy factor (SF), that is, the ratio of the observed effect (0) to the expected effect (E). The expected effect (E) for a given combination of active ingredients can be described by the so-called Colby formula, and can be calculated as follows (Colby, SR, "Calculating synergistic and antagonistic responses of herbicide combination" Weeds, volume 15, pages 20-22 (1967)): ppm = milligrams of active ingredient (= ia) per liter of spray mixture. X =% action caused by the active ingredient I at an application concentration of p ppm active ingredient, Y =% action caused by the active ingredient II, at an application concentration of q ppm of active ingredient, E = expected effect of the active ingredients I + II at an application concentration of p + q ppm of active ingredient (additive action). Colby's formula is: E = X + Y - XY 100 Example 1: Action Against Erysiphe graminis on Barley Residual protective action: Barley plants about 8 centimeters high, were sprayed to the point of drip, with an aqueous spray mixture (maximum 0.02 per percent of active ingredient), and were sprinkled 3 to 4 days later with conidia of the fungus. The infected plants were left in a greenhouse at 22 ° C. The fungus infestation was evaluated in general 10 days after infection. Systemic Action: Barley plants approximately 8 centimeters high, were irrigated with an aqueous spray mixture (maximum 0.002 percent active ingredient, based on the volume of the soil). Care was taken that the spray mixture did not come in contact with the parts of the plants that were above the ground. The plants were sprinkled with conidia of the fungus 3 to 4 days later. The infected plants were left in a greenhouse at 22 ° C. The fungus infestation was evaluated in general 10 days after infection. Table 1 Action against Erysiphe graminis in barley Component I: benzothiadiazole-7-carboxylic acid Component II: metconazole Table 2 Action against Erysiphe graminis in barley Component I: benzothiadiazole-7-carboxylic acid Table 3 Action against Erysiphe graminis in barley Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: metconazole Example 2: Action against Colletotrichum lagenarium in Cucumis sativus L. After a culture period of 10 to 14 days, the cucumber plants were sprayed with a spray mixture prepared from a wettable powder formulation of the test compound. After 3 to 4 days, the plants were infected with a spore suspension (1.0 x 105 spores / milliliter) of the fungus, and incubated for 30 hours with high humidity and at a temperature of 23 ° C. Then incubation was continued with normal humidity and from 22 ° C to 23 ° C. The evaluation of the protective action was made 7 to 10 days after the infection, and was based on the infestation of the fungus. After a culture period of 10 to 14 days, the cucumber plants were treated by applying to the soil a spray mixture prepared from a wettable powder formulation of the test compound. After 3 to 4 days, the plants were infected with a spore suspension (1.5 x 105 spores / milliliter) of the fungus, and incubated for 30 hours with high humidity and at a temperature of 23 ° C. Then the incubation with normal humidity and at 22 ° C was continued. The evaluation of the protective action was made 7 to 10 days after the infection, and was based on the infestation of the fungus. Table 4 Action against Colletotrichum lagenarium in Cucumis sativus L. / foliar application Component I: benzothiadiazole-7-carboxylic acid Component II azoxyestrobin Table 5 Action against Colletotrichum lagenarium in Cucumis sativus L. / application to the earth Component I: benzothiadiazole-7-carboxylic acid Component II: azoxyestrobin * The synergy factor SF can not be calculated Table 6 Action against Colletotrichum lagenarium in Cucumis sativus L. / foliar application Component I: benzothiadiazole-7-carboxylic acid Component II: Cresoxime-methyl Table 7 Action against Colletotrichum lagenarium in Cucumis sativus L. / foliar application Component I: benzo [1,2,3] thiadiazole-7-carbothioic acid S-methyl ester Component II: Azoxyestrobin Table 8 Action against Colletotrichum lagenarium in Cucumis sativus L. / application to soil Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid Component II: Azoxyestrobin Example 3: Action Against Cercospora nicotianae in Plants tobacco Tobacco plants (6 weeks old) are sprayed with a formulated solution of the test compound (concentration: maximum 0.02% active ingredient). Four days After treatment, the plants were inoculated with a suspension of sporangia of Cercospora nicotianae (150,000 spores / milliliter), and were kept at high humidity for 4 to 5 days, and then further incubated under a sequence normal day / night. The evaluation of the symptoms in the tests was based on the surface of the leaf infested with the fungus.

Table 9 Action against Cercospora nicotianae in tobacco plants Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid Component II: tebuconazole Table 10 Action against Cercospora nicotianae in tobacco plants Component I: benzo [1,2, 3] thiadiazole-7-carbothioic acid S-methyl ester Component II: ciproconazole Table 11 Action against Cercospora nicotianae in tobacco plants Component I: benzothiadiazole-7-carboxylic acid Component II: phenpropimorf Table 12 Action against Cercospora nicotianae in tobacco plants Component I: benzothiadiazole-7-carboxylic acid Component II: difenoconazole Example 4: Action Against Pyricularia oryzae in rice plants Rice plants of approximately 2 weeks of age were placed together with the soil around the roots in a container filled with a spray mixture (maximum 0.006 percent active ingredient). 96 hours later, the rice plants were infected with a conidia suspension of the fungus. The infestation of the fungus was evaluated after incubating the infected plants for 5 days at a relative humidity of 95 to 100 percent, and at approximately 24 ° C. Table 13 Action against Pyricularia oryzae in rice plants Component I: benzo [1, 2,3] thiadiazole-7-carbothioic acid S-methyl ester Component II: KTU 3616 On a 12-square-meter plot, rice plants were sprayed with a spray mixture prepared with a wettable powder of the active ingredient. The infection was made naturally. For the evaluation, the area of the leaf infested with the fungus was measured 44 days after the application. The following results were obtained. Table 14 Action against Pyricularia oryzae in outdoor rice plants Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid Component II: pyroquilone The rice plants of approximately 2 weeks of age were placed together with the soil around the roots in a container filled with a spray mixture. The fungus infestation was evaluated 36 days later. The infestation of the untreated plants corresponded to an action of 0 percent. Table 15 Action against Pyricularia oryzae in rice plants Component I: benzo [1, 2, 3] thiadiazole-7-carbothioic S-methalic acid ester Component II: tricyclazole Example 5: Action Against Colletotrichum sp. (Anthracnose) and Cercospora sp (Leaf spot) in Chile Effects on crop yield: In a field of approximately 10 square meters (test location: Cikampek, Java, Indonesia), chili plants were sprayed a total of 7 times at intervals of approximately 7 days, with 500 to 700 liters of spray mixture per hectare. Three days after the first spray, the plants were artificially infected with the fungus.

Table 16 Action against Colletotrichum: The evaluation was made evaluating the infestation on the fruits of chili after the fifth spray. Component I: benzo [1, 2, 3] thiadiazole-7-carbothioic acid S-methyl ester Component II: mancozeb Table 17 Action against Cercospora: The evaluation was made evaluating the infestation on the leaves after the sixth spray. Component I: benzo [1, 2, 3] thiadiazole-7-carbothioic acid S-methyl ester Component II: mancozeb Table 18 Action on crop yield: Chilies were harvested after the sixth spray. Component I: benzo [1, 2, 3] thiadiazole-7-carbothioic acid S-methyl ester Component II: mancozeb Example 6: Action against recondite puccinia in wheat. The 7-day-old wheat plants were sprayed to the drop point with a spray mixture prepared from a formulated active ingredient, or a combination of active ingredients. After 4 days, the treated plants were infected with a conidia suspension of the fungus, and the treated plants were subsequently incubated for 2 days at a relative atmospheric humidity of 90 to 100 percent, and at 20 ° C. 10 days after infection, the fungus infestation was evaluated.

Table 19 Action against Puccinia recondite in wheat. Component I: benzo [1, 2, 3] thiadiazole-7-carbothioic acid S-methyl ester Component II: propiconazole Table 20 Action against Puccinia recondite in wheat. Component I: benzothiadiazole-7-carboxylic acid Component II: fenpropidine Example 7: Action against Erysiphe graminis in wheat In field trials (10 square meters), winter wheat was sprayed in the growth phase, with a prepared spray mixture with a wettable powder of the active ingredient.

The infection was made naturally. Ten days after the infection, the fungus infestation was evaluated. The following results were obtained: Table 21 Action against Erysiphe graminis in wheat. Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: propiconazole Table 22 Action against Erysiphe graminis in wheat. Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: ciprodinil Example 8: Action against Mycosphaerella fijiensis in bananas 40 banana plants in a plot of 300 square meters were sprayed at intervals of 17 to 19 days, with a prepared spray mixture with the wettable powder of the active ingredient; in total 6 times. The infection was made naturally. For the evaluation, the leaf infested with the fungus was measured. The following results were obtained: Table 23 Action against Mycosphaerella fij iensis in bananas. Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: propiconazole Example 9: Action against Alternaria solani in tomatoes The tomato plants in a 7 square meter plot were sprayed at 7 day intervals, with a prepared spray mixture with a wettable powder of the active ingredient; altogether nine times. The infection was made naturally. For the evaluation, the leaf infested with the fungus was measured. The following results were obtained: Table 24 Action against Al ternaría solani in open-air tomatoes. Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: ciprodinil Example 10: Action against Phytophthora infestans in tomatoes. Tomato plants, variety "Roter Gnom", were sprayed to the point of dripping with a spray mixture prepared with the active ingredient formulated, or a combination of active ingredients. After 4 days, the treated plants were sprayed with a suspension of sporangia of the fungus, and subsequently incubated in a cabinet for 2 days at 18-20 ° C, and with a relative atmospheric humidity of 90 to 100 percent. 5 days after infection, the fungus infestation was evaluated. The following results were obtained: Table 25 Action against Phytophthora infestans in tomatoes. Component I: S-methyl ester of benzo [1, 2, 3] thiadiazole-7-carbothioic acid. Component II: metalaxyl Table 26 Action against Phytophthora infestans in tomatoes Component I: benzothiadiazole-7-carboxylic acid. Component II: metalaxyl Example 11: Action against Pseudoperonospora cubensis on cucumbers Cucumber plants 16 to 19 days old ("Wisconsin") were sprayed to the drip point with a spray mixture prepared with the active ingredient formulated, or a combination of the ingredient active, or a combination of active ingredients. After 4 days, the treated plants were infected with sporangia of Pseudoperonospora cubenswas (strain 365, Ciba, maximum 5,000 per milliliter), and the treated plants were subsequently incubated for 1 to 2 days at 18-20 ° C, and with a humidity . relative atmospheric from 70 to 90 percent. Ten days after the infection, 1 infestation of the fungus was evaluated, and compared with the infestation in the untreated plants. The following results were obtained: Table 27 Action against Pseudoperonospora cubensis in cucumbers. Component I: benzothiadiazole-7-carboxylic acid. Component II: metalaxyl Example 12: Action against Peronospora tabacina in tobacco plants The tobacco plants (6 weeks old) were sprayed with a formulated solution of the test compound. Four days after the treatment, the plants were inoculated with a suspension of sporangia of the fungus, kept at high humidity for 4 to 5 days, and then further incubated under a normal day / night sequence. The evaluation of the symptoms in the tests was based on the surface of the leaf infested with the fungus. The infestation of the untreated plants corresponded to an action of 0 percent. Table 28 Action against Peronospora tabacina in tobacco plants. Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: dimetomorf Example 13: Action against Peronospora parasitic on Arabidopsis thaliana The fungicides metalaxyl, fosetyl, and copper hydroxide, and the systemic resistance activator acquired S-methyl ester of benzo (1, 2, 3) -thiadiazole-7-carbothioic acid ( BTH), formulated as 25 percent, 80 percent, 70 percent, and 25 percent active ingredient (ai), respectively, with a wettable powder vehicle, were applied as a fine mist to the leaves of plants of 3 weeks of age. The wettable powder was only applied as a control. Three days later, the plants were inoculated with a suspension of conidia of Peronospora parasitica as described in Delaney et al. (1995). The Ws plants were inoculated with the compatible isolate of P. parasí tica Emwa (1-2 x 105 spores / milliliter), * the Col plants were inoculated with the compatible P. isolate. parasitic Noco2 (0.5-1 x 105 spores / milliliter). Following the inoculation, the plants were covered to maintain a high humidity, and were placed in a Percival culture chamber at 17 ° C, with a cycle of 14 hours of day / 10 hours of night (Uknes et al., 1993) . The tissue was harvested 8 days after inoculation. The progress of fungal infection was followed for 12 days, looking under a dissecting microscope to evaluate the development of conidiospores (Delaney et al., (1994), Dietrich, et al., (1994)). Dyeing with lactophenol-trypan blue from the individual leaves was performed to observe the fungal growth inside the leaf tissue. Fungal growth was quantified using a fungal rRNA probe that was obtained by the Polymerase Chain Reaction according to White et al., (1990; PCR Protocols: A guide to Methods and Application, 315-322) using the NS1 primers. and NS2, and EmWa DNA from P. parasí tica as templates. The RNA was purified from the frozen tissue by extraction with phenol / chloroform followed by precipitation with lithium chloride (Lagrimini et al., 1987: PNAS, 84: 7542-7546). The samples (7.5 micrograms) were separated by electrophoresis through formaldehyde-agarose gels, and stained on nylon membranes (Hybond-N +, Amersham) as described by Ausbel et al., (1987). Hybridizations and washing were according to Church and Gilbert (1984, PNAS, 81: 1991-1995). The relative amounts of transcription were determined using a Phosphor Imager (Molecular Dynamics, Sunnyvale, CA) following the manufacturers' instructions. The sample loading was normalized by probing the stains of the separate filter with the constitutively expressed b-tubulin Arabidopsis cDNA. The infestation of the untreated plants corresponded to an inhibition of fungal growth of 0 percent. The following results were obtained: Table 29 Action against Peronospora parasitic NoCo2 on Arabidopsis thaliana (Col-O). Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid.

Table 30 Action against Peronospora parasitic Emwa in Arabidopsis thaliana (Ws). Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid. Component II: metalaxyl Table 31 Action against Peronospora parasitic Emwa in Arabidopsis thaliana (Ws) Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid.

Table 32 Action against Peronospora parasitic Emwa in Arabidopsis thaliana (Ws). Component I: S-methyl ester of benzo [1,2,3] thiadiazole-7-carbothioic acid.

As can be seen in Table 29, the synergistic effects of disease resistance on the wild-type Arabidopsis Col-O plants were demonstrated. No inhibition of fungal growth was observed by the separate application of either 0.01 mM BTH, or 0.0001 grams / liter of metalaxyl to the plants, because these concentrations are usually insufficient to be effective. However, by applying both of these compounds to plants at these normally insufficient concentrations, an inhibition of fungal growth of 40.7 percent was observed, which is clearly a synergistic effect. Tables 30 to 32 show the synergistic effects of disease resistance in wild type Arabidopsis Ws plants. An inhibition of fungal growth was only observed from 20 to 30 percent by applying 0.01 mM BTH to the Ws plants. However, by a simultaneous application of BTH and either metalaxyl, fosetyl, or copper hydroxide to the plants, a synergistic resistance to the diseases was observed. These combined antifungal effects, which result in a decrease in the effective concentration of the fungicide and BTH required for the control of pathogens, allow to reduce the chemical dose necessary to stop the fungal growth, and consequently, mitigate the incidence of leaf damage due to tolerance to chemical products II. Synergistic Effects of Resistance to Diseases Achieved through the Application of Conventional Microbicides and / or Acquired Systemic Resistance Chemical Inductors, to Constitutive Immunity Mutant Plants (CIM) In this set of examples, a high production Northern blot screen was developed to identify mutant plants having high concentrations of PR-1 mRNA during normal growth, with the idea that these mutants also exhibit acquired systemic resistance. A number of mutants have been isolated using this screen, and have been shown to accumulate not only PR-1, but also PR-2 and PR-5 mRNAs (Lawton et al., (1993); Dietrich et al., (1994); and Weymann et al., (1995)). These mutants also have high levels of salicylic acid, and are resistant to infection by pathogens, confirming that this approach can be used to isolate transduction mutants from the acquired systemic resistance signal. Two classes of transduction mutants of the acquired systemic resistance signal have been isolated using this screen. One class has been designated as Isd mutants (Isd = simulating injury disease, simulating disease). This class of mutants is also referred to as "Class I cim", as disclosed in International Publication Number WO 94/16077, the disclosure of which is incorporated herein by reference in its entirety.This Isd class (aka Class I cim ) formed spontaneous lesions on the leaves, accumulated high concentrations of salicylic acid, high levels of mRNA of PR-1, PR-2, and PR-5, and was resistant to fungal and bacterial pathogens (Dietrich et al., 1994; Weymann and collaborators, 1995).

The second class, called cim (cim = constitutive immunity constitutive immunity), is described below, and has all the characteristics of Isd mutants, with the exception of spontaneous lesions. This second class (cim) corresponds to the "Class II cim" mutants discussed in International Publication Number WO 94/16077. The cim3 mutant plant line described below falls into this class cim (class II cim), and is a dominant mutation with a wild-type appearance expressing stable high levels of salicylic acid, mRNA of acquired systemic resistance gene, and has resistance to broad spectrum diseases.

Example 14: Isolation and Characterization of cim Mutants with Expression Constituting the Acquired Systemic Resistance Gene. 1,100 individual plants of Arabidopsis mutated with M2 (EMS) were cultured on Aracon trays (Lehie Seeds, Round Rock, TX), in sets of approximately 100. The plants were cultivated as described in Uknes et al., 1993, supra, giving special attention to avoiding over-irrigation and infection with pathogens. Put briefly, the Metro 360 Mix was saturated with water and autoclaved three times for 70 minutes in lots of 10 liters. The mixture for the putty was completely stirred between each step by autoclaving. The seeds were superficially sterilized in 20% Clorox for 5 minutes, and washed with 7 changes of sterile water before planting. The planted seeds were vernalized for 3 to 4 days, followed by growth in chambers with a cycle of 9 hours of day and 15 hours of night at 22 ° C. When the plants were 3 to 4 weeks old, one or two leaves, weighing 50 to 100 milligrams, were harvested, and the total RNA was isolated using a rapid mini-RNA preparation (Verwoerd et al., (1989) Nuc. Acid Res. 17, 2362). The expression of PR-1 gene was analyzed by Northern Blot analysis (Lagrimini et al., (1987) Proc. Nati, Acad. Sci. USA 84, 7542-7546, Ward et al., 1991). Each set of plants also contained an A. Thaliana Col-O not treated, and a control treated with INA for 2 days (0.25 milligrams / milliliter). All the plants were maintained as described in Weymann et al., (nineteen ninety five) . We identified 80 putative mutants that accumulated high levels of PR-1 mRNA. Following the descendants' test, 5 were selected for further characterization. The putative cis mutants exhibited a high expression of acquired systemic resistance gene in the absence of pathogens or inducer treatment. Proof of the descendants of the putative cim mutants confirmed that the constitutive expression of PR-1 was hereditary. Of the cim mutants, two were further characterized, cim2 and cim3, with the highest and most stable expression of PR-1.

Backcrosses with Columbia used the recessive glabrous trait as a marker for the identification of offspring Fl. Col-gil flower buds were emasculated before the pollen cover, and the pollen of the mutants was applied immediately and the next day. The Fl plants were cultivated in the soil, and the cross plants were identified by the presence of trichomes. Following crosses of cim2 and cim3 with the ecotype Col-O or La-er, a large proportion of Fl plants were identified with a high expression of the acquired systemic resistance gene, suggesting that these traits were dominant. In the case of cim2, some, but not all Fl plants had constitutive expression of the acquired systemic resistance gene. This result would be expected if the cim2 mutant were dominant and were carried as a heterozygote in the mother. An additional genetic test of cim2 showed a continuous variable segregation in generation F2, consistent with incomplete penetration. cim3 showed a segregation of 1: 1 in the Fl generation, over which, two individual Fl plants expressing a high level of PR-1 mRNA crossed themselves to form a F2 population. The segregation of F2, obtained by qualifying the accumulation of PR-1 mRNA, showed 93 F2 plants with high mRNA of PR-1, and 25 F2 plants without significant accumulation of PR-1 mRNA, giving a ratio of 3.7 : 1 (c2 = 1.77; 0.5> P> 0.1), which is consistent with the hypothesis that cim3 is a mutation of a single dominant gene. Subsequent external crosses confirmed that cim3 was inherited as a dominant mutation. For cim3, the original M2 plant identified on the screen, and the M3 population, appeared normal. However, when cim3 plants were self-crossing, some of the best expression lines had low fertility. Following the backcross with Col-gil, plants with normal appearance and fertility, and strong PR-1 expression were obtained. When initially identified, cim3 also appeared slightly dwarf with thin, distorted leaves. However, the F2 plants resulting from a cross with the Col-gil ecotype retained a high expression of the acquired systemic resistance gene, and could not be distinguished from the wild-type plants. This suggested that the distorted leaf dwarf phenotype was caused by an independent mutation that was not associated with the constitutive expression of the acquired systemic resistance gene. The mutant phenotype of cim3 was also observed when the plants were cultured under sterile conditions, confirming that the accumulation of PR-1 mRNA was not caused by a pathogen.

Example 15: Expression of the Acquired Systemic Resistance Gene In addition to PR-1, two other acquired systemic resistance genes, PR-2 and PR-5, are also highly expressed. in cim3. The levels of expression of the acquired systemic resistance gene varied among the descendants, but they were always more than 10 times higher than the untreated control, and similar to the levels obtained immediately after a treatment with INA resistance inducer 0.25 milligrams / milliliter) of wild type plants.

