ZmPRX38 protein and application thereof in improving lodging resistance of corn

Abstract

The invention discloses a ZmPRX38 protein and application thereof in improving lodging resistance of corn, and belongs to the technical field of biology. The technical problem to be solved by the invention is how to improve the stalk strength of corn. The ZmPRX38 protein disclosed by the invention is derived from Zea mays L., and is a protein of which the amino acid sequence is SEQ ID No.1. The ZmPRX38 protein disclosed by the invention is a protein of which the amino acid sequence is SEQ ID No.1. Experiments prove that the ZmPRX38 protein can improve the stalk strength of the plant, the stem diameter and the stalk bending strength of the plant can be improved after the coding gene of the ZmPRX38 protein is introduced into the plant to improve the expression quantity of the ZmPRX38 gene, and the ZmPRX38 gene and the protein coded by the ZmPRX38 gene can be used for improving the stalk strength of the plant and can be further used for cultivating lodging-resistant plants.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and particularly relates to a ZmPRX38 protein and an application thereof in improving the lodging resistance of corn. Background Art

[0002] Breeding new corn varieties with superior lodging resistance is an important solution to reducing the adverse effects of lodging on production, and improving stalk lodging resistance is a key approach to breeding lodging-resistant varieties. Previous studies have shown that corn stalk lodging resistance is affected by stalk strength, and increasing stalk strength is beneficial for improving lodging resistance.

[0003] Culm strength is closely related to lignin content. Lignin biosynthesis relies on the phenylpropanoid pathway, in which peroxidases are key enzymes influencing lignin monomer polymerization. Peroxidase genes that influence lignin content have been identified in several species, including Arabidopsis thaliana, zinnia, tobacco, and poplar. Identifying peroxidase genes in maize is crucial for improving maize culm strength and lodging resistance. Summary of the Invention

[0004] The technical problem to be solved by the present invention is how to improve the strength of corn stalks.

[0005] To solve the above technical problems, the present invention first provides at least one of the following applications of a protein or a substance that regulates the activity of the protein: D1) Regulate plant stem strength; D2) preparing products for regulating plant stem strength; D3) Regulates plant stem thickness; D4) preparing products for regulating plant stem diameter; D5) Cultivating plants with altered stem strength; D6) preparing products for cultivating plants with altered stem strength; D7) Cultivating plants with altered stem diameter; D8) preparing products of cultivating plants with altered stem diameter; The protein is derived from corn ( Zea mays L.), which is named ZmPRX38 protein, and the ZmPRX38 protein is as follows: A1), A2) or A3): A1) a protein having an amino acid sequence of SEQ ID No. 1; A2) a protein having an amino acid sequence as shown in SEQ ID No. 1 in the sequence listing, with substitutions and / or deletions and / or additions of amino acid residues, which has 98% or more identity with A1) and has the same function; A3) A fusion protein obtained by connecting a tag to the N-terminus and / or C-terminus of A1) or A2).

[0006] In the ZmPRX38 protein described in A2), the "98% or greater identity" refers to 98% or 99% identity. Identity refers to amino acid sequence identity. Amino acid sequence identity can be determined using homology search sites on the Internet, such as the BLAST page on the NCBI homepage. For example, using Advanced BLAST 2.1, blastp can be used as the program, with the Expect value set to 10, all filters set to OFF, BLOSUM62 as the matrix, and the Gap existence cost, Per residue gap cost, and Lambda ratio set to 11, 1, and 0.85 (default values), respectively. The identity (%) can be calculated for a pair of amino acid sequences.

[0007] The ZmPRX38 protein in A2) above can be artificially synthesized, or its encoding gene can be synthesized first and then expressed biologically.

[0008] The gene encoding the ZmPRX38 protein in A2) above can be obtained by deleting one or more amino acid residue codons from the DNA sequence set forth at positions 386-1426 of SEQ ID No. 3, and / or performing one or more base pair missense mutations, and / or attaching a tag coding sequence to its 5′ and / or 3′ end. The DNA molecule set forth at positions 386-1426 of SEQ ID No. 3 encodes the ZmPRX38 protein set forth in SEQ ID No. 1.

[0009] The tag in A3) can be a polypeptide or protein fused with the target protein using in vitro DNA recombination techniques to facilitate expression, detection, tracing, and / or purification of the target protein. Examples of such tags include Poly-Arg, Poly-His, FLAG, Strep-tag II, c-myc, MBP tag, HA tag, GST tag, and / or SUMO tag.