Example 16: Analysis of Salicylic Acid Endogenous concentrations of salicylic acid have been shown to be increased immediately after pathogen-induced necrosis in Arabidopsis (Uknes et al., 1993, supra). Salicylic acid and its conjugate were analyzed with glucose as described in Uknes et al., 1993. Leaf tissue was harvested from 10 plants with cim3 and 10 control plants of 4 weeks of age. The leaves of the individual plants were harvested and analyzed for the expression of the PR-1 gene. The salicylic acid levels of the plants expressing PR-1 were measured. The concentration of free salicylic acid in cim3 was 3.4 times higher than in uninfected wild type Arabidopsis (233 ± 35 versus 69 ± 8 nanograms / gram of fresh weight, respectively). Salicylic acid glucose conjugate (SAG) was 13.1 times higher in cim3 than in uninfected wild type Arabidopsis (4519 ± 473 vs. 344 ± 58 nanograms / gram of fresh weight, respectively). These increased levels of SA and SAG are comparable with the levels that have been reported either for the pathogen-infected tissue or for the cpr mutant.

Example 17: Resistance to Diseases cim3 was evaluated to determine its resistance to parasitic Peronospora (NoCo2), the causal agent of the Arabidopsis plush mold disease. Thirty cim3 (confirmed by the expression of PR-1 RNA), and thirty control plants (Columbia ecotype), each about 4 weeks old, were inoculated with P. parasí tica, as described in Uknes et al. 1992, supra. Seven days later, the plants were analyzed for sporulation, and stained with trypan blue to visualize the fungal structures, as described in Keogh et al., (1980), Trans. Br. Mycol. Soc. 74, 329-333, and in Koch and Slusarenko (1990) Plant Cell 2, 437-445. Wild-type plants (Col-O) support the growth of hyphae, conidia, and oospores, while wild-type plants treated with INA (0.25 milligrams / milliliter), and cim3 plants, showed no fungal growth. The resistance mediated by cim3 is typically seen as a small group of dead cells at the site of infection of the pathogen. This type of resistance is similar to that seen in Isd mutants (Dietrich et al., 1994, supra; Weymann et al., 1995, supra), or in wild-type plants in which acquired systemic resistance has been induced (Uknes et al., 1992, supra). Occasionally intermediate resistance phenotypes were observed, including trailing necrosis upon waking the tip of the hypha in cim3 plants. This carryover necrosis is similar to that found in wild-type plants treated with low doses of SA or INA (Uknes et al., 1992, supra; Uknes et al., 1993, supra). However, sporulation was never observed in cim3 plants, while all control plants showed sporulation. No spontaneous lesions were observed on the leaves of cim3 not inoculated when they were stained with trypan blue. In addition to resistance to fungal P. parasitic pathogen, cim3 was also resistant to infection with the bacterial pathogen Pseudomonas syringae DC3000. Wild-type 6-week-old plants (treatment ±. INA) and cim3 plants were inoculated with a suspension of P. syringae DC3000, and the progress of the disease was monitored by monitoring the growth of bacteria extracted from the infected leaves. through time. The differences in bacterial titers between Col-O, Col-O + INA, and cim3, on day 0 or on day 2, were not statistically significant. However, by day 4, there was a 31-fold decrease in bacterial growth between the wild-type plants and cim3 (P <0.003; Sokal and Rohlf, 1981). The plants were also visually inspected for disease symptoms. . The leaves of wild-type plants were severely chlorotic with symptoms of disease extending well beyond the initial injection site. In contrast, wild-type plants previously treated with INA or cim3 plants were almost without disease symptoms. For this example, cultures of Pseudomonas syringae pv. Tomato strain DC3000 on King B medium (agar or liquid plates) plus rifampin (50 micrograms / milliliter) at 28 ° C (Walen et al., (1991), Plant Cell 3, 49-59). A night culture was diluted and resuspended in 10 mM MgCl 2, to a density of 2-5 x 10 5 cells per milliliter, and injected into Arabidopsis leaves. Injections were made by creating a small hole with a 28 gauge needle half way up the leaf, and then injecting approximately 250 microliters of the diluted bacterial solution with a 1 cubic centimeter syringe. At different points of time, 10 random samples consisting of 3 random sheet perforations were taken from a # 1 rubber driller, of 10 plants of each treatment. The three leaf perforations were placed in an Eppendorf tube with 300 microliters of 10 mM MgCl2., and ground with a pistil. The resulting bacterial suspension was appropriately diluted, and coated on King's B medium plus rifampicin (50 micrograms / milliliter), and cultured for 4 days at 28 ° C. The bacterial colonies were counted, and the data were subjected to the "Student's t" statistical analysis (Sokal and Rohlf (1981), Biometry, 2nd edition, New York: W.H. Freeman and Company). Also for this example, 2,6-dichloroisonicotinic acid (INA) was suspended in sterile distilled water as a 25 percent active ingredient formulated in a wettable powder (0.25 milligrams / milliliter, 325 μM; Kessmann et al. (1994), Annu. Rev. Phytopathol 32, 439-59). All the plants were sprayed with water or INA solutions to the point of imminent overflow.

Example 18: The Role of Salicylic Acid in the Expression of the Acquired Systemic Resistance Gene and in Disease Resistance To investigate the relationship between salicylic acid, the expression of the acquired systemic resistance gene, and the resistance in cim3, crosses with Arabidopsis plants expressing the salicylate hydroxylase (nahG) gene (Delaney et al., 1994). This "NaHG plants" were made by transforming the nahG gene directed by 35S in Arabidopsis, using transformation mediated by Agrobacterium. See, Huang, H. Ma, H. (1992) Plant Mol. Biol. Rep. 10, 372-383, incorporated herein by reference; Gaffney et al. (1993) Science 261, 754-756, incorporated herein by reference; and Delaney et al., (1994) Science 266, 1247-1250, incorporated herein by reference. Col-nahG Arabidopsis carries a dominant kanamycin resistance gene in addition to the dominant nahG gene, so that Col-nahG was used as the pollen donor. The Fl seed was hydrated in water for 30 minutes, and then surface sterilized in 10 percent Clorox, 0.05 percent Tween 20 for 5 minutes, and washed thoroughly in sterile water. The seeds were placed on the germination medium (GM, Murashige and Skoog medium containing 10 grams / liter of sucrose regulated with 0.5 grams / liter of 2- (N-morpholino) ethanesulfonic acid, pH 5.7, with KOH ), containing 25 milligrams / milliliter of kanamycin, to select plants Fl. See Valvekens et al., (1988) Proc. Nati Acad. Sci. USA 85, 5536-5540. Fl plants resistant to kanamycin were transferred to the earth after 18 days. The presence of the nahG gene and the expression of PR-1 were confirmed in all experiments by Northern Blot analysis. Because both the cim3 mutant and the nahG phenotypes are dominant, epistasis could be analyzed between the two genes in the Fl plants. We analyzed 70 Fl plants from the cross of cim3 X nahG to determine the expression of PR-1 and nahG genes. In the Northern blot analysis of mRNA expression, the presence of the nahG gene was correlated with the deleted expression of the acquired systemic resistance gene. The presence of cim3 in each Fl was confirmed by evaluating the PR-1 mRNA in the resulting F2 segregates.

To determine if the cim3 mutation was epistatic for nahG with respect to disease resistance, 5 Fl plants were auto-crossed from the cross of cim3 X nahG, in which the presence of nahG had been confirmed and the absence of the PR-1 mRNA, and 20 to 30 F2 seeds were planted. The expression of nahG and PR-1 mRNA was analyzed in individuals of this F2 population, which were then stimulated with P. parasitica (NoCo2) to evaluate their susceptibility to diseases. Disease resistance conferred by cim3 was eliminated by the presence of the nahG gene, demonstrating that nahG is epistatic for cim3, for the expression of the acquired systemic resistance gene and the phenotypes of disease resistance.

Example 19: Synergistic Resistance to the Diseases Obtained through the Application of Microbicide and / or BTH to Mutants cim Three days before inoculation of the pathogen, the acquired systemic resistance chemical inducer BTH (benzo [1,2,3] thiadiazole-7-carbothioic acid S-methyl ester) was applied as the 25 percent active ingredient ( ia) with a wettable powder vehicle (Metraux et al., 1991), and / or the metalaxyl microbicide (CGA 48988) formulated as the 25 percent active ingredient, or the wettable powder alone, as a fine mist, to the leaves of plants of 4 weeks of age. The plants were inoculated with a conidia suspension (1.8 x 10? Spores / milliliter) of the compatible pathogen Peronospora parasí tica NoCo2. Following the inoculation, the plants were covered to maintain a high humidity, and were placed in a Percival culture chamber at 17 ° C, with a cycle of 14 hours a day / 10 hours at night (Uknes et al., 1993). The tissue was harvested 8 days after inoculation. Fungal growth was determined using a fungal rRNA probe obtained by Polymerase Chain Reaction, according to White et al. (1990 PCR Protocols: A guide to Methods and Application, 315-322) using the NS1 primers. and NS2, and EmWAde P. parasitic DNA as templates. The RNA was purified from the frozen tissue by extraction with phenol / chloroform, followed by precipitation with lithium chloride (Lagrimini et al 1987; PNAS, 84: 7542-7546). The samples (7.5 micrograms) were separated by electrophoresis through the formaldehyde-agarose gels, and stained on nylon membranes (Hybond-N +, Amersham) as described by Ausbel et al. (1987). Hybridizations and washing were in agreement with Church and Gilbert (1984, PNAS 81: 1991-1995). The relative amounts of transcription were determined using a Phosphor Imager (Molecular Dynamics, Sunnyvale, CA) following the manufacturers' instructions. The sample load was normalized by probing separate filter spots with the constitutively expressed β-tubulin Arabidopsis ADβc. The infestation of the untreated plants corresponded to an inhibition of the fungal growth of O percent. The application of metalaxyl alone, the "plant activator" BTH alone, or both metalaxyl and BTH, to the cim3 mutants described above, produced a disease resistance effect greater than additive, ie, synergistic. This effect was determined as the synergy factor (SF), which is the ratio of the observed effect (O) to the expected effect (E). The following results were obtained: Table 33 Action against Peronospora parasitic in Arabidopsis. Component I: mutation cim3. wt = wild type Col-O ND = not determined Table 34 Action against Peronospora parasí tica in Arabidopsis Component I: mutation cim3. Component II: BTH wt = wild type Col-O ND = not determined Table 35 Action against Peronospora parasitic in Arabidopsis Component I: mutation cim3. wt = wild type Col-O ND = not determined As can be seen from the previous tables, synergistic effects of disease resistance were demonstrated in cim3 plants, by applying metalaxyl only, by applying BTH alone, and by the application of metalaxyl and BTH in combination. For example, in the untreated cim3 plant, an inhibition of fungal growth of 12.5 percent was seen in relation to the untreated wild-type plant; this demonstrates that the constitutive expression of the acquired systemic resistance gene in the cim3 mutant correlates with disease resistance. However, as shown in Table 30, by the application of metalaxyl at 0.0001 grams / liter (a concentration normally insufficient to be effective) to the immunomodulated cim3 plant (activated systemic acquired resistance), the observed level of fungal growth inhibition It increased to 57.8 percent. The synergism factor of 4.6 calculated from these data clearly demonstrates the synergistic effect achieved through the application of a microbicide to an immunomodulated plant. The data presented in Table 31 demonstrate that synergism is also achieved by the application of a chemical inducer of acquired systemic resistance, such as BTH, to an immunomodulated cim3 plant (activated systemic acquired resistance). For example, in wild-type plants, a concentration of 0.03 mM BTH is usually insufficient to confer effective resistance to diseases, providing only an inhibition of fungal growth of 20.8 percent. However, in cim3 plants, this normally inadequate concentration of BTH, provided a fungal growth inhibition of 73.1 percent, which was almost as high as the level of inhibition provided by 0.1 mM BTH, at the recommended concentration to be effective. The synergism factor of 2.2 calculated from the data in Table 31, clearly demonstrates the synergistic effect achieved through the application of BTH to a plant that is already immunomodulated through other means. The effects on disease resistance were even more dramatic when both BTH and metalaxyl were applied to the cim3 plant. As written above in Example 13 (Table 29), in wild type plants, fungal growth inhibition is not achieved by the separate application of either 0.01 mM BTH, or 0.0001 gram / liter of metalaxyl, due to that these concentrations are usually insufficient to be effective. However, by applying both compounds to plants at these normally insufficient concentrations, an inhibition of fungal growth of 40.7 percent was observed, which is a synergistic effect with respect to wild-type plants. In cim3 plants, the simultaneous application of 0.01 mM BTH and 0.0001 grams / liter of metalaxyl resulted in 100 percent inhibition of fungal growth, clearly demonstrating a still further synergistic activity. Accordingly, the combined use of immunomodulated cim plants with usually ineffective low concentrations of chemicals to achieve disease resistance provides advantages that should be visible to those skilled in the agricultural art. The normally toxic or otherwise undesirable concentrations of chemical products can be avoided by taking advantage of the synergisms demonstrated herein. In addition, economic gains can be realized as a result of the lower amount of chemicals required to provide a given level of protection to the plants.

III. Effects of Synergistic Resistance to Diseases Achieved through the Application of Conventional Microbicides and / or Chemical Inductors of Acquired Systemic Resistance to Transgenic Plants Containing NIM1 Gene Forms The NIM1 gene is a key component of the systemic acquired resistance (ARV) pathway in plants (Ryals et al., 1996). The NIM1 gene is associated with the activation of acquired systemic resistance by chemical and biological inducers, and in conjunction with these inducers, it is required for acquired systemic resistance, and for the expression of the acquired systemic resistance gene. The localization of the NIM1 gene has been determined by molecular biological analysis of the genome of mutant plants known to carry the mutant niml gene, which gives host plants extreme sensitivity to a wide variety of pathogens, rendering them incapable of responding to pathogens and chemical inducers of acquired systemic resistance. The wild-type NJM1 gene from Arabidopsis has been mapped and sequenced (SEQ ID? 0: 1). The product of the wild-type NJM1 gene (SEQ ID? 0: 2) is involved in the signal transduction cascade that leads to both acquired systemic resistance and resistance to gene-by-gene diseases in Arabidopsis (Ryals et al. 1997). The recombinant over-expression of the wild-type form of NJM1 gives rise to immunomodulated plants with a constitutive immunity phenotype (MIC), and therefore, confers resistance to diseases in transgenic plants. Increased levels of active NIM1 protein produce the same disease resistance effect as chemical induction with chemical inducers, such as BTH, INA, and SA. See U.S. Patent Application Pending Serial Number 08 / 880,179, incorporated herein by reference. In addition, it has been demonstrated that the product of the NIM1 gene is a structural homolog of the mammalian signal transduction factor and I? B, subclass A (Ryals et al., 1997). IB mutations have been described that act as super-repressors or negative-dominants of the regulation scheme of NF-? B / l B. Accordingly, certain altered forms of NIM1 act as negative-dominant regulators of the transduction pathway of the acquired systemic resistance signal. These altered forms of NIM1 confer the opposite phenotype on plants transformed with it, such as the niml mutant; that is, the immunomodulated plants transformed with altered forms of NJM1 exhibit a constitutive expression of an acquired systemic resistance gene and a CIM phenotype. See the Pending TCP Application "METHODS OF USI? G THE NJMl GENE TO CONFER DISEASE RESISTANCE IN PLANTS" incorporated herein by reference.

Example 20: Transformation of Plants with Cosmic Clones Containing the Wild-type NIM1 Gene The cosmid D7 (deposited in the ATCC on September 25, 1996 as ATCC 97736) was generated from a clone extending the NIM1 gene region, and therefore, includes the wild-type NJM1 gene (SEQ ID? 0: 1). The cosmid El was also generated from a clone that extends the region of the NJM1 gene, and consequently, also includes the wild-type NIM1 gene (SEQ ID? O: 1). Cosmids D7 and El were moved to Agrobacterium tumefaciens AGL-1, through conjugative transfer in a triparental coupling with the helper strain HB101 (pRK2013), as described in United States Patent Application Number 08 / 880,179. These cosmids were then used to transform a niml mutant Arabidop-sis line sensitive to kanamycin, using vacuum infiltration (Mindrinos et al., 1994, Cell 78, 1089-1099). The seeds of the infiltrated plants were harvested, and allowed to germinate on GM agar plates containing 50 milligrams / milliliter of kanamycin as a selection agent. The seedlings that survived the selection were transferred to the soil approximately 2 weeks after coating. The plants transferred to the soil were cultivated in a phytotron for approximately 1 week after the transfer. 300 mM INA was applied as a fine mist to completely cover the plants, using a cronebulizer.

After two days, the leaves were harvested for RNA extraction and PR-1 expression analysis. The plants were then sprayed with parasitic Peronospora (EmWa isolate), and were cultivated under high humidity conditions in a culture chamber at temperatures of 19 ° C / day / 17 ° C at night, and 8-hour light cycles / 16 hours of darkness. From 8 to 10 days after fungal infection, the plants were evaluated and scored as positive or negative for fungal growth. The Ws and niml plants were treated in the same way to serve as controls for each experiment. Total RNA was extracted from the harvested tissue, using a LiCl / phenol extraction regulator (Verwoerd et al., 1989, Nuc Acid Res, 2362). The RNA samples were run on a formaldehyde-agarose gel, and stained on GeneScreen Plus membranes (DuPont). The spots were hybridized with a PR-1 cDNA probe labeled with 32 P. The resulting spots were exposed to a film to determine which transformants were capable of inducing the expression of PR-1 after treatment with INA. To see if any of the transformants D7 and El overexpressed NJM1 due to the effect of the insertion site (position), the primary transformants containing the cosmids D7 or El were auto-crossed, and the seeds were collected T2. The seeds of an El line and 95 D7 lines were sown in the soil, and were cultivated as described above.

When the T2 plants obtained at least 4 true leaves, a single leaf was harvested separately for each plant. The RNA was extracted from this tissue, and analyzed for the expression of PR-1 and NIM1. The plants were then inoculated with P. parasitica (EmWa), and analyzed for fungal growth ten days after the infection. A number of transformants showed fungal growth less than normal, and 4 of them, that is, lines D7-2, D7-74, D7-89, and El-1, showed no visible fungal growth. Plants that show NIM1 and PR-1 expression higher than normal, and that exhibit resistance to the fungus, demonstrate that overexpression of NIM1 confers resistance to diseases.

Example 21: Overexpression of NIM1 Under its Native Promoter Plants constitutively expressing the NIM1 gene were generated from the transformation of wild-type plants Ws with the genomic fragment of NJMl BamHl -HindIII (SEQ ID? O: 1- bases 1249- 5655), containing 1.4 kb of promoter sequence. This fragment was cloned into pSGCGOl, and transformed into Agrobacterium strain GV3101 (pMP90, Koncz and Schell (1986) Mol, Gen. Genet, 204: 383-396). The Ws plants were infiltrated as described above. The resulting seed was harvested and coated on GM agar containing 50 micrograms / milliliter of kanamycin. The surviving seedlings were transferred to the soil, and were tested as described above to determine their resistance to the isolate of Peronospora parasí tica, Emwa. The selected plants were auto-crossed and selected for two subsequent generations, in order to generate homozygous lines. The seeds of several of these lines were sown in the soil, and were cultivated from 15 to 18 plants per line for three weeks, and were tested again to determine their resistance to Emwa without a previous treatment, with an inducer chemical. Approximately 24 hours, 48 hours and 5 days after the fungal treatment, the tissue was harvested, grouped, and frozen for each line. The plants remained in the culture chamber until 10 days after the inoculation, when they were qualified to determine their resistance to Emwa. RNA was prepared from all the samples collected, and analyzed as described previously (Delaney et al., 1995). The spot was hybridized in the probe of the Arabidopsis PR-1 gene (Uknes et al., 1992). Five of the 13 transgenic lines analyzed showed an early induction of PR-1 gene expression. For these lines, the PR-1 mRNA was evident at 24 or 48 hours after the fungal treatment. These five lines also had no visible fungal growth. The leaves were stained with lactophenol blue as described (Dietrich et al., 1994), to verify the absence of fungal hyphae in the leaves. The expression of the PR-1 gene was not induced in the other 8 lines at 48 hours, and these plants did not show resistance to Emwa. A subset of resistant lines was also tested to determine a greater resistance to the bacterial pathogen Pseudomonas syringae DC3000, to evaluate the spectrum of evident resistance, as described by Uknes et al. (1993). The experiments were essentially done as described by Lawton et al., (1996). Bacterial growth was slower in these lines, which also showed a constitutive resistance to Emwa. This shows that plants that overexpress the NIM1 gene under its native promoter have a constitutive immunity against pathogens. To evaluate the additional characteristics of the CIM phenotype in these lines, uninfected plants were evaluated to determine free salicylic acid and conjugated with glucose, and the leaves were stained with lactophenol blue to evaluate the presence of microscopic lesions. Plants with resistance cross sexually with the acquired systemic resistance mutants, such as NahG and ndrl, to establish the epistatic relationship of the resistance phenotype with other mutants, and evaluate how these dominant negative mutants of NJMl can influence the feedback cycle dependent on salicylic acid.

Example 22: Overexpression of NIM1 Powered by 35S The full-length NJM1 AD? C (SEQ ID NO: 6) was cloned into the EcoRi site of pCGN1761 ENX (Comai et al. (1990) Plant Mol. Biol. 15, 373- 381). From the resulting plasmid, an Xbal fragment containing an improved CaMV 35S promoter, the NJM1 cDNA in the correct orientation for transcription, and a 3 'terminator tml was obtained. This fragment was cloned into the binary vector pCIB200 and transformed into GV3101. The Ws plants were infiltrated as described above. The resulting seed was harvested and coated on GM agar containing 50 micrograms / milliliter of kanamycin. The surviving seedlings were transferred to the soil, and tested as described above. The selected plants were auto-crossed, and were selected for two subsequent generations, in order to generate homozygous lines. Nine of the 58 tested lines demonstrated resistance when treated with Emwa without prior chemical treatment. Accordingly, overexpression of the NIM1 cDNA also results in disease resistant plants.