[0010] In the above application, the substance that regulates the content and activity of ZmPRX38 protein may be any one of the following B1) to B7): B1) Nucleic acid molecule encoding ZmPRX38 protein; B2) an expression cassette containing the nucleic acid molecule described in B1); B3) a recombinant vector containing the nucleic acid molecule described in B1), or a recombinant vector containing the expression cassette described in B2); B4) a recombinant microorganism containing the nucleic acid molecule described in B1), or a recombinant microorganism containing the expression cassette described in B2), or a recombinant microorganism containing the recombinant vector described in B3); B5) a transgenic plant cell line containing the nucleic acid molecule described in B1) or a transgenic plant cell line containing the expression cassette described in B2); B6) transgenic plant tissue containing the nucleic acid molecule described in B1) or transgenic plant tissue containing the expression cassette described in B2); B7) A transgenic plant organ containing the nucleic acid molecule described in B1) or a transgenic plant organ containing the expression cassette described in B2).

[0011] The nucleic acid molecule may be DNA, such as cDNA, genomic DNA or recombinant DNA; the nucleic acid molecule may also be RNA, such as mRNA or hnRNA.

[0012] Those skilled in the art can readily mutate the nucleotide sequence encoding the ZmPRX38 protein of the present invention using known methods, such as directed evolution and point mutagenesis. Artificially modified nucleotide sequences that share 75% or greater identity with the nucleotide sequence of the isolated ZmPRX38 protein of the present invention are derived from and are equivalent to the nucleotide sequence of the present invention, as long as they encode the ZmPRX38 protein and possess the function of the ZmPRX38 protein.

[0013] In the above application, the nucleic acid molecule in B1) may be a DNA molecule whose coding sequence is positions 386-1426 of SEQ ID No. 3 in the sequence listing.

[0014] Specifically, the nucleic acid molecule in B1) may be the DNA molecule shown at positions 386-1426 of SEQ ID No. 3, or the DNA molecule shown in SEQ ID No. 3 or the DNA molecule shown in SEQ ID No. 2.

[0015] In the above application, the expression cassette ( ZmPRX38 Gene expression cassette), refers to DNA capable of expressing ZmPRX38 protein in host cells, which may include not only promoter ZmPRX38 The promoter of gene transcription may also include the terminator ZmPRX38The terminator of gene transcription. Further, the expression cassette may also include an enhancer sequence. Promoters that can be used in the present invention include but are not limited to: constitutive promoters, tissue-, organ- and development-specific promoters, and inducible promoters. Examples of promoters include, but are not limited to, the constitutive promoter 35S of cauliflower mosaic virus; a wound-inducible promoter from tomato, leucine aminopeptidase ("LAP," Chao et al. (1999) Plant Physiol 120:979-992); a chemically inducible promoter from tobacco, pathogenesis-related 1 (PR1) (induced by salicylic acid and BTH (benzothiadiazole-7-thiocarboxylic acid S-methyl ester)); the tomato proteinase inhibitor II promoter (PIN2) or the LAP promoter (both inducible by methyl jasmonate); a heat shock promoter ( U.S. Pat. No. 5,187,267 ); a tetracycline-inducible promoter ( U.S. Pat. No. 5,057,422 ); seed-specific promoters, such as the millet seed-specific promoter pF128 ( CN101063139B (China Patent 200710099169.7 )), promoters specific for seed storage proteins (e.g., phaseolin, napin, The promoters for oleosin and soybean beta-conglycin (Beachy et al. (1985) EMBO J. 4: 3047-3053) can be used alone or in combination with other plant promoters. All references cited herein are incorporated by reference in their entirety. Suitable transcription terminators include, but are not limited to, the Agrobacterium nopaline synthase terminator (NOS terminator), the cauliflower mosaic virus CaMV 35S terminator, the tml terminator, the pea rbcS E9 terminator, and the nopaline and octopine synthase terminators (see, e.g., Odell et al. (1985) Nature 313:810; Rosenberg et al. (1987) Gene, 56:125; Guerineau et al. (1991) Mol. Gen. Genet, 262:141; Proudfoot (1991) Cell, 64:671; Sanfacon et al. Genes Dev., 5:141; Mogen et al. (1990) Plant Cell, 2:1261; Munroe et al. (1990) Gene, 91:151; Ballad et al. (1989) Nucleic Acids Res. 17:7891; Joshi et al. (1987) Nucleic Acid Res., 15:9627).