Example 23: NIM1 is an IKBOI Homologue Multiple sequence alignment was performed between the NJM1 and I? B protein gene products, whereby it was determined which product of the NIM1 gene is a homologue of I? B (Figure 1) . Sequence homology searches were performed using BLAST (Altschul et al., J., Mol. Biol. 215, 403-410 (1990).) Multiple sequence alignment was constructed using Clustal V (Higgins et al., CABIOS 5, 151 -153 (1989)) as part of the Lasergene Biocom-putación Software package from DNASTAR (Madison, Wl) The sequences used in the alignment were NIM1 (SEQ ID N0: 2), and mouse I? B (SEQ ID. N0: 3), GenBank Accession Number: 1022734), I? Ba of rat (SEQ ID N0: 4, GenBank, Accession Nos. 57674 and X63594; Tewari et al., Nucleic Acids Res. 20, 607 (1992)), and Pig I Ba (SEQ ID NO: 5, GenBank, Accession number Z21968, de Martin et al, EMBO J. 12, 2773-2779 (1993); GenBank accession number 517193, by Martin et al., Gene 152 , 253-255 (1995).) The parameters used in the Clustal analysis had a gap violation of 10, and a hole length infraction of 10. The distances were calculated s of evolutionary divergence using the PAM250 weight table (Dayhoff et al., "A model of evolutionary change in proteins. Matrices for detecting distant relationships. "In Atlas of Protein Sequence and Structure, volume 5, Supplement 3, MO, Dayhoff, editors (National Biomedical Research Foundation, Washington, DC), pages 345-358 (1978).) Similarity was calculated of residues using a modified Dayhoff table (Schwartz and Dayhoff, "A model of evolutionary change in proteins." In Atlas of Protein Sequence and Structure, MO, Dayhoff, editors (National Biomedical Research Foundation, Washington, DC). pages 353- 358 (1979), Gribskov and Burgess, Nucleic Acids Res. 14. 6745-6763 (1986)).

The homology searches indicate similarity of NIM1 with the ankyrin domains of several proteins, including: ankyrin NF-KB and I? B. The best global homology is for I B, and the related molecules (Figure 1). NIM1 contains two serines at amino acid positions 55 and 59; the serine in position 59 is in a context (D / ExxxxS) and in the position (N-terminal) consistent with a role in an inducible degradation dependent on phosphorylation, mediated by ubiquitin. All IKBOÍS have these N-terminal serines, and are required for the inactivation of I? B and the subsequent release of NF-KB. NIM1 has ankyrin domains (amino acids 262-290 and 323-371). It is believed that the ankyrin domains are involved in protein-protein interactions, and are a ubiquitous feature for the I? B and NF-KB molecules. The C ends of IKBS may be different. NIM1 has some homology with a region rich in QL (amino acids 491-499) which is found in terms C of some IKBS.

Example 24: Generation of Altered Forms of NIM1 - Changes of Serine Residues 55 and 59 to Alanine Residues Phosphorylation of serine residues in human I? B is required for degradation activated by stimulation of I? Ba, activating from this way NF-KB. The mutagenesis of serine residues (S32-S36) in human IKBOI, to alanine residues, inhibits stimulus-induced phosphorylation, thus blocking IKBOI proteosome-mediated degradation (E.Britta-Mareen Traenckner IKBCÜ, EMBO J ".14: 2876-2883 (1995); Brown et al., Science 267: 1485-1488 (1996); Brockman et al., Molecular and Cellular Biology 15: 2809-2818 (1995); Wang et al., Science 274: 784-787 (1996).) This altered form of I? Ba functions as a dominant negative form, retaining NF-KB in the cytoplasm, thereby blocking downstream signaling events, based on sequence comparisons between NIM1 and IKB, serines 55 (S55) and 59 (S59) of NIM1 are homologous to S32 and S36 in human IKBQ !. To construct the negative-dominant forms of NIM1, serines at amino acid positions 55 and 59 are mutagenized to residues of alanine, this can be done by any cone method This is done by experts in the field, such as, for example, by using the Mutagenesis Kit Directed to the QuikChange Site (# 200518: Strategene). Using a full-length NJM1 cDNA (SEQ ID? O: 6), including 42 base pairs of the 5 'untranslated sequence (UTR), and 187 base pairs of 3' UTR, the mutagenized construct can be made according to the manufacturer's instructions, using the following primers (SEQ ID? 0: 6, positions 192-226): 5 '- CAA CAG CTT CGA AGC CGT CTT TGA CGC GCC GGA TG-3' (SEQ ID? O: 25), and 5 '-CAT CCG GCG CGT CAA AGA CGG CTT CGA AGC TGT TG-3' (SEQ ID? O: 26), where the underlined bases denote the mutations. The strategy is as follows. * The AD? of NJMl cloned into the vector pSE936 (Elledge et al., Proc. Nat. Acad. Sci. USA 88: 1731-1735 (1991)) is denatured, and the primers containing the altered bases are quenched. The 7AD polymerase? (Pfu) extends the primers by a non-chain offset, resulting in tight circular chains. The AD? it is subjected to digestion with restriction endonuclease, with Dpnl, which only cuts the methylated sites (AD? of non-mutagenized template). The remaining circular AD? Ds is transformed into E. coli strain XLl-Blue. The plasmids from the resulting colonies are extracted and sequenced to verify the presence of the mutated bases, and to confirm that no other mutations are present. The AD? C of mutagenized NIM1 is digested with the restriction endonuclease EcoRI and cloned into pCG? 1761 under the transcription regulation of the double 35S promoter of the cauliflower mosaic virus. The transformation cassette including the 35S promoter, the AD? C of NIM1, and the tmJ terminator is released from pCG? 1761 by partial restriction digestion with Xbal, and ligated into the Xbal site of dephosphorylated pCIB200. The SEQ ID? Os: 7 and 8 show the coding sequence of the AD ?, and the encoded amino acid sequence, respectively, of this altered form of the NIM1 gene.

Example 25: Generation of Altered N-terminal Suppression Forms of NIM1 Deletion of amino acids 1-36 (Brockman et al., Sun et al.), Or 1-72 (Sun et al.) Of human I? B including K21 , k22, S32, and S36, results in a negative-dominant I? B phenotype in transfected human cell cultures. An N-terminal deletion of approximately the first 125 amino acids of the NIM1 cDNA encoded product removes 8 lysine residues that can serve as potential sites of ubiquitination and also removes the putative phosphorylation sites at S55 and S59 (see Example 2). This altered genetic construct can be produced by any means known to those skilled in the art. For example, using the method of Ho and collaborators, Gene 77: 51-59 (1989), a form of NIM1 can be generated in which the 7DNA encoding approximately the first 125 amino acids is deleted. The following primers produce a polymerase chain reaction product of 1.612 base pairs (SEQ ID NO: 6: 418 to 2011): 5 '-gg aat tca-AIG GAT TCG GTT GTG ACT GTT TTG-3' (SEQ ID NO: 27) and 5 '-gga att ATTA ATT TGT ATA CCA TTG G-3' (SEQ ID NO: 28) where the synthetic start codon is underlined (? GI) and the EcoRI linker sequence is in lower case. Amplification of the fragments uses a reaction mixture comprising from 0.1 to 100 nanograms of the template DNA, 10 mM Tris, pH of 8.3 / 50 mM KCl / 2 mM MgCl 2 / 0.001% gelatine / 0.25 mM of each dNTP / 0.2 mM of each primer, and one unit of rTth DNA polymerase in final volume of 50 milliliters, and a Perkin Elmer Cetus 9600 polymerase chain reaction machine. The conditions of the Polymerase Chain Reaction are as follows: ° C for 3 minutes: 35 times (94 ° C for 30 seconds: 52 ° C for 1 minute: 72 ° C for 2 minutes): 72 ° C for 10 minutes. The product of the Polymerase Chain Reaction is cloned directly into the vector pCR2.1 (Invitrogen). The insert generated by the Polymerase Chain Reaction, in the vector of the Polymerase Chain Reaction, is released by digestion with restriction endonuclease, using EcoRI, and is ligated into the EcoRI site of dephosphorylated pCGN1761, under the regulation transcription of the double 35S promoter. The construction is sequenced to verify the presence of the synthetic start ATG, and to confirm that no other mutations occur during the Polymerase Chain Reaction. The transformation cassette including the 35S promoter, the modified NIM1 cDNA, and the tmJ terminator is released from pCGN1761 by partial restriction digestion with Xbal, and ligated into the Xbal site of pCIB200. SEQ ID NOs: 9 and 10 show the coding sequence of the DNA, and the encoded amino acid sequence, respectively, of an altered form of the NJM1 gene, which has a β-terminal amino acid deletion.

Example 26: Generation of Altered Forms of C-terminal Suppression of NJMl It is believed that suppression of amino acids 261-317 of human I? B results in better intrinsic stability, by blocking the constitutive phosphorylation of the residues of serine and trionine in the term C. A region rich in serine and trionine is present in amino acids 522-593, in the C-terminus of? IM1. The C-terminal coding region of the NIM1 gene can be modified by deleting the nucleotide sequences encoding amino acids 522-593. Using the method of Ho et al. (1989), the C-terminal and 31 UTR coding region of the AD? C of NJMl (SEQ ID? O: 6: 1606-2011) is suppressed by polymerase chain reaction, generating a fragment of 1623 base pairs, using the following primers: 5 '-cggaattcGATCTCTTTAATTTGTGAATTT C-3' (SEQ ID O: 29), and 5 '-ggaattc ^ AACAGTT CATAATCTGGTCG-3' (SEQ ID? O: 30), where a synthetic stop codon is underlined (TGA in the complementary strand), and the EcoRI linker sequences are in lower case. The components of the polymer chain reaction are as described above, and the parameters of the cyclization are as follows: 94 ° C for 3 minutes; 35 times (94 ° C for 30 seconds: 52 ° C for 30 seconds: 72 ° C for 2 minutes); 72 ° C for 10 minutes. The product of the Polymerase Chain Reaction is cloned directly into the vector pCR2.1 (Invitrogen). The insert generated by the Polymerase Chain Reaction, in the vector of the Polymerase Chain Reaction, is released by restriction endonuclease digestion, using EcoRI, and ligated into the EcoRI site of dephosphorylated pCGN1761, which contains double 35S promoter. The construct is sequenced to verify the presence of the synthetic stop codon within the frame, and to confirm that no other mutations are present during the Polymerase Chain Reaction. The transformation cassette including the promoter, the modified NJMl cDNA, and the tml terminator is released from pCG? 1761 by partial restriction digestion with Xbal, and ligated into the Xbal site of the dephosphorylated pCIB200. SEQ ID: Os: 11 and 12 show the coding sequence of the AD ?, and the encoded amino acid sequence, respectively, of an altered form of the NJM1 gene, which has a C-terminal amino acid deletion.

Example 27: Generation of Altered Forms of Suppression Chimera? -terminal / C-terminal of NIM1 A form of suppression? -terminal and C-terminus of NIM1 is generated, using a unique Kpnl restriction site at position 819 (SEQ ID ? O: 6). The? -terminal suppression form (Example 25) is the restriction endonuclease digested with EcoRl / Kpnl, and the 415 base pair fragment corresponding to the term? modified is recovered by gel electrophoresis. In the same manner, the C-terminal deletion form (Example 26) is digested with restriction endonuclease, with EcoRI / Kpnl, and the 790 base pair fragment corresponding to the modified C terminus is recovered by gel electrophoresis. The fragments are ligated at 15 ° C, digested with EcoRI to remove EcoRI concatemers, and cloned into the EcoRI site of dephosphorylated pCGN1761. The N / C-terminal deletion form of NJMl is under the transcription regulation of the double 35S promoter. In a similar manner, a chimeric form of NIM1 consisting of the mutagenized phosphorylation sites S55 / S59 (Example 24) fused with the C-terminal deletion (Example 26) is generated. The construction is generated as described above. The constructs are sequenced to verify the fidelity of the start and stop codons, and to confirm that no mutations occur during cloning. The respective transformation cassettes, including the 35S promoter, the NIM1 chimera, and the tmJ terminator are released from pCG? 1761 by partial restriction digestion with Xbal, and ligated into the Xbal site of the dephosphorylated pCIB200. SEQ ID: Os: 13 and 14 show the coding sequence of the AD ?, and the encoded amino acid sequence, respectively, of an altered form of the NIM1 gene that has both -terminal and C-terminal amino acid deletions.

Example 28: Generation of Altered Forms of Ankirin Domains of NIM1 NIM1 exhibits homology with ankyrin motifs at approximately amino acids 103-362. Using the method of Ho et al., (1989), the DNA sequence encoding the putative ankyrin domains (SEQ ID NO: l: 3093-3951) is amplified by the Polymerase Chain Reaction (conditions: 94 ° C for 3 minutes: 35 times (94 ° C for 30 seconds: 62 ° C for 30 seconds: 72 ° C for 2 minutes): 72 ° C for 10 minutes) from NIM1 cDNA (SEQ ID NO: 6: 349 -1128) using the following primers: 5'-ggaattaaAIG.GACTCCAACAACACCGCCGC-3 '(SEQ ID NO: 31), and 5'-ggaattcICAACCTTCCAAAGTTGCTTCTGATG-3' (SEQ ID NO: 32). The resulting product is digested with restriction endonuclease, with EcoRI, and then spliced into the EcoRI site of dephosphorylated pCGN1761 under the transcription regulation of the double 35S promoter. The construction is sequenced to verify the presence of the synthetic start codon (ATG), a stop codon in the frame (TGA), and to confirm that no other mutations occur during the Polymerase Chain Reaction. The transformation cassette including the 35S promoter, the ankyrin domains, and the tml terminator is released from pCGN1761 by partial restriction digestion with Xbal, and ligated into the XfoaJ site of the dephosphorylated pCIB200. SEQ ID NOs: 15 and 16 show the coding sequence of the DNA, and the encoded amino acid sequence, respectively, of the ankyrin domain of NJM1.

Example 29: Construction of Chimeric Genes To increase the possibility of appropriate spatial and temporal expression of the altered NJM1 forms, a HindIII / BamHI fragment of 4.407 base pairs (SEQ ID? O: l: bases 1249-5655) is used, and / or an EcoRV / BamHI fragment of 5,655 base pairs (SEQ ID? O: 1: bases 1-5655), which contains the NJM1 promoter and the gene is used for the creation of the altered NIM1 forms of Examples 24 to 28 previous Although the construction steps may be different, the concepts are comparable with the Examples previously described herein. Strong overexpression of altered forms can be potentially lethal. Accordingly, the altered forms of the NJM1 gene described in Examples 24 to 28 can be placed under the regulation of promoters different from the endogenous NIM1 promoter, including, but not limited to, promoter nos, or the small subunit of the Rubisco promoter. In the same manner, altered NIM1 forms can be expressed under the regulation of the promoter that responds to the pathogen PR-1 (U.S. Patent Number 5,614,395). This expression allows to have a strong expression of the NIM1 forms altered only under an attack of pathogen or other activating conditions of the acquired systemic reaction. In addition, resistance to diseases in transformants expressing the NJM1 forms altered under the regulation of the PR-1 promoter may be evident, when treated with concentrations of acquired systemic resistance activating compounds (ie, BTH or I? A) that normally do not activate acquired systemic resistance, thus activating a feedback loop (Weymann et al., (1995) Plant Cell 7: 3013-2022).

Example 30: Transformation of Altered Forms of NIM1 in Arabidopsis thaliana Generated constructs (Examples 24 to 29) are moved to Agrobacterium tumefaciens by electroporation to strain GV3101. These constructions are used to transform Arabidopsis, ecotypes Col-O and Ws-0 by vacuum infiltration (Mindrinos et al., CelJ 78, 1089-1099 (1994)) or by conventional root transformation. The seeds of these plants are harvested and allowed to germinate on agar dishes with kanamycin (or another appropriate antibiotic) as a selection agent. Only the seedlings that are transformed can detoxify the selection agent and survive. The seedlings that survive the selection are transferred to the soil and tested to determine a CIM phenotype (constitutive immunity). The plants are evaluated to determine the observable phenotypic differences, compared to the wild-type plants.

Example 31: Evaluation of the CIM phenotype in Plants Transformed with the Wild-type NIM1 Gene, or an Altered Form of the NIM1 Gene One leaf is harvested from each primary transformant, the RNA is isolated (Verwoerd et al., 1989, Nuc Acid Res., 2362) and is tested for the constitutive expression of PR-1 by RNA stain analysis (Uknes et al., 1992). Each transformant is evaluated to determine an improved disease resistance response, which indicates the analysis of the constitutive expression of acquired systemic resistance (Uknes et al., 1992). Conidia suspensions of 5-10 x 10 spores / milliliter are prepared from 2 compatible isolates of P. parasí tica, Emwa and Ñoco (ie, these fungal strains cause the disease in wild-type plants Ws-0 and Col-O, respectively), and the transformants are sprayed with the appropriate isolate, depending on the ecotype of the transformant. The inoculated plants are incubated under high humidity for 7 days. The disease of the plants is evaluated on day 7, and a single leaf is harvested for RNA stain analysis, using a probe that provides a means to measure fungal infection. Transformants that exhibit a CIM phenotype are taken as the TI generation, and homozygous plants are identified. The transformants are subjected to a battery of resistance tests against diseases, as described below. The fungal infection is repeated with Ñoco and Emwa, and the leaves are stained with lactophenol blue to identify the presence of fungal hyphae, as described in Dietrich et al., (1994). The transformants are infected with the bacterial pathogen Pseudomonas syringae DC3000 to evaluate the spectrum of evident resistance, as described in Uknes et al., (1993). Uninfected plants are evaluated for both free salicylic acid and glucose conjugate, and the leaves are stained with lactophenol blue to evaluate the presence of microscopic lesions. Resistant plants are sexually crossed with mutants of acquired systemic resistance, such as NaHG (U.S. Patent Number 5,614,395), and ndrl, to establish the epistatic relationship of the resistance phenotype with other mutants, and to evaluate the manner in which these negative-dominant mutants of NIM1 can influence the feedback cycle dependent on salicylic acid.

Example 32: Isolation of NIM1 Homologs NIM1 homologs can be obtained that hybridize under moderately stringent conditions, either to the entire NJMl gene of Arabidopsis, or preferably, to an oligonucleotide probe derived from the NIM1 gene of Arabidopsis that comprises a contiguous portion of its coding sequence, at least about 10 nucleotides in length. The factors that affect the stability of the hybrids determine the stringency of the hybridization. One of these factors is the melting temperature Tf which can be easily calculated according to the formula provided in DNA PROBES, George H. Keller and Mark M. Manak, Macmillan Publishers Ltd, 1993, section 1: Molecular Hybridization Technology; pages 8 and following. The preferred hybridization temperature is in the range of about 25 ° C below the calculated melting temperature Tf which can be easily calculated according to the formula provided in DNA PROBES, George H. Keller and Mark M. Manak, Tf preferably on the scale of about 12 to 15 ° C below the calculated melting temperature Tf and in the case of the oligonucleotides, on the scale of about 5 to 10 ° C below the melting temperature Tf. Using NJM1 cDNA (SEQ ID? 0: 6) as a probe, NJMl homologs of Arabidopsis are identified through the selection of genomic or AD? C libraries from different cultures, such as, but not limited to, those mentioned above. further in Example 33. Standard techniques for accomplishing this include selection by hybridization of AD libraries. coated (either plates or colognes; see, for example, Sambrook et al., Molecular Cloning, editors, Cold Spring Harbor Laboratory Press. (1989)) and amplification by polymerase chain reaction, using oligonucleotide primers (see, for example, Innis et al., PCR in the expression vectors herein, and transformed into the above-mentioned cultures.) Transformants are evaluated To determine its best resistance to diseases, using relevant pathogens of the crop plant being tested, homologues of NIM1 have been detected in the genomes of cucumber, tomato, tobacco, corn, wheat, and barley, by analysis of DNA Genomic DNA was isolated from cucumber, tomato, tobacco, corn, wheat, and barley, digested by cooling with the enzymes BamH1, HindIII, Xbal, or SalI, separated electrophoretically on 0.8 percent agarose gels, and it was transferred to a nylon membrane by capillary stain, then the ultraviolet crosslinking to fix the DNA, the membrane was hybridized under low stringent conditions ncia [(1 percent bovine serum albumin, * NaP04, 520 mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; EDTA lmM; 250 mM sodium chloride) at 55 ° C for 18 to 24 hours], with 3P radiolabelled Arabidopsis thaliana NIM1 cDNA. Following the hybridization, the spots were washed under conditions of low stringency [SSC6X for 15 minutes (3 times) SSC3X for 15 minutes (1 time) at 55 ° C; SSCIX is 0.15 M NaCl, 15 mM sodium citrate, (pH 7.0), and exposed to X-ray film to visualize the bands corresponding to NIM1. In addition, expressed sequence tags (ESTs) identified with similarity to the NIM1 gene can be used to isolate homologues. For example, several expressed sequence tags (ESTs) of rice with similarity to the NIM1 gene have been identified. A multiple sequence alignment was constructed using Clustal V (Higgins, Desmond G. and Paul M. Sharp (1989), Fast and sensitive multiple sequence alignments on to microcomputer, CHANGES 5: 151-153) as part of the DNA * ( 1228 South Park Street, Madison Wisconsis, 53715) Lasercom Biocom-putación Software package for Macintosh (1994). Certain regions of the NJM1 protein are homologous in the amino acid sequence to 4 different rice AD? C protein products. Homologies were identified using the NJM1 sequences in a BLAST search of GenBank. Comparisons of the regions of homology in NJMl and in the rice AD? C products are shown in Figure 2 (See also, SEQ ID O: 2 and SEQ ID? Os: 17-24). Fragments of the? IM1 protein show 36 to 48 percent identical amino acid sequences with the four rice products. This rice ESTs can be especially useful for the isolation of NIM1 homologs, from other monocots. Homologs can also be obtained by Polymerase Chain Reaction. In this method, comparisons are made between known counterparts (eg, rice and Arabidopsis). Then the regions of high similarity or identity of amino acids and AD are used. to make the primers of the Polymerase Chain Reaction. Regions rich in amino acid residues M and W are best followed by regions rich in amino acid residues F, Y, C, H, Q, K, and E, because these amino acids are encoded by a limited number of codons. Once a suitable region is identified, primers are made for that region with a variety of substitutions at the position of the third codon. This diversity of substitution in the third position can be limited, depending on the species to which it is directed. For example, because corn is rich in GC, primers are designed that use a G or a C in the third position, if possible. The Polymerase Chain Reaction is made from cDNA, or genomic DNA, under a variety of conventional conditions. When a band can be seen, it is cloned and / or sequenced to determine if it is a NIM1 homologue.

Example 33: Expression of an NJM1 Form in Crop Plants These constructs that confer a CIM phenotype in Col-O or Ws-0, are transformed into crop plants for evaluation. Alternatively, the native NJM1 genes altered from the cultures of the previous example are returned to the respective cultures. Although the NIM1 gene can be inserted into any plant cell that falls within these broad classes, it is particularly useful in cells of crop plants, such as rice, wheat, barley, rye, corn, potato, carrot, sweet potato, beet sugar, beans, peas, chicory, lettuce, cabbage, cauliflower, broccoli, turnip, radish, spinach, asparagus, onion, garlic, eggplant, pepper, celery, carrot, chayote, pumpkin, zucchini, cucumber, apple, pear, quince, melon, plum, cherry, peach, nectarine, apricot, strawberry, grape, raspberry, blackberry, pineapple, avocado, papaya, mango, banana, soybean, tobacco, tomato, sorghum, and sugar cane. Transformants are evaluated to determine their best resistance to diseases. In a preferred embodiment of the invention, expression of the NIM1 gene is at a level that is at least two times higher than the expression level of the native NIM1 gene in wild-type plants, and is preferably 10 times higher than the level of expression of the wild type.

Example 34: Synergistic Resistance to Diseases Obtained through the Application of a Conventional Microbicide to Transgenic Plants Overexpressing NIM1 The plant lines used in this example (6E and 7C) were generated from the transformation of wild type Arabidopsis thaliana plants (ecotype Ws), with the genomic fragment of NJMl, BamHl -HindIII (SEQ ID? 0: 1 - bases 1249-5655), as described above in Example 21. The fungicides metalaxyl, fosetyl, and copper hydroxide, formulated as 25 percent, 80 percent, and 70 percent active ingredient ( ia), respectively with a wettable powder carrier, such as a fine mist, to the leaves of transgenic Ws plants 3 weeks of age constitutively expressing the NJM1 gene. The wettable powder was applied only as a control. Three days later the plants were inoculated with a conidia suspension of Peronospora parasitic isolate, Emwa (1-2 x 105 spores / milliliters), as described in Delaney et al. (1995). Following the inoculation, the plants were covered to maintain a high humidity, and were placed in a Percival culture chamber at 17 ° C, with a cycle of 14 hours of day / 10 hours of night (Uknes et al., 1993). . The tissue was harvested 8 days after inoculation. The progress of fungal infection was followed for 12 days, looking under a dissecting microscope, to qualify the development of conidiospores (Delaney et al., (1994), Dietrich et al. (1994)). Dyeing with lactophenol-trypan blue from the individual leaves was performed to observe the fungal growth inside the leaf tissue. Fungal growth was quantified using a fungal probe of AR? R obtained by means of Polymerase Chain Reaction, according to White et al., (1990, PCR Protocols: A guide to Methods and Application, 315-322) using the primers? S1 and? S2, and the AD? of P. parasí tica EmWa as templates. The AR? it was purified from the frozen tissue by extraction with phenol / chloroform, followed by precipitation with lithium chloride (Lagrimini et al., 1987: PNAS, 84: 7542-7546). Samples (7.5 micrograms) were separated by electrophoresis through formaldehyde-agarose gels, and stained on nylon membranes (Hybond-N +, Amersham), as described by Ausbel et al., (1987). Hybridizations and washing were according to Church and Gilbert (1984, PNAS, 81: 1991-1995). The relative amounts of transcription were determined using a Phosphor Imager (Molecular Dynamics, Sunnyva-le, CA) following the instructions of the manufacturers. The sample loading was normalized by probing separate filter spots with the constitutively expressed b-tubulin Arabidopsis cDNA. The infestation of the untreated plants corresponded to a fungal growth inhibition of 0 percent. The application of metalaxyl, fosetyl, or copper hydroxide to the lines of plants that overexpress NIM1, produced a disease resistance effect greater than additive, that is, synergistic. This effect was determined as the synergy factor (SF), which is the proportion of the observed effect (0) to the expected effect (E). The following results were obtained: Table 36 Action against Peronospora parasitic on Arabidopsis Component I: overexpression of NIM1 (line 6E). Component II: metalaxyl wt Ws wild type Table 37 Action against Peronospora parasitic in Arabidopsis Component I: overexpression of NIM1 (line 6E).