[0016] Existing expression vectors can be used to constructZmPRX38 Recombinant vectors containing gene expression cassettes. These plant expression vectors include binary Agrobacterium vectors and vectors that can be used for plant microprojectile bombardment. Examples include pAHC25, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa, PSN1301, or pCAMBIA1391-Xb (CAMBIA). These plant expression vectors may also contain the 3′ untranslated region of the foreign gene, i.e., a polyadenylation signal and any other DNA fragments involved in mRNA processing or gene expression. The polyadenylation signal can direct the addition of polyadenylic acid to the 3′ end of an mRNA precursor, such as genes from the Agrobacterium crown gall-inducing (Ti) plasmid (e.g., the nopaline synthase gene). Nos ), the 3' end transcribed non-translated regions of plant genes (such as soybean storage protein genes) all have similar functions. When using the gene of the present invention to construct a plant expression vector, enhancers can also be used, including translation enhancers or transcription enhancers. These enhancer regions can be ATG start codons or adjacent region start codons, etc., but must be the same as the reading frame of the coding sequence to ensure the correct translation of the entire sequence. The sources of the translation control signals and start codons are wide-ranging and can be natural or synthetic. The translation initiation region can come from the transcription initiation region or the structural gene. In order to facilitate the identification and screening of transgenic plant cells or plants, the plant expression vector used can be processed, such as adding genes that can be expressed in plants and encode enzymes or luminescent compounds that can produce color changes ( GUS genes, luciferase genes, etc.), antibiotic marker genes (such as those that confer resistance to kanamycin and related antibiotics) nptII Gene that confers resistance to the herbicide phosphinothricin bar Gene that confers resistance to the antibiotic hygromycin hph genes, and those that confer resistance to methotrexate dhfr Genes such as the EPSPS gene that confers glyphosate resistance, chemical resistance marker genes (such as herbicide resistance genes), and the mannose-6-phosphate isomerase gene that provides the ability to metabolize mannose can be added. For the safety of transgenic plants, it is possible to directly screen transformed plants using stress without adding any selectable marker genes.

[0017] In the above applications, the microorganism may be yeast, bacteria, algae or fungi. Among them, the bacteria may be Agrobacterium, such as Agrobacterium EHA105.

[0018] In the above applications, the transgenic plant cell lines, transgenic plant tissues and transgenic plant organs do not include reproductive materials.

[0019] In the above application, the substance that regulates the activity of the ZmPRX38 protein content may be a substance that increases the activity of the ZmPRX38 protein content, the substance that regulates the plant stem strength may be an increase in the plant stem strength, the substance that regulates the plant stem thickness may be an increase in the plant stem thickness, the substance that changes the plant stem strength may be an increase in the plant stem strength, and the substance that changes the plant stem thickness may be an increase in the plant stem thickness.

[0020] The present invention also provides any of the following methods: X1) A method for increasing plant stem strength, comprising: expressing a ZmPRX38 protein in a plant, or increasing the content or activity of the ZmPRX38 protein in the plant, to obtain a plant with increased stem strength, thereby increasing plant stem strength; X2) A method for cultivating plants with increased culm strength, comprising: expressing a ZmPRX38 protein in the plant, or increasing the content or activity of the ZmPRX38 protein in the plant, to obtain a plant with increased culm strength; X3) A method for increasing plant stem diameter, comprising: expressing a ZmPRX38 protein in a plant, or increasing the content or activity of the ZmPRX38 protein in a plant, to obtain a plant having increased stem diameter, thereby achieving increased plant stem diameter; X4) A method for cultivating plants with increased stem thickness, comprising: expressing ZmPRX38 protein in the plant, or increasing the content or activity of ZmPRX38 protein in the plant, to obtain the target plant with increased stem thickness.

[0021] Among the above methods, methods X1) to X4) can be achieved by introducing a gene encoding the ZmPRX38 protein into the plant and expressing the gene encoding the gene.

[0022] In the above method, the encoding gene may be the nucleic acid molecule described in B1).

[0023] The gene encoding ZmPRX38 can be introduced into a recipient plant using a recombinant expression vector containing the gene encoding ZmPRX38.

[0024] The recombinant expression vector can be introduced into plant cells by conventional biotechnology methods such as Ti plasmid, plant virus vector, direct DNA transformation, microinjection, electroporation, etc. (Weissbach, 1998, Method for Plant Molecular Biology VIII, Academy Press, New York, pp.411-463; Geiserson and Corey, 1998, Plant Molecular Biology (2nd Edition)).