Component II: fosetyl Wt Ws of wild type Table 38 Action against Peronospora parasitic in Arabidopsis Component I: overexpression of NIM1 (line 7C). Component II: fosetyl wt = Wild type Ws Table 39 Action against Peronospora parasitic in Arabidopsis Component I: overexpression of NJMl (line 6E). Component II: copper hydroxide wt = wild type Ws Table 40 Action against Peronospora parasitic on Arabidopsis Component I: overexpression of NIM1 (line 7C). Component II: copper hydroxide wt = wild-type Ws As can be seen from the previous tables, the effects of synergistic resistance to diseases in plants that over-express NIM1 were demonstrated by the application of metalaxyl, fosetyl, and copper hydroxide. For example, in the untreated NIM1 plant (line 6E), a fungal growth inhibition of 10 percent was seen in relation to the untreated wild type plant; this demonstrates that the constitutive expression of the systemic resistance gene acquired in this overexpressor of NIM1 correlates with disease resistance. However, as shown above in Table 37, by the application of fosetyl at 5.0 grams / liter (a concentration normally insufficient to be effective) to the plant overexpressing immunomodulated NIM1 (activated systemic acquired resistance), the observed level of inhibition of fungal growth increased to 93 percent. The synergism factor of 5.5 calculated from these data clearly demonstrates the synergistic effect achieved by applying a microbicide to an immunomodulated plant (activated systemic acquired resistance). In another example, in the untreated NIM1 plant (line 7C), a fungal growth inhibition of 14 percent was seen in relation to the untreated wild-type plant, demonstrating that the constitutive expression of the acquired systemic resistance gene in this NIM1 overexpressor, it correlates with disease resistance. However, as shown above in Table 40, by the application of copper hydroxide in 2.0 grams / liter (a concentration normally insufficient to be effective) to the plant overexpressing immunomodulated NIM1 (activated systemic acquired resistance), the observed level of inhibition of fungal growth was increased to 77 percent. The synergism factor of 5.5 calculated from these data further demonstrates the synergistic effect achieved by applying a microbicide to an immunomodulated plant (activated systemic acquired resistance). Accordingly, the combined use of immunomodulated flats expressing NIM1 with generally ineffective low concentrations of microbicides to achieve disease resistance provides advantages that should be apparent to those skilled in the agricultural arts. The normally toxic or otherwise undesirable concentrations of microbicides can be avoided, taking advantage of the synergies demonstrated herein. In addition, economic gains can be realized as a result of the lower amount of microbicides required to provide a given level of protection to the plants. Example 35: Synergistic Resistance to Diseases Obtained by Application of a Chemical Inductor of Acquired Systemic Resistance to Transgenic Plants Overexpressing NIM1 Transgenic plants containing the genomic DNA fragment of NIM1 were also analyzed under their own promoter (Example 21), for determine its response to different concentrations of BTH in relation to the wild-type line Ws. Seeds were planted from each line, and were cultured as described above. Approximately three weeks after planting, leaf samples were harvested from each line (controls on day 0), and the remaining plants were treated with H20, 10 μM BTH, or 100 μM BTH. Additional samples were harvested on days 1, 3, and 5 following treatment. After harvesting the samples from day 3, a subset of plants was removed for each line, and treated with Peronospora parasitic isolate, Emwa, as described above. RNA was prepared from the harvested tissue, and Northern analysis was performed using the PR-1 Arabidopsis gene probe. The plants were graded to determine their fungal resistance 8 days after the infection. The results of the Northern analysis for Ws, and four of the lines overexpressing NIM1 (3A, 5B, 6E, and 7C) are shown in Figure 3. The expression of the PR-1 gene in the wild-type Ws line was barely detectable after the low level of treatment with BTH lOμM (a BTH concentration of 100 to 300 μM is usually required to be effective). The flat Ws of this treatment were also still susceptible to fungal pathogen P. parasí tica (Emwa). However, in all the lines that overexpressed NIM1, there was a much stronger response for the expression of the PR-1 gene following treatment with BTH at a low level. In addition, all the lines that expressed about NJMl treated with BTH lOμM, showed a complete or almost complete resistance to P. parasí tica. Leaves stained with lactophenol blue to identify the presence of fungal hyphae (Dietrich et al., (1994)) confirmed the absence of fungal growth in lines overexpressing NIM1. The expression of the PR-1 gene in leaf tissue following treatment with 100 μM BTH, was also much stronger and faster in the lines overexpressing NJM1 in relation to the wild type. Therefore, immunomodulated plants are able to respond much faster and at much lower doses of BTH, as shown by PR-1 gene expression and P. parasitic resistance, than wild-type plants. These data demonstrate that synergistic resistance to diseases is achieved by the application of a chemical inducer of acquired systemic resistance, such as BTH to an immunomodulated plant (activated systemic acquired resistance), such as a plant that overexpresses NIM1. Therefore, the combined use of immunomodulated plants that overexpress NIM1 with low, usually ineffective, concentrations of chemical inducers of acquired systemic resistance, such as BTH, to achieve resistance to diseases, provides advantages that should be apparent to those skilled in the art. agricultural technique The normally toxic or otherwise undesirable concentrations of the acquired systemic resistance inducing chemicals can be avoided, taking advantage of the synergies demonstrated herein. In addition, economic gains can be realized as a result of the lower amount of acquired systemic resistance inducing chemicals required to provide a given level of protection to the plants.UENCES (1) GENERAL INFORMATION: (i) APPLICANT: (A) NAME: Novartis AG (B) STREET: Schwarzwaldallee 215 (C) CITY: Basel (E) COUNTRY: Switzerland (F) POSTAL CODE: (ZIP): 4002 (G) ) TELEPHONE: +41 61 69 11 11 (H) TELEFAX: +41 61 696 79 76 (I) TELEX: 962 991 (v) COMPUTER-FRIENDLY FORM (A) TYPE OF MEDIUM: flexible disk (B) COMPUTER: IBM PC compatible (C) OPERATING SYSTEM: PC-DOS / MS-DOS (D) SOFTWARE: Patentln Relay # 1.0, Version # 1.30 (ii) TITLE OF THE INVENTION: METHOD FOR THE PROTECTION OF PLANTS (iii) NUMBER OF SEQUENCES: 32 (2) INFORMATION FOR SEQ ID NO: 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 5655 base pairs (B) TYPE: Nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETIC: NO (iv) ANTI-SENSE: NO (ix) CHARACTERISTICS: (A) NAME / KEY: exon (B) LOCATION: 2787..3347 (D) OTHER INFORMATION: / product = "first exon of NIM1" (ix) CHARACTERISTICS: (A) NAME / KEY: exon (B) LOCATION: 3427..4162 (D) OTHER INFORMATION: / product = "second exon of NIM1" ( ix) CHARACTERISTICS: (A) NAME / KEY: exon (B) LOCATION: 4271..4474 (D) OTHER INFORMATION: / product = "third exon of NIM1" (ix) FEATURES: (A) NAME / KEY: exon ( B) LOCATION: 4586..4866 (D) OTHER INFORMATION: / product = "fourth exon of NIM1" (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: union (2787..3347, 3427. .4162, 4271..4474, 4586..4866) (xi) DESCRIPTION OF THE SEQUENCE CIA: SEQ ID NO: 1: TGTGATGCAA GTCATGGGAT ATTGCTTTGT GT AAGTATA CAAAACCATC ACGTGGATAC 60 ATAGTCTTCA AACCAACCAC TAAACAGTAT CAGGTCATAC CAAAGCCAGA AGTGAAGGGT 120 TGGGATATGT CATTGGGTTT AGCGGTAATC GGATTGAACC CTTTCCGGTA TAAAATACAA 180 AGGCTTTCGC AGTCTCGGCG TATGTGTATG TCTCGGGGTA TCTACCATTT GAATCACAGA 240 ACTTTTATGT GCGAAGTTTT CGATTCTGAT TCGTTTACCT GGAAGAGATT AGAAAATTTG 300 CGTCTACCAA AAACAGACAG ATTAATTTTT TCCAACCCGA TACAAGTTTC GGGGTTCTTG 360 CATTGGATAT CACGGAACAA CAATGTGATC CGGTTTTGTC TCAAAACCGA AACTTGGTCC 420 TTCTTCCATA CTCCGAACTC TGATGTTTTC TCAGGATTAG TCAGATACGA AGGGAAGCTA 480 GGTGCTATTC GTCAGTGGAC AAACAAAGAT CAAGAAGATG TTCACGAGTT ATGGGTTTTA 540 AAGAGCAGTT TTGAAAAGTC GTGGGTTAAA GTGAAAGATA TTAAAAGCAT TGGAGTAGAT 600 TTGATTACGT GGACTCCAAG CAACGACGTT GTATTGTTTC GTAGTAGTGA TCGTGGTTGC 660 CTCTACAACA TAAACGCAGA GAAGTTGAAT TTAGTTTATG CAAAAAAAGA GGGATCTGAT 720 TGTTCTTTCG TTTGTTTTCC GTTTTGTTCT GATTACGAGA GGGTTGATCT GAACGGAAGA 780 AGCAACGGGC CGACACTTTA AAAAAAAAAT AAAAAAAATG GGCCGACAAA TGCAAACGTA 840 GTTGACAAGG ATCTCAAGTC TCAAGTCTCA ATTGGCTCGC TCATTGTGGG GCATAAATAT 900 ATCTAGTGAT GTTTAATTGT TTTTTATAAG GTAAAAAGGA ATATTGAATT TTGTTTCTTA 960 GGTTTATGTA ATAATACCAA ACATTGTTTT ATGAATATTT AATCTGATTT TTTGGCTAGT 1020 TATTTTATTA TATCAAGGGT TCCTGTTTAT AGTTGAAAAC AGTTACTGTA TAGAAAATAG_1080_TGTCCCAATT TTCTCTCTTA AATAATATAT TAGTTAATAA AAGATATTTT AATATATTAG_1140_ATATACATAA TATCTAAAGC AACACATATT TAGACACAAC ACGTAATATC TTACTATTGT 1200 TTACATATAT TTATAGCTTA CCAATATAAC CCGTATCTAT GTTTTATAAG CTTTTATACA 1260 ATATATGTAC GGTATGCTGT CCACGTATAT ATATTCTCCA AAAAAAACGC ATGGTACACA 1320 AAATTTATTA AATATTTGGC AATTGGGTGT TTATCTAAAG TTTATCACAA TATTTATCAA 1380 CTATAATAGA TGGTAGAAGA TAAAAAAATT ATATCAGATT GATTCAATTA AATTTTATAA 1440 TATATCATTT TAAAAAATTA AT AAAAGAA AACTATTTCA TAAAATTGTT CAAAAGATAA 1500 TTAGTAAAAT TAATTAAATA TGTGATGCTA TTGAGTTATA GAGAGTTATT GTAAATTTAC 1560 TTAAAATCAT ACAAATCTTA TCCTAATTTA ACTTATCATT TAAGAAATAC AAAAGTAAAA 1620 AACGCGGAAA GCAATAATTT ATTTACCTTA TTATAACTCC TATATAAAGT ACTCTGTTTA 1680 TTCAACATAA TCTTACGTTG TTGTATTCAT AGGCATCTTT AACCTATCTT TTCATTTTCT 1740 GATCTCGATC GTTTTCGATC CAACAAAATG AGTCTACCGG TGAGGAACCA AGAGGTGATT 1800 ATGCAGATTC CTTCTTCTTC TCAGTTTCCA GCAACATCGA GTCCGGAAAA CACCAATCAA 1860 GTGAAGGATG AGCCAAATTT GTTTAGACGT GTTATGAATT TGCTTTTACG TCGTAGTTAT 1920 TGAAAAAGCT GATTTATCGC ATGATTCAGA ACGAGAAGTT GAAGGCAAAT AACTAAAGAA 1980 GTCTTTTATA TGTATACAAT AATTGTTTTT AAATCAAATC CTAATTAAAA AAATATATTC 2040 ATTATGACTT TCATGTTTTT AATGTAATTT ATTCCTATAT CTATAATGAT TTTGTTGTGA 2100 AGAGCGTTTT CATTTGCTAT AGAACAAGGA GAATAGTTCC AGGAAATATT CGACTTGATT 2160 TAATTATAGT GTAAACATGC TGAACACTGA AAATTACTTT TTCAATAAAC GAAAAATATA 2220 ATATACATTA CAAAACTTAT GTGAATAAAG CATGAAACTT AATATACGTT CCCTTTATCA 2280 TTTTACTTCA AAGAAAATAA ACAGAAATGT AACTTTCACA TGTAAATCTA ATTCTTAAAT 2340 TTAAAAAATA ATATTTATAT ATTTATATGA AAATAACGAA CCGGATGAAA AATAAATTTT 2400 ATATATTTAT ATCATCTCCA AATCTAGTTT GGTTCAGGGG CTTACCGAAC CGGATTGAAC 2460 TTCTCATATA CAAAAATTAG CAACACAAAA TGTCTCCGGT ATAAATACTA ACATTTATAA 2520 CCCGAACCGG TTTAGCTTCC TGTTATATCT TTTTAAAAAA GATCTCTGAC AAAGATTCCT 2580 TTCCTGGAAA TTTACCGGTT TTGGTGAAAT GTAAACCGTG GGACGAGGAT GCTTCTTCAT 2640 ATCTCACCAC CACTCTCGTT GACTTGACTT GGCTCTGCTC GTCAATGGTT ATCTTCGATC 2700 TTTAACCAAA TCCAGTTGAT AAGGTCTCTT CGTTGATTAG CAGAGATCTC TTTAATTTGT 2760 GAATTTCAAT TCATCGGAAC CTGTTG ATG GAC ACC ACC ATT GAT GGA TTC GCC 2813 Met Asp Thr Thr He Asp Gly Phe Wing 1 5 GAT TCT TAT GAA ATC AGC AGC ACT AGT TTC GTC GCT ACC GAT AAC ACC 2861 Asp Ser Tyr Glu Be Ser Ser Thr Be Phe Val Wing Thr Asp Asn Thr 10 15 20 25 GAC TCC TCT ATT GTT TAT CTG GCC GCC GAA CAA GTA CTC ACC GGA CCT 2909 Asp Ser Ser He Val Tyr Leu Ala Wing Glu Gln Val Leu Thr Gly Pro 30 35 40 GAT GTA TCT GCT CTG CAA TTG CTC TCC AAC AGC TTC GAA TCC GTC TTT 2957 Asp Val Ser Wing Leu Gln Leu Leu Ser Asn Ser Phe Glu Ser Val Phe 45 50 55 GAC TCG CCG GAT GAT TTC TAC AGC GAC GCT AAG CTT GTT CTC TCC GAC 3005 Asp Ser Pro Asp Asp Phe Tyr Ser Asp Ala Lys Leu Val Leu Ser Asp 60 65 70 GGC CGG GAA GTT TCT TTC CAC CGG TGC GTT TTG TCA GCG AGA AGC TCT 3053 Gly Arg Glu Val Ser Phe His Arg Cys Val Leu Ser Ala Arg Ser Ser 75 80 85 TTC TTC AAG AGC GCT TTA GCC GCC GCT AAG AAG GAG AAA GAC TCC AAC 3101 Phe Phe Lys Ser Ala Leu Ala Ala Ala Lys Glu Lys Asp Ser Asn 90 95 100 105 AAC ACC GCC GCC GTG AAG CTC GAG CTT AAG GAG ATT GCC AAG GAT TAC 3149 Asn Thr Wing Wing Val Lys Leu Glu Leu Lys Glu He Wing Lys Asp Tyr 110 115 120 GAA GTC GGT TTC GAT TCG GTT GTG ACT GTT TTG GCT TAT GTT TAC AGC 3197 Glu Val Gly Phe Asp Ser Val Val Thr Val Leu Ala Tyr Val Tyr Ser 125 130 135 AGC AGA GTG AGA CCG CCG CCT AAA GGA GTT TCT GAA TGC GAC GAC GAG 3245 Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser Glu Cys Wing Asp Glu 140 145 150 AAT TGC TGC CAC GTG GCT TGC CGG CCG GCG GTG GAT TTC ATG TTG GAG 3293 Asn Cys Cys His Val Wing Cys Arg Pro Wing Val Asp Phe Met Leu Glu 155 160 165 GTT CTC TAT TTG GCT TTC ATC TTC AAG ATC CCT GAA TTA ATT ACT CTC 3341 Val Leu Tyr Leu Wing Phe He Phe Lys He Pro Glu Leu He Thr Leu 170 175 180 185 TAT CAG GTAAAACACC ATCTGCATTA AGCTATGGTT ACACATTCAT GAATATGTTC 3397 Tyr Gln TTACTTGAGT ACTTGTATTT GTATTTCAG AGG CAC TTA TTG GAC GTT GTA GAC 3450 Arg His Leu Leu Asp Val Val Asp 190 195 AAA GTT GTT ATA GAG GAC ACA TTG GTT ATA CTC AAG CTT GCT AAT ATA 3498 Lys Val Val He Glu Asp Thr Leu Val He Leu Lys Leu Ala Asn He 200 205 210 TGT GGT AAA GCT TGT ATG AAG CTA TTG GAT AGA TGT AAA GAG ATT ATT 3546 Cys Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He 215 220 225 GTC AAG TCT AAT GTA GAT ATG GTT AGT CTT GAA AAG TCA TTG CCG GAA 3594 Val Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu 230 235 240 GAG CTT GTT AAA GAG ATA ATT GAT AGA CGT AAA GAG CTT GGT TTG GAG 3642 Glu Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu 245 250 255 GTA CCT AAA GTA AAG AAA CAT GTC TCG AAT GTA CAT AAG GCA CTT GAC 3690 Val Pro Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp 260 265 270 275 TCG GAT GAT ATT GAG TTA GTC AAG TTG CTT TTG AAA GAG GAT CAC ACC 3738 Ser Asp Asp He Glu Leu Val Lys Leu Leu Leu Lys Glu Asp His Thr 280 285 290 AAT CTA GAT GAT GCG TGT GCT CTT CAT TTC GCT GTT GCA TAT TGC AAT 3786 Asn Leu Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn 295 300 305 GTG AAG ACC GCA ACA GAT CTT TTA AAA CTT GAT CTT GCC GAT GTC AAC 3834 Val Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn 310 315 320 CAT AGG AAT CCG AGG GGA TAT ACG GTG CTT CAT GTT GCT GCG ATG CGG 3882 His Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg 325 330 335 AAG GAG CCA CAA TTG ATA CTA TCT CTA TTG GAA AAA GGT GCA AGT GCA 3930 Lys Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Ala Ser Wing 340 345 350 355 TCA GAA GCA ACT TTG GAA GGT AGA ACC GCA CTC ATG ATC GCA AAA CAA 3978 Ser Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He Wing Lys Gln 360 365 370 GCC 'ACT ATG GCG GTT GAA TGT AAT AAT ATC CCG GAG CA TGC AAG CAT 4026 Wing Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln Cys Lys His 375 380 385 TCT CTC AAA GGC CGA CTA TGT GTA GAA ATA CTA GAG CAA GAA GAC AAA 4074 Ser Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln Glu Asp Lys 390 395 400 CGA GAA CAA ATT CCT AGA GAT GTT CCT CCC TCT TTT GCA GTG GCG GCC 4122 Arg Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Ala Val Ala Ala 405 410 415 GAT GAA TTG AAG ATG ACG CTG CTC GAT CTT GAA AAT AGA G 4162 Asp Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg 420 425 430 GTATCTATCA AGTCTTATTT CTTATATGTT TGAATTAAAT TTATGTCCTC TCTATTAGGA 4222 AACTGAGTGA ACTAATGATA ACTATTCTTT GTGTCGTCCA CTGTTTAG TT GCA CTT 4278 Val Ala Leu 435 GCT CA CT CGT CTT TTT CCA ACG GAA GCA CA GCT GCA ATG GAG ATC GCC 4326 Wing Gln Arg Leu Phe Pro Thr Glu Wing Gln Wing Wing Met Glu He Wing 440 445 450 GAA ATG AAG GGA ACA TGT GAG TTC ATA GTG ACT AGC CTC GAG CCT GAC 4374 Glu Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu Glu Pro Asp 455 460 465 CGT CTC ACT GGT ACG AAG AGA ACA TCA CCG GGT GTA AAG ATA GCA CCT 4422 Arg Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys He Wing Pro 470 475 480 TTC AGA ATC CTA GAA GAG CAT CA AGT AGA CTA AAA GCG CTT TCT AAA 4470 Phe Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Ala Leu Ser Lys 485 490 495 ACC G GTATGGATTC TCACCCACTT CATCGGACTC CTTATCACAA AAAACAAAAC 4524 Thr 500 TAAATGATCT TTAAACATGG TTTTGTTACT TGCTGTCTGA CCTTGTTTTT TTTATCATCA 4584 G TG GAA CTC GGG AAA CGA TTC TTC CCG CGC TGT TCG GCA GTG CTC 4629 Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Val Val Leu 505 510 515 GAC CAG ATT ATG AAC TGT GAG GAC TTG ACT CA CTG GCT TGC GGA GAA 4677 Asp Gln He Met Asn Cys Glu Asp Leu Thr Gln Leu Ala Cys Gly Glu 520 525 530 GAC GAC ACT GCT GAG AAA CGA CTA CAA AAG AAG CA AGG TAC ATG GAA 4725 Asp Asp Thr Ala Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr Met Glu 535 540 545 ATA CAA GAG ACA CTA AAG AAG GCC TTT AGT GAG GAC AAT TTG GAA TTA 4773 He Gln Glu Thr Leu Lys Lys Wing Phe Ser Glu Asp Asn Leu Glu Leu 550 555 560 GGA AAT TCG TCC CTG ACA GAT TCG ACT TCT TCC ACA TCG AAA TCA ACC 4821 Gly Asn Being Ser Leu Thr Asp Ser Thr Ser Ser Thr Ser Lys Ser Thr 565 570 575 GGT GGA AAG AGG TCT AAC CGT AAA CTC TCT CAT CGT CGT CGG TGA 4866 Gly Gly Lys Arg Ser Asn Arg Lys Leu Ser His Arg Arg Arg * 580 585 590 GACTCTTGCC TCTTAGTGTA ATTTTTTGCTG TACCATATAA TTCTGTTTTC ATGATGACTG 4926 TAACTGTTTA TGTCTATCGT TGGCGTCATA TAGTTTCGCT CTTCGTTTTG CATCCTGTGT 4986 ATTATTGCTG CAGGTGTGCT TCAAACAAAT GTTGTAACAA TTTGAACCAA TGGTATACAG 5046 ATTTGTAATA TATATTTATG TACATCAACA ATAACCCATG ATGGTGTTAC AGAGTTGCTA 5106 GAATCAAAGT GTGAAATAAT GTCAAATTGT TCATCTGTTG GATATTTTCC ACCAAGAACC 5166 AAAAGAATAT TCAAGTTCCC TGAACTTCTG GCAACATTCA TGTTATATGT ATCTTCCTAA 5226 TTCTTCCTTT AACCTTTTGT AACTCGAATT ACACAGCAAG TTAGTTTCAG GTCTAGAGAT 5286 AAGAGAACAC TGAGTGGGCG TGTAAGGTGC ATTCTCCTAG TCAGCTCCAT TGCATCCAAC 5346 ATTTGTGAAT GACACAAGTT AACAATCCTT TGCACCATTT CTGGGTGCAT ACATGGAAAC 5406 TTCTTCGATT GAAACTTCCC ACATGTGCAG GTGCGTTCGC TGTCACTGAT AGACCAAGAG 5466 ACTGAAAGCT TTCACAAATT GCCCTCAAAT CTTCTGTTTC TATCGTCATG ACTCCATATC 5526 TCCGACCACT GGTCATGAGC CAGAGCCCAC TGATTTTGAG GGAATTGGGC TAACCATTTC 5586 CGAGCTTCTG AGTCCTTCTT TTTGATGTCC TTTATGTAGG AATCAAATTC TTCCTTCTGA 5646 CTTGTGGAT 5655 (2) INFORMATION FOR SEQ ID NO: 2: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 594 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE SEQ ID NO: 2: Met Asp Thr Thr He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser 1 5 10 15 Thr Ser Phe Val Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu 20 25 30 Ala Ala Glu Gln Val Leu Thr Gly Pro Asp Val Ser Ala Leu Gln Leu 35 40 45 Leu Ser Asn Ser Phe Glu Ser Val Phe Asp Ser Pro Asp Asp Phe Tyr 50 55 60 Be Asp Ala Lys Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His 65 70 75 80 Arg Cys Val Leu Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala 85 90 95 Ala Ala Lys Lys Glu Lys Asp Ser Asn Asn Thr Ala Ala Val Lys Leu 100 105 110 Glu Leu Lys Glu He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val 115 120 125 Val Thr Val Leu Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro 130 135 140 Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys 145 150 155 160 Arg Pro Wing Val Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He 165 170 175 Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp 180 185 190 Val Val Asp Lys Val Val He Glu Asp Thr Leu Val He Leu Lys Leu 195 200 205 Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys 210 215 220 Glu He He Val Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser 225 230, 235 240 Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu 245 250 255 Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser Asn Val His Lys 260 265 270 Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu Leu Leu Lys Glu 275 280 285 Asp His Thr Asn Leu Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala 290 295 300 Tyr Cys Asn Val Lys Thr Ala Thr Asp Leu Leu Lys Leu Asp Leu Ala 305 310 315 320 Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala 325 330 335 Wing Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly 340 345 350 Wing Being Wing Being Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He 355 360 365 Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln 370 375 380 Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln 385 390 395 400 Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Wing 405 410 415 Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg 420 425 430 Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala Gln Ala Ala Met 435 440 445 Glu He Wing Glu Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu 450 455 460 Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys 465 470 475 480 He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Wing 485 490 495 Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser 500 505 510 Ala Val Leu Asp Gln He Met Asn Cys Glu Asp Leu Thr Gln Leu Ala 515 520 525 Cys Gly Glu Asp Asp Thr Ala Glu Lys Arg Leu Gln Lys Lys Gln Arg 530 535 540 Tyr Met Glu He Gln Glu Thr Leu Lys Lys Wing Phe Ser Glu Asp Asn 545 550 555 560 Leu Glu Leu Gly Asn Be Ser Leu Thr Asp Ser Thr Ser Ser Thr Ser 565 570 575 Lys Ser Thr Gly Gly Lys Arg Ser Asn Arg Lys Leu Ser His Arg Arg 580 585 590 Arg * INFORMATION FOR SEQ ID NO: 3: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 314 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) ) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE SEQ ID NO: 3 Met Phe Gln Pro Wing Gly His Gly Gln Asp Trp Wing Met Glu Gly Pro 1 5 10 15 Arg Asp Gly Leu Lys Lys Glu Arg Leu Val Asp Asp Arg His Asp Ser 25 30 Gly Leu Asp Ser Met Lys Asp Glu Glu Tyr Glu Gln Met Val Lys Glu 35 40 45 Leu Arg Glu He Arg Leu Gln Pro Gln Glu Pro Wing Leu Ala Wing Glu 50 55 60 Pro Trp Lys Gln Gln Leu Thr Glu Asp Gly Asp Ser Phe Leu His Leu 65 70 75 80 Wing He He His Glu Glu Lys Pro Leu Thr Met Glu Val He Gly Gln 85 90 95 Val Lys Gly Asp Leu Wing Phe Leu Asn Phe Gln Asn Asn Leu Gln Gln 100 105 110 Thr Pro Leu His Leu Wing Val He Thr Asn Gln Pro Gly He Wing Glu 115 120 125 Ala Leu Leu Lys Ala Gly Cys Asp Pro Glu Leu Arg Asp Phe Arg Gly 130 135 140 Asn Thr Pro Leu His Leu Wing Cys Glu Gln Gly Cys Leu Wing Ser Val 145 150 155 160 Wing Val Leu Thr Gln Thr Cys Thr Pro Gln His Leu His Ser Val Leu 165 170 175 Gln Ala Thr Asn Tyr Asn Gly His Thr Cys Leu His Leu Wing Ser Thr 180 185 190 His Gly Tyr Leu Wing He Val Glu His Leu Val Thr Leu Gly Wing Asp 195 200 205 Val Asn Ala Gln Glu Pro Cys Asn Gly Arg Thr Ala Leu His Leu Ala 210 215 220 Val Asp Leu Gln Asn Pro Asp Leu Val Ser Leu Leu Leu Lys Cys Gly 225 230 235 240 Wing Asp Val Asn Arg Val Thr Tyr Gln Gly Tyr Ser Pro Tyr Gln Leu 245 250 255 Thr Trp Gly Arg Pro Be Thr Arg He Gln Gln Gln Leu Gly Gln Leu 260 265 270 Thr Leu Glu Asn Leu Gln Met Leu Pro Glu Ser Glu Asp Glu Glu Ser 275 280 285 Tyr Asp Thr Glu Ser Glu Phe Thr Glu Asp Glu Leu Pro Tyr Asp Asp 290 295 300 Cys Val Phe Gly Gly Gln Arg Leu Thr Leu 305 310 INFORMATION FOR SEQ ID NO: 4: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 314 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 4 Met Phe Gln Pro Wing Gly His Gly Gln Asp Trp Wing Met Glu Gly Pro 1 5 10 15 Arg Asp Gly Leu Lys Lys Glu Arg Leu Val Asp Asp Arg His Asp Ser 20 25 30 Gly Leu Asp Ser Met Lys Asp Glu Asp Tyr Glu Gln Met Val Lys Glu 35 40 45 Leu Arg Glu He Arg Leu Gln Pro Gln Glu Pro Wing Leu Ala Wing Glu 50 55 60 Pro Trp Lys Gln Gln Leu Thr Glu Asp Gly Asp Ser Phe Leu His Leu 65 70 75 80 Wing He He His Glu Glu Lys Thr Leu Thr Met Glu Val He Gly Gln 85 90 95 Val Lys Gly Asp Leu Wing Phe Leu Asn Phe Gln Asn Asn Leu Gln Gln 100 105 110 Thr Pro Leu His Leu Wing Val He Thr Asn Gln Pro Gly He Wing Glu 115 120 125 Wing Leu Leu Lys Wing Gly Cys Asp Pro Glu Leu Arg Asp Phe Arg Gly 130 135 140 Asn Thr Pro Leu His Leu Wing Cys Glu Gln Gly Cys Leu Wing Ser Val 145. 150 155 160 Wing Val Leu Thr Gln Thr Cys Thr Pro Gln His Leu His Ser Val Leu 165 170 175 Gln Ala Thr Asn Tyr Asn Gly His Thr Cys Leu His Leu Ala Ser He 180 185 190 His Gly Tyr Leu Gly He Val Glu His Leu Val Thr Leu Gly Ala Asp 195 200 205 Val Asn Ala Gln Glu Pro Cys Asn Gly Arg Thr Ala Leu His Leu Ala 210 215 220 Val Asp Leu Gln Asn Pro Asp Leu Val Ser Leu Leu Leu Lys Cys Gly 225 230 235 240 Wing Asp Val Asn Arg Val Thr Tyr Gln Gly Tyr Ser Pro Tyr Gln Leu 245 250 255 Thr Trp Gly Arg Pro Be Thr Arg He Gln Gln Gln Leu Gly Gln Leu 260 265 270 Thr Leu Glu Asn Leu Gln Thr Leu Pro Glu Ser Glu Asp Glu Glu Ser 275 280 285 Tyr Asp Thr Glu Ser Glu Phe Thr Glu Asp Glu Leu Pro Tyr Asp Asp 290 295 300 Cys Val Phe Gly Gly Gln Arg Leu Thr Leu 305 310 (2) INFORMATION FOR SEQ ID NO: 5: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 314 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant ( ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 5 Met Phe Gln Pro Wing Glu Pro Gly Gln Glu Trp Wing Met Glu Gly Pro 1 5 10 15 Arg Asp Ala Leu Lys Lys Glu Arg Leu Leu Asp Asp Arg His Asp Ser 20 25 30 Gly Leu Asp Ser Met Lys Asp Glu Glu Tyr Glu Gln Met Val Lys Glu 35 40 45 Leu Arg Glu He Arg Leu Glu Pro Gln Glu Pro Wing Arg Gly Ala Glu 50 55 60 Pro Trp Lys Gln Gln Leu Thr Glu Asp Gly Asp Ser Phe Leu His Leu 65 70 75 80 Ala He He His Glu Glu Lys Ala Leu Thr Met Glu Val Val Arg Gln 85 90 95 Val Lys Gly Asp Leu Wing Phe Leu Asn Phe Gln Asn Asn Leu Gln Gln 100 105 110 Thr Pro Leu His Leu Wing Val He Thr Asn Gln Pro Glu He Wing Glu 115 120 125 Ala Leu Leu Glu Ala Gly Cys Asp Pro Glu Leu Arg Asp Phe Arg Gly 130 135 140 Asn Thr Pro Leu His Leu Wing Cys Glu Gln Gly Cys Leu Wing Ser Val 145 150 155 160 Gly Val Leu Thr Gln Pro Arg Gly Thr Gln His Leu His Ser He Leu 165 170 175 Gln Ala Thr Asn Tyr Asn Gly His Thr Cys Leu His Leu Ala Ser He 180 185 190 His Gly Tyr Leu Gly He Val Glu Leu Leu Val Ser Leu Gly Ala Asp 135 200 '205 Val Asn Ala Gln Glu Pro Cys Asn Gly Arg Thr Ala Leu His Leu Ala 210 215 220 Val Asp Leu Gln Asn Pro Asp Leu Val Ser Leu Leu Leu Lys Cys Gly 225 230 235 240 Wing Asp Val Asn Arg Val Thr Tyr Gln Gly Tyr Ser Pro Tyr Gln Leu 245 250 255 Thr Trp Gly Arg Pro Be Thr Arg He Gln Gln Gln Leu Gly Gln Leu 260 265 270 Thr Leu Glu Asn Leu Gln Met Leu Pro Glu Ser Glu Asp Glu Glu Ser 5 280 285 Tyr Asp Thr Glu Ser Glu Phe Thr Glu Asp Glu Leu Pro Tyr Asp Asp 9 295 300 Cys Val Leu Gly Gly Gln Arg Leu Thr Leu 305 3io (2) INFORMATION FOR SEQ ID NO: 6: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 2011 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: cDNA (vi) ORIGINAL SOURCE: (A) ORGANISMS: Arabidopsis thaliana (ix) CHARACTERISTICS: (A) NAME / KEY: various_ characteristics (B) LOCATION: 1..2011 (D) OTHER INFORMATION: / note = "NIM1 cDNA sequence" (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 43. . 1824 (D) OTHER INFORMATION: / product = "protein NIM1" (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 6: GATCTCTTTA ATTTGTGAAT TTCAATTCAT CGGAACCTGT TG ATG GAC ACC ACC 54 Met Asp Thr Thr 1 ATT GAT GGA TTC GCC GAT TCT TAT GAA ATC AGC AGC ACT AGT TTC GTC 102 He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser Thr Ser Phe Val 5 10 15 20 GCT ACC GAT AAC ACC GAC TCC TCT ATT GTT TAT CTG GCC GCC GAA CAA 150 Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu Ala Wing Glu Gln 25 30 35 GTA CTC ACC GGA CCT GAT GTA TCT GCT CTG CAA TTG CTC TCC AAC AGC 198 Val Leu Thr Gly Pro Asp Val Ser Wing Leu Gln Leu Leu Ser Asn Ser 40 45 50 TTC GAA TCC GTC TTT GAC TCG CCG GAT GAT TTC TAC AGC GAC GCT AAG 246 Phe Glu Ser Val Phe Asp Ser Pro Asp Asp Phe Tyr Ser Asp Ala Lys 55 60 65 CTT GTT CTC TCC GAC GGC CGG GAA GTT TCT TTC CAC CGG TGC GTT TTG 294 Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His Arg Cys Val Leu 70 75 80 TCA GCG AGA AGC TCT TTC TTC AAG AGC GCT TTA GCC GCC GCT AAG AAG 342 Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala Ala Ala Lys Lys 85 90 95 100 GAG AAA GAC TCC AAC AAC ACC GCC GCC GTG AAG CTC GAG CTT AAG GAG 390 Glu Lys Asp Ser Asn Asn Thr Wing Wing Val Lys Leu Glu Leu Lys Glu 105 110 115 ATT GCC AAG GAT TAC GAA GTC GGT TTC GAT TCG GTT GTG ACT GTT TTG 438 He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val Val Thr Val Leu 120 125 130 GCT TAT GTT TAC AGC AGC AGA GTG AGA CCG CCG CCT AAA GGA GTT TCT 486 Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser 135 140 145 GAA TGC GAC GAC GAG AAT TGC TGC CAC GTG GCT TGC CGG CCG GCG GTG 534 Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys Arg Pro Wing Val 150 155 160 GAT TTC ATG TTG GAG GTT CTC TAT TTG GCT TTC ATC TTC AAG ATC CCT 582 Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He Phe Lys He Pro 165 170 175 180 GAA TTA ATT ACT CTC TAT CAG AGG CAC TTA TTG GAC GTT GTA GAC AAA 630 Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp Val Val Asp Lys 185 190 195 GTT GTT ATA GAG GAC ACA TTG GTT ATA CTC AAG CTT GCT AAT ATA TGT 678 Val Val He Glu Asp Thr Leu Val He Leu Lys Leu Ala Asn He Cys 200 205 210 GGT AAA GCT TGT ATG AAG CTA TTG GAT AGA TGT AAA GAG ATT ATT GTC 726 Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He Val 215 220 225 AAG TCT AAT GTA ATG GTT AGT CTT GAA AAG TCA TTG CCG GAA GAG 774 Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu Glu 230 235 240 CTT GTT AAA GAG ATA ATT GAT AGA CGT AAA GAG CTT GGT TTG GAG GTA 822 Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu Val 245 250 255 260 CCT AAA GTA AAG AAA CAT GTC TCG AAT GTA CAT AAG GCA CTT GAC TCG 870 Pro Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp Ser 265 270 275 GAT GAT ATT GAG TTA GTC AAG TTG CTT TTG AAA GAG GAT CAC ACC AAT 918 Asp Asp He Glu Leu Val Lys Leu Leu Leu Glu Asp His Thr Asn 280 285 290 CTA GAT GAT GCG TGT GCT CTT CAT TTC GCT GTT GCA TAT TGC AAT GTG 966 Leu Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn Val 295 300 305 AAG ACC GCA ACA GAT CTT TTA AAA CTT GAT CTT GCC GAT GTC AAC CAT 1014 Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn His 310 315 320 AGG AAT CCG AGG GGA TAT ACG GTG CTT CAT GTT GCT GCG ATG CGG AAG 1062 Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg Lys 325 330 335 340 GAG CCA CAA TTG ATA CTA TCT CTA TTG GAA AAA GGT GCA AGT GCA TCA 1110 Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Wing Ser Ala Ser 345 350 355 GAA GCA ACT TTG GAA GGT AGA ACC GCA CTC ATG ATC GCA AAA CAA GCC 1158 Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He Wing Lys Gln Wing 360 365 370 ACT ATG GCG GTT GAA TGT AAT AAT ATC CCG GAG CA TGC AAG CAT TCT 1206 Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln Cys Lys His Ser 375 380 385 CTC AAA GGC CGA CTA TGT GTA GAA ATA CTA GAG CA GAA GAC AAA CGA 1254 Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln Glu Asp Lys Arg 390 395 400 GAA CAA ATT CCT AGA GAT GTT CCT CCC TCT TTT GCA GTG GCG GCC GAT 1302 Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Ala Val Ala Ala Asp 405 410 415 420 GAA TTG AAG ATG ACG CTG CTC GAT CTT GAA AAT AGA GTT GCA CTT GCT 1350 Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg Val Ala Leu Ala 425 430 435 CAA CGT CTT TTT CCA ACG GAA GCA CA CA GCT GCA ATG GAG ATC GCC GAA 1398 Gln Arg Leu Phe Pro Thr Glu Wing Gln Wing Wing Met Glu He Wing Glu 440 445 450 ATG AAG GGA ACA TGT GAG TTC ATA GTG ACT AGC CTC GAG CCT GAC CGT 1446 Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu Glu Pro Asp Arg 455 460 465 CTC ACT GGT ACG AAG AGA ACA TCA CCG GGT GTA AAG ATA GCA CCT TTC 1494 Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys He Wing Pro Phe 470 475 480 AGA ATC CTA GAA GAG CAT CA AGT AGA CTA AAA GCG CTT TCT AAA ACC 1542 Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Ala Leu Ser Lys Thr 485 490 495 500 GTG GAA CTC GGG AAA CGA TTC TTC CCG CGC TGT TCG GCA GTG CTC GAC 1590 Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Wing Val Leu Asp 505 510 515 CAG ATT ATG AAC TGT GAG GAC TTG ACT CA CTG GCT TGC GGA GAA GAC 1638 Gln He Met Asn Cys Glu Asp Leu Thr Gln Leu Ala Cys Gly Glu Asp 520 525 530 GAC ACT GCT AAA CGA CTA CAA AAG AAG CA AGG TAC ATG GAA ATA 1686 Asp Thr Ala Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr Met Glu He 535 540 545 CAA GAG ACA CTA AAG AAG GCC TTT AGT GAG GAC AAT TTG GAA TTA GGA 1734 Gln Glu Thr Leu Lys Lys Wing Phe Ser Glu Asp Asn Leu Glu Leu Gly 550 555 560 AAT TTG TCC CTG ACA GAT TCG ACT TCT TCC ACA TCG AAA TCA ACC GGT 1782 Asn Leu Ser Leu Thr Asp Ser Thr Ser Ser Thr Ser Lys Ser Thr Gly 565 570 575 580 GGA AAG AGG TCT AAC CGT AAA CTC TCT CAT CGT CGT CGG TGA 1824 Gly Lys Arg Ser Asn Arg Lys Leu Ser His Arg Arg Arg * 585 590 GACTCTTGCC TCTTAGTGTA ATTTTTTGCTG TACCATATAA TTCTGTTTTC ATGATGACTG 1884 TAACTGTTTA TGTCTATCGT TGGCGTCATA TAGTTTCGCT CTTCGTTTTG CATCCTGTGT 1944 ATTATTGCTG CAGGTGTGCT TCAAACAAAT GTTGTAACAA TTTGAACCAA TGGTATACAG 2004 ATTTGTA 2011 (2) INFORMATION FOR SEQ ID NO: 7: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 2011 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY : linear (ii) TYPE OF MOLECULE: cDNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 43..1824 (D) OTHER INFORMATION: / product = "altered form of NIM1". / note = "the serine residues in the positions of amino acids 55 and 59 in the product of the wild type NIM1 gene have changed to Alanine residues" (ix) FEATURES: (A) NAME / KEY: characteristic_varies (B) LOCATION: 205..217 (D) OTHER INFORMATION: / note = "nucleotides 205 and 217 changed from T to G, compared to wild-type sequence" (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 7: GATCTCTTTA ATTTGTGAAT TTCAATTCAT CGGAACCTGT TG ATG GAC ACC ACC 54 Met Asp Thr Thr 1 ATT GAT GGA TTC GCC GAT TCT TAT GAA ATC AGC AGC ACT AGT TTC GTC 102 He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser Thr Ser Phe Val 5 10 15 20 GCT ACC GAT AAC ACC GAC TCC TCT ATT GTT TAT CTG GCC GCC GAA CAA 150 Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu Ala Wing Glu Gln 25 30 35 GTA CTC ACC GGA CCT GAT GTA TCT GCT CTG CAA TTG CTC TCC AAC AGC 198 Val Leu Thr Gly Pro Asp Val Ser Wing Leu Gln Leu Leu Ser Asn Ser 40 45 50 TTC GAA GCC GTC TTT GAC GCG CCG GAT GAT TTC TAC AGC GAC GCT AAG 246 Phe Glu Ala Val Phe Asp Ala Pro Asp Asp Phe Tyr Ser Asp Ala Lys 55 60 65 CTT GTT CTC TCC GAC GGC CGG GAA GTT TCT TTC CAC CGG TGC GTT TTG 294 Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His Arg Cys Val Leu 70 75 80 TCA GCG AGA AGC TCT TTC TTC AAG AGC GCT TTA GCC GCC GCT AAG AAG 342 Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala Ala Ala Lys Lys 85 90 95 100 GAG AAA GAC TCC AAC AAC ACC GCC GCC GTG AAG CTC GAG CTT AAG GAG 390 Glu Lys Asp Ser Asn Asn Thr Wing Wing Val Lys Leu Glu Leu Lys Glu 105 110 115 ATT GCC AAG GAT TAC GAA GTC GGT TTC GAT TCG GTT GTG ACT GTT TTG 438 He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val Val Thr Val Leu 120 125 130 GCT TAT GTT TAC AGC AGC AGA GTG AGA CCG CCG CCT AAA GGA GTT TCT 486 Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser 135 140 145 GAA TGC GAC GAC GAG AAT TGC TGC CAC GTG GCT TGC CGG CCG GCG GTG 534 Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys Arg Pro Wing Val 150 155 160 GAT TTC ATG TTG GAG GTT CTC TAT TTG GCT TTC ATC TTC AAG ATC CCT 582 Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He Phe Lys He Pro 165 170 175 180 GAA TTA ATT ACT CTC TAT CAG AGG CAC TTA TTG GAC GTT GTA GAC AAA 630 Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp Val Val Asp Lys 185 190 195 GTT GTT ATA GAG GAC ACA TTG GTT ATA CTC AAG CTT GCT AAT ATA TGT 678 Val Val He Glu Asp Thr Leu Val He Leu Lys Leu Ala Asn He Cys 200 205 210 GGT AAA GCT TGT ATG AAG CTA TTG GAT AGA TGT AAA GAG ATT ATT GTC 726 Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He Val 215 220 225 AAG TCT AAT GTA ATG GTT AGT CTT GAA AAG TCA TTG CCG GAA GAG 774 Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu Glu 230 235 240 CTT GTT AAA GAG ATA ATT GAT AGA CGT AAA GAG CTT GGT TTG GAG GTA 822 Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu Val 245 250 255 260 CCT AAA GTA AAG AAA CAT GTC TCG AAT GTA CAT AAG GCA CTT GAC TCG 870 Pro Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp Ser 265 270 275 GAT GAT ATT GAG TTA GTC AAG TTG CTT TTG AAA GAG GAT CAC ACC AAT 918 Asp Asp He Glu Leu Val Lys Leu Leu Lys Glu Asp His Thr Asn 280 285 290 CTA GAT GAT GCG TGT GCT CTT CAT TTC GCT GTT GCA TAT TGC AAT GTG 966 Leu Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn Val 295 300 305 AAG ACC GCA ACA GAT CTT TTA AAA CTT GAT CTT GCC GAT GTC AAC CAT 1014 Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn His 310 315 320 AGG AAT CCG AGG GGA TAT ACG GTG CTT CAT GTT GCT GCG ATG CGG AAG 1062 Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg Lys 325 330 335 340 GAG CCA CAA TTG ATA CTA TCT CTA TTG GAA AAA GGT GCA AGT GCA TCA 1110 Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Wing Ser Ala Ser 345 350 355 GAA GCA ACT TTG GAA GGT AGA ACC GCA CTC ATG ATC GCA AAA CAA GCC 1158 Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He Wing Lys Gln Wing 360 365 370 ACT ATG GCG GTT GAA TGT AAT AAT ATC CCG GAG CA TGC AAG CAT TCT 1206 Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln Cys Lys His Ser 375 380 385 CTC AAA GGC CGA CTA TGT GTA GAA ATA CTA GAG CA GAA GAC AAA CGA 1254 Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln Glu Asp Lys Arg 390 395 400 GAA CAA ATT CCT AGA GAT GTT CCT CCC TCT TTT GCA GTG GCG GCC GAT 1302 Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Ala Val Ala Ala Asp 405 410 415 420 GAA TTG AAG ATG ACG CTG CTC GAT CTT GAA AAT AGA GTT GCA CTT GCT 1350 Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg Val Ala Leu Ala 425 430 435 CAA CGT CTT TTT CCA ACG GAA GCA CA CA GCT GCA ATG GAG ATC GCC GAA 1398 Gln Arg Leu Phe Pro Thr Glu Wing Gln Wing Wing Met Glu He Wing Glu 440 445 450 ATG AAG GGA ACA TGT GAG TTC ATA GTG ACT AGC CTC GAG CCT GAC CGT 1446 Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu Glu Pro Asp Arg 455 460 465 CTC ACT GGT ACG AAG AGA ACA TCA CCG GGT GTA AAG ATA GCA CCT TTC 1494 Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys He Wing Pro Phe 470 475 480 AGA ATC CTA GAA GAG CAT CA AGT AGA CTA AAA GCG CTT TCT AAA ACC 1542 Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Ala Leu Ser Lys Thr 485 490 495 500 GTG GAA CTC GGG AAA CGA TTC TTC CCG CGC TGT TCG GCA GTG CTC GAC 1590 Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Wing Val Leu Asp 505 510 515 CAG ATT ATG AAC TGT GAG GAC TTG ACT CA CTG GCT TGC GGA GAA GAC 1638 Gln He Met Asn Cys Glu Asp Leu Thr Gln Leu Ala Cys Gly Glu Asp 520 525 530 GAC ACT GCT AAA CGA CTA CAA AAG AAG CA AGG TAC ATG GAA ATA 1686 Asp Thr Ala Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr Met Glu He 535 540 545 CAA GAG ACA CTA AAG AAG GCC TTT AGT GAG GAC AAT TTG GAA TTA GGA 1734 Gln Glu Thr Leu Lys Lys Wing Phe Ser Glu Asp Asn Leu Glu Leu Gly 550 555 560 AAT TTG TCC CTG ACA GAT TCG ACT TCT TCC ACA TCG AAA TCA ACC GGT 1782. Asn Leu Ser Leu Thr Asp Ser Thr Be Ser Thr Ser Lys Ser Thr Gly 565 570 575 580 GGA AAG AGG TCT AAC CGT AAA CTC TCT CAT CGT CGT CGG TGA 1824 Gly Lys Arg Ser Asn Arg Lys Leu Ser His Arg Arg Arg * 585 590 GACTCTTGCC TCTTAGTGTA ATTTTTTGCTG TACCATATAA TTCTGTTTTC ATGATGACTG 1884 TAACTGTTTA TGTCTATCGT TGGCGTCATA TAGTTTCGCT CTTCGTTTTG CATCCTGTGT 1944 ATTATTGCTG CAGGTGTGCT TCAAACAAAT GTTGTAACAA TTTGAACCAA TGGTATACAG 2004 ATTTGTA 2011 (2) INFORMATION FOR SEQ ID NO: 8 (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 594 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) ) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 8: Met Asp Thr Thr He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser 1 5 10 15 Thr Ser Phe Val Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu 20 25 30 Ala Ala Glu Gln Val Leu Thr Gly Pro Asp Vai Ser Ala Leu Gln Leu 35 40 45 Leu Ser Asn Ser Phe Glu Wing Val Phe Asp Wing Pro Asp Asp Phe Tyr 50 55 60 Be Asp Ala Lys Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His 65 70 75 80 Arg Cys Val Leu Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala 85 90 95 Ala Ala Lys Lys Glu Lys Asp Ser Asn Asn Thr Ala Ala Val Lys Leu 100 105 110 Glu Leu Lys Glu He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val 115 120 125 Val Thr Val Leu Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro 130 135 140 Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys 145 150 155 160 Arg Pro Wing Val Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He 165 170 175 Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp 180 185 190 Val Val Asp Lys Val Val He Glu Asp Thr Leu Val He Leu Lys Leu 195 200 205 Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys 210 215 220 Glu He He Val Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser 225 230 235 240 Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu 245 250 255 Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser Asn Val His Lys 260 265 270 Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu Leu Leu Lys Glu 275 280 285 Asp His Thr Asn Leu Asp Asp Wing Cys Wing Leu His Phe Wing Val Wing 290 295 300 Tyr Cys Asn Val Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing 305 310 315 320 Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala 325 330 335 Wing Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly 340 345 350 Wing Being Wing Being Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He 355 360 365 Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln 370 375 380 Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln 385 390 395 400 Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Wing 405 410 415 Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg 420 425 430 Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala Gln Ala Ala Met 435 440 445 Glu He Wing Glu Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu 450 455 460 Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys 465 470 475 480 He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Wing 485 490 495 Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser 500 505 510 Ala Val Leu Asp Gln He Met Asn Cys Glu Asp Leu Thr Gln Leu Ala 515 520 525 Cys Gly Glu Asp Asp Thr Wing Glu Lys Arg Leu Gln Lys Lys Gln Arg 530 535 540 Tyr Met Glu He Gln Glu Thr Leu Lys Lys Wing Phe Ser Glu Asp Asn 545 550 555 560 Leu Glu Leu Gly Asn Leu Ser Leu Thr Asp Ser Thr Ser Ser Thr Ser 565 570 575 Lys Ser Thr Gly Gly Lys Arg Ser Asn Arg Lys Leu Ser His Arg Arg 580 535 590 Arg * (2) INFORMATION FOR SEQ ID NO: 9: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1597 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY : linear (ii) TYPE OF MOLECULE: cDNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..1410 (D) OTHER INFORMATION: / product = "altered form of NIM1". / note = "N-terminal deletion compared to wild-type NIM1 sequence" (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 9:nd.