[0025] The target plant is understood to include not only first-generation plants in which the ZmPRX38 protein or its encoding gene has been altered, but also its progeny. The target plant can be propagated within the species in which the gene was altered, or it can be transferred into other varieties of the same species, particularly commercial varieties, using conventional breeding techniques. The target plant includes seeds, callus tissue, whole plants, and cells.

[0026] The ZmPRX38 protein, or the substance that regulates the content and activity of the ZmPRX38 protein, also falls within the scope of protection of the present invention.

[0027] In the present invention, the plant may be M1) or M2) or M3): M1) Monocots; M2) grasses; M3) Corn.

[0028] Experiments have shown that the ZmPRX38 protein of the present invention can increase the stem strength of plants. After introducing the coding gene of the ZmPRX38 protein into plants to increase the expression level of the ZmPRX38 gene, the stem diameter and stem bending strength of the plants can be increased. The ZmPRX38 gene and the protein encoded by it can be used to increase the stem strength of plants and can be further used to cultivate lodging-resistant plants.

[0029] The present invention will be further described in detail below in conjunction with specific embodiments. The examples provided are only for illustrating the present invention and are not intended to limit the scope of the present invention. The examples provided below can serve as a guide for further improvements by those skilled in the art and are not intended to limit the present invention in any way. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 . ZmPRX38 Schematic diagram of the structure of gene overexpression plasmid wmv072-ZmPRX38.

[0031] Figure 2 .genetically modified ZmPRX38 Phenotype and expression level of corn. A and B are transgenic ZmPRX38 Appearance of corn. C shows the appearance of different transgenic events and wild type (WT). ZmPRX38 Expression level detection results. D is the stem diameter of the third node of different transgenic events and the wild type. *, **, and *** indicate significant differences at the 0.05, 0.01, and 0.001 levels, respectively.

[0032] Figure 3 .genetically modified ZmPRX38Corn stalk bending strength. AC represents the bending strength test results for sections 3, 4, and 5, respectively. ** and *** indicate significant differences at the 0.01 and 0.001 levels, respectively. DETAILED DESCRIPTION

[0033] Unless otherwise specified, the experimental methods in the following examples are conventional methods and were performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, and instruments used in the following examples are all commercially available unless otherwise specified.

[0034] In the following examples, unless otherwise specified, the first position of each nucleotide sequence in the sequence listing is the 5' terminal nucleotide of the corresponding DNA / RNA, and the last position is the 3' terminal nucleotide of the corresponding DNA / RNA.

[0035] Example 1: ZmPRX38 protein can improve corn stalk strength This example found that the Zea mays Peroxidase genes of L. ZmPRX38 It can regulate the stalk strength of corn. In the corn inbred line B73, ZmPRX38 The genomic sequence of the gene is shown in SEQ ID No. 2, the T01 transcript sequence is shown in SEQ ID No. 3, and its CDS sequence is positions 386-1426 of SEQ ID No. 3, encoding the ZmPRX38 protein shown in SEQ ID No. 1.

[0036] The artificially synthesized SEQ ID No.3 ZmPRX38 The T01 transcript (2422 bp) was ligated into plasmid wmv072 (Weimi Biotechnology (Jiangsu) Co., Ltd.) to obtain ZmPRX38 The structure of the gene expression plasmid wmv072-ZmPRX38 is as follows: T-DNA region right-border, Ubi promoter, ZmPRX38 gene, 3×HA tag, Nos terminator, 35S promoter, BlpR gene, CaMV35S polyA signal, T-DNA region Left-border ( Figure 1 wmv072-ZmPRX38 can express a fusion protein formed by a 3×HA tag and the ZmPRX38 protein shown in SEQ ID No. 1, and the expression of the ZmPRX38 gene is driven by the Ubi promoter.

[0037] Will ZmPRX38The gene expression plasmid wmv072-ZmPRX38 was introduced into Agrobacterium EHA105 and transformed into immature embryos of the maize inbred line B73 by Agrobacterium infection. The transformation method was based on the article published by Ishida et al. in 2007 (Ishida Y, HieiY, Komari T. Agrobacterium-mediated transformation of maize. Nat Protoc. 2007; 2(7): 1614-1621.), resulting in T0 generation transgenic ZmPRX38 maize.