ATG GAT TCG GTT GTG ACT GTT TTG GCT TAT GTT TAC AGC AGC AGA GTG 48 Met Asp Ser Val Val Thr Val Leu Ala Tyr Val Tyr Ser Ser Arg Val 1 5 10 15 AGA CCG CCG CCT AAA GGA GTT TCT GAA TGC GCA GAC GAG AAT TGC TGC 96 Arg Pro Pro Pro Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys 20 25 30 CAC GTG GCT TGC CGG CCG GCG GTG GAT TTC ATG TTG GAG GTT CTC TAT 144 His Val Wing Cys Arg Pro Wing Val Asp Phe Met Leu Glu Val Leu Tyr 35 40 45 TTG GCT TTC ATC TTC AAG ATC CCT GAA TTA ATT ACT CTC TAT CAG AGG 192 Leu Wing Phe He Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg 50 55 60 CAC TTA TTG GAC GTT GTA GAC AAA GTT GTT ATA GAG GAC ACA TTG GTT 240 His Leu Leu Asp Val Val Asp Lys Val Val He Glu Asp Thr Leu Val 65 70 75 80 ATA CTC AAG CTT GCT AAT ATA TGT GGT AAA GCT TGT ATG AAG CTA TTG 288 He Leu Lys Leu Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu 85 90 95 GAT AGA TGT AAA GAG ATT ATT GTC AAG TCT AAT GTA GAT ATG GTT AGT 336 Asp Arg Cys Lys Glu He HV Val Lys Ser Asn Val Asp Met Val Ser 100 105 110 CTT GAA AAG TCA TTG CCG GAA GAG CTT GTT AAA GAG ATA ATT GAT AGA 384 Leu Glu Lys Ser Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg 115 120 125 CGT AAA GAG CTT GGT TTG GAG GTA CCT AAA GTA AAG AAA CAT GTC TCG 432 Arg Lys Glu Leu Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser 130 135 140 AAT GTA CAT AAG GCA CTT GAC TCG GAT GAT ATT GAG TTA GTC AAG TTG 480 Asn Val His Lys Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu 145 150 155 160 CTT TTG AAA GAG GAT CAC ACC AAT CTA GAT GAT GCG TGT GCT CTT CAT 528 Leu Leu Lys Glu Asp His Thr Asn Leu Asp Asp Ala Cys Ala Leu His 165 170 175 TTC GCT GTT GCA TAT TGC AAT GTG AAG ACC GCA ACA GAT CTT TTA AAA 576 Phe Wing Val Wing Tyr Cys Asn Val Lys Thr Wing Thr Asp Leu Leu Lys 180 185 190 CTT GAT CTT GCC GAT GTC GTC AAC CAT AGG AAT CCG AGG GGA TAT ACG GTG 624 Leu Asp Leu Wing Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val 195 200 205 CTT CAT GTT GCT GCG ATG CGG AAG GAG CCA CAA TTG ATA CTA TCT CTA 672 Leu His Val Ala Ala Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu 210 215 220 TTG GAA AAA GGT GCA AGT GCA TCA GAA GCA ACT TTG GAA GGT AGA ACC 720 Leu Glu Lys Gly Wing Ser Wing Ser Glu Wing Thr Leu Glu Gly Arg Thr 225. 230 235 240 GCA CTC ATG ATC GCA AAA CAA GCC ACT ATG GCG GTT GAA TGT AAT AAT 768 Wing Leu Met He Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn 245 250 255 ATC CCG GAG CA TGC AAG CAT TCT CTC AAA GGC CGA CTA TGT GTA GAA 816 He Pro Glu Gln Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu 260 265 270 ATA CTA GAG CA GAA GAC AAA CGA GAA CAA ATT CCT AGA GAT GTT CCT 864 He Leu Glu Gln Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro 275 280 285 CCC TCT TTT GCA GTG GCG GCC GAT GAA TTG AAG ATG ACG CTG CTC GAT 912 Pro Ser Phe Ala Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp 290 295 300 CTT GAA AAT AGA GTT GCA CTT GCT CAA CGT CTT TTT CCA ACG GAA GCA 960 Leu Glu Asn Arg Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala 305 310 315 320 CAQ GCT GCA ATG GAG ATC GCC GAA ATG AAG GGA ACA TGT GAG TTC ATA 1008 Gln Wing Wing Met Glu He Wing Glu Met Lys Gly Thr Cys Glu Phe He 325 330 335 GTG ACT AGC CTC GAG CCT GAC CGT CTC ACT GGT ACG AAG AGA ACA TCA 1056 Val Thr Ser Leu Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser 340 345 350 CCG GGT GTA AAG ATA GCA CCT TTC AGA ATC CTA GAA GAG CAT CAA AGT 1104 Pro Gly Val Lys He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser 355 360 365 AGA CTA AAA GCG CTT TCT AAA ACC GTG GAA CTC GGG AAA CGA TTC TTC 1152 Arg Leu Lys Ala Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe 370 375 380 CCG CGC TGT TCG GCA GTG CTC GAC CAG ATT ATG AAC TGT GAG GAC TTG 1200 Pro Arg Cys Ser Wing Val Leu Asp Gln He Met Asn Cys Glu Asp Leu 385 390 395 400 ACT CA CTG GCT TGC GGA GAA GAC GAC ACT GCT GAG AAA CGA CTA CAA 1248 Thr Gln Leu Wing Cys Gly Glu Asp Asp Thr Wing Glu Lys Arg Leu Gln 405 410 415 AAG AAG CAG AGA TAC ATG GAA ATA CAA GAG ACA CTA AAG AAG GCC TTT 1296 Lys Lys Gln Arg Tyr Met Glu He Gln Glu Thr Leu Lys Lys Wing Phe 420 425 430 AGT GAG GAC AAT TTG GAA TTA GGA AAT TTG TCC CTG ACA GAT TCG ACT 1344 Ser Glu Asp Asn Leu Glu Leu Gly Asn Leu Ser Leu Thr Asp Ser Thr 435 440 445 TCT TCC ACA TCG AAA TCA ACC GGT GGA AAG AGG TCT AAC CGT AAA CTC 1392 Ser Ser Thr Ser Lys Ser Thr Gly Gly Lys Arg Ser Asn Arg Lys Leu 450 455 460 TCT CAT CGT CGT CGG TGA GACTCTTGCC TCTTAGTGTA ATTTTTGCTG 1440 Ser His Arg Arg Arg * 465 470 TACCATATAA TTCTGTTTTC ATGATGACTG TAACTGTTTA TGTCTATCGT TGGCGTCATA 1500 TAGTTTCGCT CTTCGTTTTG CATCCTGTGT ATTATTGCTG CAGGTGTGCT TCAAACAAAT 1560 GTTGTAACAA TTTGAACCAA TGGTATACAG ATTTGTA 1597 (2) INFORMATION FOR SEQ ID NO: 10: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 470 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: linear (ii) TYPE OF MOLECULE: protein ( xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 10 Met Asp Ser Val Val Thr Val Leu Ala Tyr Val Tyr Ser Ser Arg Val 1 5 10 15 Arg Pro Pro Pro Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys 20 25 30 His Val Ala Cys Arg Pro Ala Val Asp Phe Met Leu Glu Val Leu Tyr 35 40 45 Leu Ala Phe He Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg 50 55 60 His Leu Leu Asp Val Val Asp Lys Val Val He Glu Asp Thr Leu Val 65 70 75 80 He Leu Lys Leu Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu 85 90 95 Asp Arg Cys Lys Glu He He Val Val Lys Ser Asn Val Asp Met Val Ser 100 105 110 Leu Glu Lys Ser Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg 115 120 125 Arg Lys Glu Leu Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser 130 135 140 Asn Val His Lys Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu 145 150 155 160 Leu Leu Lys Glu Asp His Thr Asn Leu Asp Asp Ala Cys Ala Leu His 165 170 175 Phe Ala Val Ala Tyr Cys Asn Val Lys Thr Ala Thr Asp Leu Leu Lys 180 185 190 Leu Asp Leu Ala Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val 195 200 205 Leu His Val Ala Ala Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu 210 215 220 Leu Glu Lys Gly Wing Ser Wing Ser Glu Wing Thr Leu Glu Gly Arg Thr 225 230 235 240 Wing Leu Met He Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn 245 250 255 He Pro Glu Gln Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu 260 265 270 He Leu Glu Gln Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro 275 280 285 Pro Ser Phe Ala Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp 290 295 300 Leu Glu Asn Arg Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala 305 310 315 320 Gln Ala Ala Met Glu He Ala Glu Met Lys Gly Thr Cys Glu Phe He 325 330 335 Val Thr Ser Leu Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser 340 345 350 Pro Gly Val Lys He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser 355 360 365 Arg Leu Lys Ala Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe 370 375 380 Pro Arg Cys Ser Ala Val Leu Asp Gln He Met Asn Cys Glu Asp Leu 385 390 395 400 Thr Gln Leu Wing Cys Gly Glu Asp Asp Thr Wing Glu Lys Arg Leu Gln 405 410 415 Lys Lys Gln Arg Tyr Met Glu He Gln Glu Thr Leu Lys Lys Wing Phe 420 425 430 Ser Glu Asp Asn Leu Glu Leu Gly Asn Leu Ser Leu Thr Asp Ser Thr 435 440 445 Ser Ser Thr Ser Lys Ser Thr Gly Gly Lys Arg Ser Asn Arg Lys Leu 450 455 460 Ser His Arg Arg Arg * 465 470 (2) INFORMATION FOR SEQ ID NO: 11: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1608 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: l ineal (ii) TYPE OF MOLECULE: cDNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCALIZATION: 43. . 1608 (D) OTHER INFORMATION: / product = "altered form of NIMl'V / note =" C-terminal deletion compared to wild-type NIM1"(i) DESCRIPTION OF SEQUENCE: SEQ ID NO: 11: GATCTCTTTA ATTTGTGAAT TTCAATTCAT CGGAACCTGT TG ATG GAC ACC ACC 54 Met Asp Thr Thr 1 ATT GAT GGA TTC GCC GAT TCT TAT GAA ATC AGC AGC ACT AGT TTC GTC 102 He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser Thr Ser Phe Val 5 10 15 20 GCT ACC GAT AAC ACC GAC TCC TCT ATT GTT TAT CTG GCC GCC GAA CAA 150 Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu Ala Wing Glu Gln 25 30 35 GTA CTC ACC GGA CCT GAT GTA TCT GCT CTG CAA TTG CTC TCC AAC AGC 198 Val Leu Thr Gly Pro Asp Val Ser Wing Leu Gln Leu Leu Ser Asn Ser 40 45 50 TTC GAA TCC TTC GTC TTG GCT TCG CCG GAT TTC TAC AGC GAC GCT AAG 246 Phe Glu Ser Val Phe Asp Ser Pro Asp Asp Phe Tyr As Asp Ala Lys 55 60 65 CTT GTT CTC TCC GAC GGC CGG GAA GTT TCT TTC CAC CGG TGC GTT TTG 294 Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His Arg Cys Val Leu 70 75 80 TCA GCG AGA AGC TCT TTC TTC AAG AGC GCT TTA GCC GCC GCT AAG AAG 342 Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala Ala Ala Lys Lys 85 90 95 100 GAG AAA GAC TCC AAC AAC ACC GCC GCC GTG AAG CTC GAG CTT AAG GAG 390 Glu Lys Asp Ser Asn Asn Thr Ala Ala Val Lys Leu Glu Leu Lys Glu 105 110 115 ATT GCC AAG GAT TAC GAA GTC GGT TTC GAT TCG GTT GTG ACT GTT TTG 438 He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val Val Thr Val Leu 120 125 130 GCT TAT GTT TAC AGC AGC AGA GTG AGA CCG CCG CCT AAA GGA GTT TCT 486 Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser 135 140 145 GAA TGC GAC GAC GAG AAT TGC TGC CAC GTG GCT TGC CGG CCG GCG GTG 534 Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys Arg Pro Wing Val 150 155 160 GAT TTC ATG TTG GAG GTT CTC TAT TTG GCT TTC ATC TTC AAG ATC CCT 582 Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He Phe Lys He Pro 165 170 175 180 GAA TTA ATT ACT CTC TAT CAG AGG CAC TTA TTG GAC GTT GTA GAC AAA 630 Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp Val Val Asp Lys 185 190 195 GTT GTT ATA GAG GAC ACA TTG GTT ATA CTC AAG CTT GCT AAT ATA TGT 678 Val Val He Glu Asp Thr Leu Val He Leu Lys Leu Ala Asn He Cys 200 205 210 GGT AAA GCT TGT ATG AAG CTA TTG GAT AGA TGT AAA GAG ATT ATT GTC 726 Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He Val 215 220 225 AAG TCT AAT GTA ATG GTT AGT CTT GAA AAG TCA TTG CCG GAA GAG 774 Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu Glu 230 235 240 CTT GTT AAA GAG ATA ATT GAT AGA CGT AAA GAG CTT GGT TTG GAG GTA 822 Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu Val 245 250 255 260 CCT AAA GTA AAG AAA CAT GTC TCG AAT GTA CAT AAG GCA CTT GAC TCG 870 Pro Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp Ser 265 270 275 GAT GAT ATT GAG TTA GTC AAG TTG CTT TTG AAA GAG GAT CAC ACC AAT 918 Asp Asp He Glu Leu Val Lys Leu Leu Lys Glu Asp His Thr Asn 280 285 290 CTA GAT GAT GCG TGT GCT CTT CAT TTC GCT GTT GCA TAT TGC AAT GTG 966 Leu Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn Val 295 300 305 AAG ACC GCA ACA GAT CTT TTA AAA CTT GAT CTT GCC GAT GTC AAC CAT 1014 Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn His 310 315 320 AGG AAT CCG AGG GGA TAT ACG GTG CTT CAT GTT GCT GCG ATG CGG AAG 1062 Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg Lys 325 330 335 340 GAG CCA CAA TTG ATA CTA TCT CTA TTG GAA AAA GGT GCA AGT GCA TCA 1110 Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Wing Ser Ala Ser 345 350 355 GAA GCA ACT TTG GAA GGT AGA ACC GCA CTC ATG ATC GCA AAA CAA GCC 1158 Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He Wing Lys Gln Wing 360 365 370 ACT ATG GCG GTT GAA TGT AAT AAT ATC CCG GAG CA TGC AAG CAT TCT 1206 Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln Cys Lys His Ser 375 380 385 CTC AAA GGC CGA CTA TGT GTA GAA ATA CTA GAG CA GAA GAC AAA CGA 1254 Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln Glu Asp Lys Arg 390 395 400 GAA CAA ATT CCT AGA GAT GTT CCT CCC TCT TTT GCA GTG GCG GCC GAT 1302 Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Ala Val Ala Ala Asp 405 410 415 420 GAA TTG AAG ATG ACG CTG CTC GAT CTT GAA AAT AGA GTT GCA CTT GCT 1350 Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg Val Ala Leu Ala 425 430 435 CAA CGT CTT TTT CCA ACG GAA GCA CA CA GCT GCA ATG GAG ATC GCC GAA 1398 Gln Arg Leu Phe Pro Thr Glu Wing Gln Wing Wing Met Glu He Wing Glu 440 445 450 ATG AAG GGA ACA TGT GAG TTC ATA GTG ACT AGC CTC GAG CCT GAC CGT 1446 Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu Glu Pro Asp Arg 455 460 465 CTC ACT GGT ACG AAG AGA ACA TCA CCG GGT GTA AAG ATA GCA CCT TTC 1494 Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys He Wing Pro Phe 470 475 480 AGA ATC CTA GAA GAG CAT CA AGT AGA CTA AAA GCG CTT TCT AAA ACC 1542 Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Ala Leu Ser Lys Thr 485 490 495 500 GTG GAA CTC GGG AAA CGA TTC TTC CCG CGC TGT TCG GCA GTG CTC GAC 1590 Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Wing Val Leu Asp 505 510 515 CAG ATT ATG AAC TGT TGA 1608 Gln He Met Asn Cys * 520 (2) INFORMATION FOR SEQ ID NO: 12: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 522 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 12 Met Asp Thr Thr He Asp Gly Phe Wing Asp Ser Tyr Glu He Ser Ser 1 5 10 15 Thr Ser Phe Val Wing Thr Asp Asn Thr Asp Ser Ser He Val Tyr Leu 20 25 30 Ala Ala Glu Gln Val Leu Thr Gly Pro Asp Val Ser Ala Leu Gln Leu 35 40 45 Leu Ser Asn Ser Phe Glu Ser Val Phe Asp Ser Pro Asp Asp Phe Tyr 50 55 60 Be Asp Ala Lys Leu Val Leu Ser Asp Gly Arg Glu Val Ser Phe His 65 70 75 80 Arg Cys Val Leu Ser Ala Arg Ser Ser Phe Phe Lys Ser Ala Leu Ala 85 90 95 Ala Ala Lys Lys Glu Lys Asp Ser Asn Asn Thr Ala Ala Val Lys Leu 100 105 110 Glu Leu Lys Glu He Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val 115 120 125 Val Thr Val Leu Wing Tyr Val Tyr Ser Ser Arg Val Arg Pro Pro Pro 130 135 140 Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys His Val Wing Cys 145 150 155 160 Arg Pro Wing Val Asp Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He 165 170 175 Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp 180 185 190 Val Val Asp Lys Val Val He Glu Asp Thr Leu Val He Leu Lys Leu 195 200 205 Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lye 210 215 220 Glu He He Val Lys Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser 225 230 235 240 Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg Arg Lys Glu Leu 245 250 255 Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser Asn Val His Lys 260 265 270 Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu Leu Leu Lys Glu 275 280 285 Asp His Thr Asn Leu Asp Asp Wing Cys Wing Leu His Phe Wing Val Wing 290 295 300 Tyr Cys Asn Val Lys Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing 305 310 315 320 Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala 325 330 335 Wing Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly 340 345 350 Wing Being Wing Being Glu Wing Thr Leu Glu Gly Arg Thr Wing Leu Met He 355 360 365 Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn He Pro Glu Gln 370 375 380 Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu He Leu Glu Gln 385 390 395 400 Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro Pro Ser Phe Wing 405 410 415 Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp Leu Glu Asn Arg 420 425 430 Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala Gln Ala Ala Met 435 440 445 Glu He Wing Glu Met Lys Gly Thr Cys Glu Phe He Val Thr Ser Leu 450 455 460 Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser Pro Gly Val Lys 465 470 475 480 He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser Arg Leu Lys Wing 485 490 495 Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser 500 505 510 Wing Val Leu Asp Gln He Met Asn Cys * 515 520 (2) INFORMATION FOR SEQ ID NO: 13: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1194 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: cDNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..1194 (D) OTHER INFORMATION: / product = "altered form of NIM1". / note = "N-terminal chimera / C-terminal" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 13: ATG GAT TCG GTT GTG ACT GTT TTG GCT TAT GTT TAC AGC AGC AGA GTG 48 Met Asp Ser Val Val Thr Val Leu Ala Tyr Val Tyr Ser Ser Arg Val 1 5 10 15 AGA CCG CCG CCT AAA GGA GTT TCT GAA TGC GCA GAC GAT AAT TGC TGC 96 Arg Pro Pro Pro Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys 20 25 30 CAC GTG GCT TGC CGG CCG GCG GTG GAT TTC ATG TTG GAG GTT CTC TAT 144 His Val Wing Cys Arg Pro Wing Val Asp Phe Met Leu Glu Val Leu Tyr 35 40 45 TTG GCT TTC ATC TTC AAG ATC CCT GAA TTA ATT ACT CTC TAT CAG AGG 192 Leu Wing Phe He Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg 50 55 60 CAC TTA TTG GAC GTT GTA GAC AAA GTT GTT ATA GAG GAC ACA TTG GTT 240 His Leu Leu Asp Val Val Asp Lys Val Val He Glu Asp Thr Leu Val 65 70 75 80 ATA CTC AAG CTT GCT AAT ATA TGT GGT AAA GCT TGT ATG AAG CTA TTG 288 He Leu Lys Leu Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu 85 90 95 GAT AGA TGT AAA GAG ATT ATT GTC AAG TCT AAT GTA GAT ATG GTT AGT 336 Asp Arg Cys Lys Glu He HV Val Lys Ser Asn Val Asp Met Val Ser 100 105 110 CTT GAA AAG TCA TTG CCG GAA GAG CTT GTT AAA GAG ATA ATT GAT AGA 384 Leu Glu Lys Ser Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg 115 120 125 CGT AAA GAG CTT GGT TTG GAG GTA CCT AAA GTA AAG AAA CAT GTC TCG 432 Arg Lys Glu Leu Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser 130 135 140 AAT GTA CAT AAG GCA CTT GAC TCG GAT GAT ATT GAG TTA GTC AAG TTG 480 Asn Val His Lys Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu 145 150 155 160 CTT TTG AAA GAG GAT CAC ACC AAT CTA GAT GAT GCG TGT GCT CTT CAT 528 Leu Leu Lys Glu Asp His Thr Asn Leu Asp Asp Ala Cys Ala Leu His 165 170 175 TTC GCT GTT GCA TAT TGC AAT GTG AAG ACC GCA ACA GAT CTT TTA AAA 576 Phe Wing Val Wing Tyr Cys Asn Val Lys Thr Wing Thr Asp Leu Leu Lys 180 185 190 CTT GAT CTT GCC GAT GTC AAC CAT AGG AAT CCG AGG GGA TAT ACG GTG 624 Leu Asp Leu Wing Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val 195 200 205 CTT CAT GTT GCT GCG ATG CGG AAG GAG CCA CAA TTG ATA CTA TCT CTA 672 Leu His Val Ala Wing Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu 210 215 220 TTG GAA AAA GGT GCA AGT GCA TCA GAA GCA ACT TTG GAA GGT AGA ACC 720 Leu Glu Lys Gly Wing Ser Wing Ser Glu Wing Thr Leu Glu Gly Arg Thr 225 230 235 240 GCA CTC ATG ATC GCA AAA CAA GCC ACT ATG GCG GTT GAA TGT AAT AAT 768 Wing Leu Met He Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn 245 250 255 ATC CCG GAG CA TGC AAG CAT TCT CTC AAA GGC CGA CTA TGT GTA GAA 816 He Pro Glu Gln Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu 260 265 270 ATA CTA GAG CAA GAA GAC AAA CGA GAA CAA ATT CCT AGA GAT GTT CCT 864 He Leu Glu Gln Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro 275 280 285 CCC TCT TTT GCA GTG GCG GCC GAT GAA TTG AAG ATG ACG CTG CTC GAT 912 Pro Ser Phe Ala Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp 290 295 300 CTT GAA AAT AGA GTT GCA CTT GCT CAA CGT CTT TTT CCA ACG GAA GCA 960 Leu Glu Asn Arg Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala 305 310 315 320 CAQ GCT GCA ATG GAG ATC GCC GAA ATG AAG GGA ACA TGT GAG TTC ATA 1008 Gln Wing Wing Met Glu He Wing Glu Met Lys Gly Thr Cys Glu Phe He 325 330 335 GTG ACT AGC CTC GAG CCT GAC CGT CTC ACT GGT ACG AAG AGA ACA TCA 1056 Val Thr Ser Leu Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser 340 345 350 CCG GGT GTA AAG ATA GCA CCT TTC AGA ATC CTA GAA GAG CAT CAA AGT 1104 Pro Gly Val Lys He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser 355 360 365 AGA CTA AAA GCG CTT TCT AAA ACC GTG GAA CTC GGG AAA CGA TTC TTC 1152 Arg Leu Lys Ala Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe 370 375 380 CCG CGC TGT TCG GCA GTG CTC GAC CAG ATT ATG AAC TGT TGA 1194 Pro Arg Cys Ser Wing Val Leu Asp Gln He Met Asn Cys * 385 390 395 (2) INFORMATION FOR SEQ ID NO: 14: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 398 amino acids (B) TYPE: amino acid (D) TOPOLOGY: l ineal (ii) TYPE OF MOLECULE: protein (xi) ) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 14: Met Asp Ser Val Val Thr Val Leu Ala Tyr Val Tyr Ser Ser Arg Val 1 5 10 15 Arg Pro Pro Pro Lys Gly Val Ser Glu Cys Wing Asp Glu Asn Cys Cys 20 25 30 His Val Ala Cys Arg Pro Ala Val Asp Phe Met Leu Glu Val Leu Tyr 35 40 45 Leu Ala Phe He Phe Lys He Pro Glu Leu He Thr Leu Tyr Gln Arg 50 55 60 His Leu Leu Asp Val Val Asp Lys Val Val He Glu Asp Thr Leu Val 65 70 75 80 He Leu Lys Leu Wing Asn He Cys Gly Lys Wing Cys Met Lys Leu Leu 85 90 - 95 Asp Arg Cys Lys Glu He He Val Val Lys Ser Asn Val Asp Met Val Ser 100 105 110 Leu Glu Lys Ser Leu Pro Glu Glu Leu Val Lys Glu He He Asp Arg 115 120 125 Arg Lys Glu Leu Gly Leu Glu Val Pro Lys Val Lys Lys His Val Ser 130 135 140 Asn Val His Lys Ala Leu Asp Ser Asp Asp He Glu Leu Val Lys Leu 145 150 155 160 Leu Leu Lys Glu Asp His Thr Asn Leu Asp Asp Ala Cys Ala Leu His 165 170 175 Phe Ala Val Ala Tyr Cys Asn Val Lys Thr Ala Thr Asp Leu Leu Lys 180 185 190 Leu Asp Leu Ala Asp Val Asn His Arg Asn Pro Arg Gly Tyr Thr Val 195 200 205 Leu His Val Ala Ala Met Arg Lys Glu Pro Gln Leu He Leu Ser Leu 210 215 220 Leu Glu Lys Gly Wing Ser Wing Ser Glu Wing Thr Leu Glu Gly Arg Thr 225 230 235 240 Wing Leu Met He Wing Lys Gln Wing Thr Met Wing Val Glu Cys Asn Asn 245 250 255 He Pro Glu Gln Cys Lys His Ser Leu Lys Gly Arg Leu Cys Val Glu 260 265 270 He Leu Glu Gln Glu Asp Lys Arg Glu Gln He Pro Arg Asp Val Pro 275 280 285 Pro Ser Phe Ala Val Ala Ala Asp Glu Leu Lys Met Thr Leu Leu Asp 290 295 300 Leu Glu Asn Arg Val Ala Leu Ala Gln Arg Leu Phe Pro Thr Glu Ala 305 310 315 320 Gln Ala Ala Met Glu He Ala Glu Met Lys Gly Thr Cys Glu Phe He 325 330 335 Val Thr Ser Leu Glu Pro Asp Arg Leu Thr Gly Thr Lys Arg Thr Ser 340 345 350 Pro Gly Val Lys He Wing Pro Phe Arg He Leu Glu Glu His Gln Ser 355 360 365 Arg Leu Lys Ala Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe 370 375 380 Pro Arg Cys Ser Wing Val Leu Asp Gln He Met Asn Cys * 385 390 395 (2) INFORMATION FOR SEQ ID NO: 15: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 786 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: cDNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..786 (D) OTHER INFORMATION: / product = "altered form of NIM1". / note = "Ankyrin domains of NIM1" (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 15: ATG GAC TCC AAC AAC ACC GCC GCC GTG AAG CTC GAG CTT AAG GAG ATT 48 Met Asp Ser Asn Asn Thr Wing Wing Val Lys Leu Glu Leu Lys Glu He 1 5 10 15 GCC AAG GAT TAC GAA GTC GGT TTC GAT TCG GTT GTG ACT GTT TTG GCT 96 Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val Val Thr Val Leu Ala 20 25 30 TAT GTT TAC AGC AGC AGA GTG AGA CCG CCG CCT AAA GGA GTT TCT GAA 144 Tyr Val Tyr Being Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser Glu 35 40 45 TGC GCA GAC GAG AAT TGC TGC CAC GTG GCT TGC CGG CCG GCG GTG GAT 192 Cys Ala Asp Glu Asn Cys Cys His Val Ala Cys Arg Pro Ala Val Asp 50 55 60 TTC ATG TTG GAG GTT CTC TAT TTG GCT TTC ATC TTC AAG ATC CCT GAA 240 Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He Phe Lys He Pro Glu 65 70 75 80 TTA ATT ACT CTC TAT CAG AGG CAC TTA TTG GAC GTT GTA GAC AAA GTT 288 Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp Val Val Asp Lys Val 85 90 95 GTT ATA GAG GAC ACA TTG GTT ATA CTC AAG CTT GCT AAT ATA TGT GGT 336 Val He Glu Asp Thr Leu Val He Leu Lys Leu Wing Asn He Cys Gly 100 105 110 AAA GCT TGT ATG AAG CTA TTG GAT AGA TGT AAA GAG ATT ATT GTC AAG 384 Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He Val Lys 115 120 125 TCT AAT GTA ATG GTT AGT CTT GAA AAG TCA TTG CCG GAA GAG CTT 432 Ser Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu Glu Leu 130 135 140 GTT AAA GAG ATA ATT GAT AGA CGT AAA GAG CTT GGT TTG GAG GTA CCT 480 Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu Val Pro 145 150 155 160 AAA GTA AAG AAA CAT GTC TCG AAT GTA CAT AAG GCA CTT GAC TCG GAT 528 Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp Ser Asp 165 170 175 GAT ATT GAG TTA GTC AAG TTG CTT TTG AAA GAG GAT CAC ACC AAT CTA 576 Asp He Glu Leu Val Lys Leu Leu Leu Glu Asp His Thr Asn Leu 180 185 190 GAT GAT GCG TGT GCT CTT CAT TTC GCT GTT GCA TAT TGC AAT GTG AAG 624 Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn Val Lys 195 200 205 ACC GCA ACA GAT CTT TTA AAA CTT GAT CTT GCC GAT GTC AAC CAT AGG 672 Thr Wing Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn His Arg 210 215 220 AAT CCG AGG GGA TAT ACG GTG CTT CAT GTT GCT GCG ATG CGG AAG GAG 720 Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg Lys Glu 225 230 235 240 CCA CA TTG ATA CTA TCT CTA TTG GAA AAA GGT GCA AGT GCA TCA GAA 768 Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Wing Ser Wing Ser Glu 245 250 255 GCA ACT TTG GAA GGT TGA 7gg Ala Thr Leu Glu Gly * 260 (2) INFORMATION FOR SEQ ID NO: 16: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 262 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 16: Met Asp Ser Asn Asn Thr Wing Wing Val Lys Leu Glu Leu Lys Glu He 1 5 10 15 Ala Lys Asp Tyr Glu Val Gly Phe Asp Ser Val Val Thr Val Leu Ala 20 25 30 Tyr Val Tyr Being Ser Arg Val Arg Pro Pro Pro Lys Gly Val Ser Glu 35 40 45 Cys Ala Asp Glu Asn Cys Cys His Val Ala Cys Arg Pro Ala Val Asp 50 55 60 Phe Met Leu Glu Val Leu Tyr Leu Wing Phe He Phe Lys He Pro Glu 65 70 75 80 Leu He Thr Leu Tyr Gln Arg His Leu Leu Asp Val Val Asp Lys Val 85 90 95 Val He Glu Asp Thr Leu Val He Leu Lys Leu Wing Asn He Cys Gly 100 105 110 Lys Wing Cys Met Lys Leu Leu Asp Arg Cys Lys Glu He He Val Lys 115 120 125 As Asn Val Asp Met Val Ser Leu Glu Lys Ser Leu Pro Glu Glu Leu 130 135 140 Val Lys Glu He He Asp Arg Arg Lys Glu Leu Gly Leu Glu Val Pro 145 150 155 160 Lys Val Lys Lys His Val Ser Asn Val His Lys Ala Leu Asp Ser Asp 165 170 175 Asp He Glu Leu Val Lys Leu Leu Leu Lys Glu Asp His Thr Asn Leu 180 185 190 Asp Asp Ala Cys Ala Leu His Phe Ala Val Ala Tyr Cys Asn Val Lys 195 200 205 Thr Ala Thr Asp Leu Leu Lys Leu Asp Leu Wing Asp Val Asn His Arg 210 215 220 Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Wing Met Arg Lys Glu 225 230 235 240 Pro Gln Leu He Leu Ser Leu Leu Glu Lys Gly Wing Ser Wing Ser Glu 245 250 255 Ala Thr Leu Glu Gly * 260 (2) INFORMATION FOR SEQ ID NO: 17: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 41 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 17 He Arg Arg Met Arg Arg Ala Leu Asp Ala Ala Asp He Glu Leu Val 1 5 10 15 Lys Leu Met Val Met Gly Glu Gly Leu Asp Leu Asp Asp Ala Leu Ala 20 25 30 Val His Tyr Ala Val Gln His Cys Asn 35 40 (2) INFORMATION FOR SEQ ID NO: 18: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 38 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant ( ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 18 Pro Thr Gly Lys Thr Ala Leu His Leu Ala Ala Glu Met Val Ser Pro 1 5 10 15 Asp Met Val Ser Val Leu Leu Asp His His Wing Asp Xaa Asn Phe Arg 20 25 30 Thr Xaa Asp Gly Val Thr 35 (2) INFORMATION FOR SEQ ID NO: 19: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 41 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 19 He Arg Arg Met Arg Arg Ala Leu Asp Ala Wing Asp He Glu Leu Val 1 5 10 15 Lys Leu Met Val Met Gly Glu Gly Leu Asp Leu Asp Asp Ala Leu Ala 20 25 30 Val His Tyr Ala Val Gln His Cys Asn 35 40 (2) INFORMATION FOR SEQ ID NO: 20: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 27 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN : not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 20: Arg Arg Pro Asp Ser Lys Thr Ala Leu His Leu Ala Ala Glu Met Val 1 5 10 15 Ser Pro Asp Met Val Ser Val Leu Leu Asp Gln 20 25 [2) INFORMATION FOR SEQ ID NO: 21: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 41 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant ( ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 21: He Arg Arg Met Arg Arg Ala Leu Asp Ala Wing Asp He Glu Leu Val 1 5 10 15 Lys Leu Met Val Met Gly Glu Gly Leu Asp Leu Asp Asp Ala Leu Ala 20 25 30 Val His Tyr Ala Val Gln His Cys Asn 35 40 (2) INFORMATION FOR SEQ ID NO: 22: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 27 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN : not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 22: Arg Arg Pro Asp Ser Lys Thr Ala Leu His Leu Ala Ala Glu Met Val 1 5 10 15 Ser Pro Asp Met Val Ser Val Leu Leu Asp Gln 20 25 (2) INFORMATION FOR SEQ ID NO: 23: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 41 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 23 He Arg Arg Met Arg Arg Ala Leu Asp Ala Wing Asp He Glu Leu Val 1 5 10 15 Lys Leu Met Val Met Gly Glu Gly Leu Asp Leu Asp Asp Ala Leu Ala 20 25 30 Val His Tyr Ala Val Gln His Cys Asn 35 40 (2) INFORMATION FOR SEQ ID NO: 24: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 19 amino acids (B) TYPE: amino acid (C) TYPE OF CHAIN: not relevant (D) TOPOLOGY: not relevant (ii) TYPE OF MOLECULE: peptide (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 24: Pro Thr Gly Lys Thr Ala Leu His Leu Ala Ala Glu Met Val Ser Pro 1 5 10 15 Asp Met Val (2) INFORMATION FOR SEQ ID NO: 25: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 35 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 25: CAACAGCTTC GAAGCCGTCT TTGACGCGCC GGATG 35 (2) INFORMATION FOR SEQ ID NO: 26: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 35 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 26: CATCCGGCGC GTCAAAGACG GCTTCGAAGC TGTTG 35 (2) INFORMATION FOR SEQ ID NO: 27: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 32 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 27: GGAATTCAAT GGATTCGGTT GTGACTGTTT TG 32 (2) INFORMATION FOR SEQ ID NO: 28: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 28 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 28: GGAATTCTAC AAATCTGTAT ACCATTGG 28 (2) INFORMATION FOR SEQ ID NO: 29: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 31 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 29: CGGAATTCGA TCTCTTTAAT TTGTGAATTT C 3 1 (2) INFORMATION FOR SEQ ID NO: 30: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 30: GGAATTCTCA ACAGTTCATA ATCTGGTCG 29 (2) INFORMATION FOR SEQ ID NO: 31: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 31 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 31: GGAATTCAAT GGACTCCAAC AACACCGCCG C 31 (2) INFORMATION FOR SEQ ID NO: 32: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 33 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: other nucleic acid (A) DESCRIPTION / desc = "oligonucleotide" (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 32: GGAATTCTCA ACCTTCCAAA GTTGCTTCTG ATG 33

Claims (31)

1. A method to protect a plant from pathogen attack, through synergistic resistance to diseases, which comprises the steps of: (a) providing an immunomodulated plant having a first level of resistance to diseases; and (b) apply to this immunomodulated plant at least one microbicide that confers a second level of disease resistance, * (c) whereby the application of the microbicide to the immunomodulated plant confers a third, synergistically improved level of resistance. to diseases that is greater than the sum of the first and second levels of disease resistance.

2. A method according to claim 1, wherein the immunomodulated plant is: (a) a mutant plant of constitutive immunity (cim); (b) a mutant plant that mimics injuries; (c) obtained by the recombinant expression in a plant, of an acquired systemic resistance gene; (d) obtained by applying to a plant a chemical capable of inducing acquired systemic resistance.

3. A method according to claim 2, wherein the cim mutant plant is selected from a population of plants according to the following steps: (a) evaluating the expression of acquired systemic resistance genes in uninfected plants that are phenotypically normal, in which the non-infected plants lack an imitation lesion phenotype; and (b) selecting uninfected plants that constitutively express acquired systemic resistance genes in the absence of a viral, bacterial, or fungal infection.

4. A method according to claim 3, wherein the mutant plant that mimics injury is selected from a population of plants according to the following steps: (a) evaluating the expression of systemic resistance genes acquired in plants not infected that have a phenotype that mimics injury; and (b) selecting uninfected plants that constitutively express acquired systemic resistance genes in the absence of a viral, bacterial, or fungal infection.

A method according to claim 2, wherein the acquired systemic resistance gene is a functional form of a NJM1 gene that encodes an? IM1 protein involved in the signal transduction cascade leading to systemic resistance acquired in plants .

6. A method according to claim 5, wherein the NIM1 protein comprises the amino acid sequence stipulated in SEQ ID NO: 2.

7. A method according to claim 5, wherein the NJM1 gene comprises the stipulated coding sequence. in SEQ ID? 0: 1.

A method according to claim 3, wherein the acquired systemic resistance gene encodes an altered form of an? IM1 protein that acts as a negative-dominant regulator of the transduction path of the acquired systemic resistance signal.

9. A method according to claim 8, wherein the altered form of the? IM1 protein has alanines in place of serines at amino acid positions corresponding to positions 55 and 59 of SEQ ID? 0: 2.

10. A method according to claim 9, wherein the altered form of the? IM1 protein comprises the amino acid sequence shown in SEQ ID? 0.-8.

11. A method according to claim 9, wherein the AD? comprises the nucleotide sequence shown in SEQ ID? O: 7.

12. A method according to claim 8, wherein the altered form of the? IM1 protein has a? -terminal truncation of amino acids corresponding approximately to the positions of amino acids 1-125 of SEQ ID? 0: 2.

13. A method according to claim 13, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 10.

14. A method according to claim 13, wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 9.

15. A method according to claim 8, wherein the altered form of the NIM1 protein has a C-terminal truncation of amino acids that corresponds approximately to the amino acid positions 522-593 of SEQ ID NO: 2.

16. A method according to claim 15, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 12.

A method according to claim 15, wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 11.

18. A method according to claim 8, wherein the altered form of the NIM1 protein has an N-terminal truncation of amino acids corresponding approximately to the positions of amino acids 1-125 of SEQ ID NO: 2, and a C-terminal truncation of amino acids corresponding approximately to the positions is amino acid 522-593 of SEQ ID NO: 2.

19. A method according to claim 18, wherein the altered form of the NIM1 protein comprises the amino acid sequence shown in SEQ ID NO: 14.

20. A method according to claim 18, wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 13.

21. A method according to claim 8, wherein the altered form of the NIM1 protein consists of essentially of ankyrin motifs corresponding approximately to the amino acid positions 103-362 of SEQ ID NO: 2.

22. A method according to claim 21, wherein the altered form of the NIM1 protein comprises the sequence of amino acids shown in SEQ ID NO: 16.

23. A method according to claim 21, wherein the DNA molecule comprises the nucleotide sequence shown in SEQ ID NO: 15.

24. A method according to any of claims 7, 11, 14, 17, 20, and 23, wherein the molecule of DNA is hybridized under the following conditions to the nucleotide sequence stipulated in SEQ ID Nos: 1, 7, 9, 11, 13, or 15: hybridization in 1 percent bovine serum albumin; NaP04, 520mM, pH 7.2; 7 percent lauryl sulfate, sodium salt; EDTA lmM; 250 mM sodium chloride at 55 ° C for 18 to 24 hours, and washed in SSC6X for 15 minutes (3 times) SSC3X for 15 minutes (1 time) at 55 ° C.

25. A method according to any of claims 1 to 24, wherein the microbicide is a fungicide selected from the following group: 4 - [3- (4-chlorophenyl) -3- (3,4-dimethoxyphenyl) acryloyl ] morpholine ("dimetomorf"); 5-methyl-1,2,4-triazolo [3,4-b] [1,3] benzothiazole ("tricyclazole"), 1, l-dioxide of 3-allyloxy-1,2-benzothiazole ("probonazole"); c- [2 - (4-chlorophenyl) ethyl] -α- (1,1-dimethylethyl) -1H-1,2,4-triazole-1-ethanol, ("tebuconazole"); 1- [[3- (2-chlorophenyl) -2-- (4-fluorophenyl) oxiran-2-yl] methyl] -1H- 1, 2,4-triazole, ("epoxiconazole"); cu- (4-chlorophenyl) -α- (1-cyclopropylethyl) -1 H-1,2,4-triazole-1-ethanol ("cyproconazole"); 5- (4-chlorobenzyl) -2,2-dimethyl-1- (1H-1,2,4-triazol-1-ylmethyl) -cyclopentanol, ("metconazole"); 2- (2,4-dichlorophenyl) -3- (1H-1,2,4-triazol-1-yl) -propyl-1, 2,2-tetrafluoroethyl ether ("tetraconazole"); (E) -2-. { 2- [6- (2-Cyanophenoxy) pyrimidin-4-yloxy] phenyl} -3-methoxyacrylate methyl ("ICI A 5504", "azoxyestrobin"); (E) -2-methoxy-imino-2- [Oi - (o-tolyloxy) -o-tolyl] methyl acetate; ("BAS 490 F", "cresoxima-methyl"); acetamide 2- (2-phenoxyphenyl) - (E) -2-methoxy-imino-N-methyl, acetamide [2- (2,5-dimethylphenoxymethyl) -phenyl] - (E) -2-methoxy-imino- N-methyl; (1R, 3S / 1S, 3R) -2,2-dichloro-N- [(R) -1- (4-chlorophenyl) ethyl] -1-ethyl-3-methylcyclopropanecarboxamide, ("KTU 3616") complex manganese-polymer of ethylenebis (dithiocarbamate) -zinc ("mancozeb"); 1- [2- (2,4-dichlorophenyl) -4-propyl-l, 3-dioxolan-2-ylmethyl] -1 H-1, 2,4-triazole, ("propiconazole"); 1- . { 2- [2-chloro-4- (4-chlorophenoxy) phenyl] -4-methyl-l, 3-dioxolan-2-ylmethyl) -1 H-1, 2,4-triazole ("difenoconazole"); 1- [2- (2,4-dichlorophenyl) pentyl-lH-1, 2,4-triazole ("penconazole"); cis-4- [3- (4-butyl-tertiary-phenyl) -2-methylpropyl] -2,6-dimethylmorpholine, ("phenpropimorf"); 1- [3- (4-tertiary butyl-phenyl) -2-methylpropyl] -piperidine, ("fenpropidine"), * 4-cyclopropyl-6-methyl-N-phenyl-2-pyrimidinamine ("cyprodinil"); methyl ester of (RS) -N- (2,6-dimethylphenyl-N- (methoxyacetyl) -alanine ("metalaxyl"); methyl ester of (R) -N- (2,6-dimethylphenyl-N- ( methoxyacetyl) -alanine ("R-metalaxyl"); 1,2,5,6-tetrahydro-4H-pyrrolo [3, 2, 1-ij] quinolin-4-one ("pyroquinone"), * and Ethyl acid phosphonate ("fosetyl")

26. A method according to claim 25, wherein the fungicide is metalaxyl

27. A method according to any of claims 2 to 26, wherein the chemical capable of inducing acquired systemic resistance is a benzothiadiazole compound, an isonicotinic acid compound, or a salicylic acid compound

28. A method according to any one of claims 1 to 26, wherein the microbicide is a benzothiadiazole compound, an isonicotinic acid compound, or a salicylic acid compound.

29. A method according to any of claims 1 to 28, wherein two microbicides are applied concurrently to the immunomodulated plant.

30. A method according to claim 29, wherein one of the microbicides is a fungicide selected from the following group: 4- [3- (4-chlorophenyl) -3- (3,4-dimethoxyphenyl) acryloyl] morpholine ("dimetomorf"), * 5-methyl-l, 2,4-triazolo [3,4-b] [1,3] benzothiazole ("tricyclazole"), 1, l-dioxide of 3-allyloxy-1,2-benzothiazole ("probonazole"); oc- [2- (4-chlorophenyl) ethyl] -ae- (1,1-dimethylethyl) -1H-1,2,4-triazole-1-ethanol, ("tebuconazole"); 1- [[3- (2-chlorophenyl) -2-- (4-fluorophenyl) oxiran-2-yl] methyl] -1H- 1, 2,4-triazole, ("epoxiconazole"); - (4-chlorophenyl) -ae- (1-cyclopropylethyl) -1 H -1,2-triazole-1-ethanol, ("cyproconazole"); 5- (4-chlorobenzyl) -2,2-dimethyl-1- (1H-1,2,4-triazol-1-ylmethyl) -cyclopentanol, ("metconazole"), * 2- (2, 4-dichlorophenyl) -3- (1 H-1,2,4-triazol-1-yl) -propyl-1,4,1,2-tetrafluoroethyl ("tetraconazole"); (E) -2-. { 2- [6- (2-cyanophenoxy) pyrimidin-4-yloxy] phenyl} -3-methoxyacrylate methyl ("ICI A 5504", "azoxyestrobin"); (E) -2-methoxy-imino-2- [- (o-tolyloxy) -o-tolyl] methyl acetate; ("BAS 490 F", "cresoxima-methyl"); acetamide 2- (2-phenoxyphenyl) - (E) -2-methoxy-imino- -N-methyl, acetamide [2- (2,5-dimethylphenoxymethyl) -phenyl] - (E) -2-methoxy-imino- N-methyl; (1R, 3S / 1S, 3R) -2,2-dichloro-N- [(R) -1- (4-chlorophenyl) ethyl] -1-ethyl-3-methylcyclopropanecarboxamide, ("KTU 3616") complex manganese-polymer of ethylenebis (dithiocarbamate.) -zinc ("mancozeb"), * 1- [2- (2,4-dichlorophenyl) -4-propyl-l, 3-dioxolan-2-ylmethyl] -1 H-1, 2,4-triazole, (" propiconazole "); 1- . { 2- [2-chloro-4- (4-chlorophenoxy) phenyl] -4-methyl-1,3-dioxolan-2-ylmethyl) -1 H-1, 2,4-triazole ("difenoconazole"); 1- [2- (2,4-dichlorophenyl) pentyl-lH-1, 2,4-triazole ("penconazole"); cis-4- [3- (4-butyl-tertiary-phenyl) -2-methylpropyl] -2,6-dimethylmorpholine, ("phenpropimorf"); 1- [3- (4-tertiary butyl-phenyl) -2-methylpropyl] -piperidine, ("fenpropidine"); 4-cyclopropyl-6-methyl-N-phenyl-2-pyrimidinamine ("cyprodinil"); methyl ester of (RS) -N- (2,6-dimethylphenyl-N- (methoxyacetyl) -alanine ("metalaxyl"); methyl ester of (R) -N- (2,6-dimethylphenyl- -N- ( methoxyacetyl) -alanine ("R-metalaxyl"); 1,2,5,6-tetrahydro-4H-pyrrolo [3, 2, 1-ij] quinolin-4-one ("pyroquinone"); and phosphonate ethyl acid ("fosetyl") and the other microbicide is a benzothiadiazole compound, an isonicotinic acid compound, or a salicylic acid compound

31. A method according to claim 30, wherein the fungicide is metalaxyl , and the other "microbicide is a benzothiadiazole compound.