[0038] Identification of transgenic positive plants: Using bar gene to identify T0 transgenic plants ZmPRX38 Corn was tested and the primer information used is as follows: Bar-F1: 5′-CCATCGTCAACCACTACATCGAGACA-3′; Bar-R1: 5′-CTTCAGCAGGTGGGTGTAGAGCGT-3′.

[0039] The material from which the Bar gene fragment was successfully amplified was the positive transgenic ZmPRX38 maize. The T0 generation of the three different transgenic events were positive transgenic ZmPRX38 Maize self-pollination resulted in the T1 generation of plants OE1, OE2 and OE3.

[0040] The T1 lines were planted in the field, and RNA was extracted from the third node at the base of the stem at the early stage of tasseling. ZmPRX38 Gene-specific qRT-PCR primers for wild type (WT, i.e., inbred line B73) and positive transgenic ZmPRX38 Corn ZmPRX38 The expression of the gene was detected. The detection primers used are as follows: PRX3-RTF: 5′-GTCCTGTGTTCGATTGTCTTGC-3′; PRX3-RTR: 5′-CTCTGTGGTGGGGGATTGCTT-3′.

[0041] The internal reference is the actin gene, and its primer sequences are as follows: ZmActin-F: 5′-TCACCCTGTGCTGGCTGACCG-3′; ZmActin-R: 5′-GAACCGTTGGCTCACACCA-3′.

[0042] The results showed that OE1, OE2 and OE3 ZmPRX38 The expression levels of the genes were 2.26, 2.13, and 1.56 times that of the wild type (Figure 2 ).

[0043] T2 plants were obtained by self-pollinating T1 plants OE1, OE2, and OE3. T2 plants and a wild-type control (WT: B73) were planted in Sanya, Hainan. Stem bending strength and stem diameter at the 3rd, 4th, and 5th nodes at the base of plants with good growth and normal fruit set were assessed 30 days after pollination. A total of 27, 21, 24, and 39 plants were tested for OE1, OE2, OE3, and WT, respectively.

[0044] The stem diameter was measured as follows: a tape measure was used to circle the cross section of the stem at the third node at the base. The circumference of the stem was the stem diameter (in cm). The bending strength was measured as follows: The stem strength was determined according to the literature (Wang X, Chen Y, Sun X, et al. Characteristics and candidate genes associated with excellent stalk strength in maize ( Zea mays L.). Front Plant Sci. 2022;13:957566.) The specific measurement steps are as follows: the stem is fixed horizontally on a stem strength measuring instrument (model YYD-1, Zhejiang Top Yunnong Technology Co., Ltd.), and the instrument is used to apply a vertical force to the stem. The maximum instantaneous force at which the stem breaks is the stem strength, measured in Newtons.

[0045] Field observations revealed that the plant types of OE1, OE2, and OE3 were consistent with the wild type, but their stem diameters were significantly larger than those of the wild type. The stem diameters of the third node of OE1, OE2, OE3, and WT were 7.08±0.68 cm, 6.93±0.56 cm, 6.72±0.51 cm, and 6.47±0.48 cm, respectively. Figure 2 ). Investigation of stem strength revealed that the bending strength of the third, fourth, and fifth nodes of OE1, OE2, and OE3 were significantly greater than those of the wild type. The third node stem strengths of OE1, OE2, OE3, and WT were 637.3±156.63 N, 550.33±89.58 N, 515.00±130.60 N, and 412.11±111.34 N, respectively ( Figure 3 ). It shows that improving ZmPRX38 The expression of the gene can increase the stem thickness and bending strength of corn. ZmPRX38 The gene and the protein it encodes can be used to improve the stalk strength of corn and can be further used to breed lodging-resistant corn.

[0046] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be practiced over a wide range of parameters, concentrations, and conditions without departing from the spirit and scope of the present invention and without unnecessary experimentation. Although specific embodiments have been given herein, it should be understood that further modifications may be made to the present invention. In summary, this application is intended to encompass any variations, uses, or improvements to the present invention, including those made by conventional techniques known in the art that depart from the scope of the present invention. Applications of the essential features may be made within the scope of the following claims.

Claims

1. At least one of the following uses of a protein or a substance that regulates the content or activity of the protein: D1) Regulate plant stem strength; D2) preparing products for regulating plant stem strength; D3) Regulates plant stem thickness; D4) preparing products for regulating plant stem diameter; D5) Cultivating plants with altered stem strength; D6) preparing products for cultivating plants with altered stem strength; D7) Cultivating plants with altered stem diameter; D8) preparing products of cultivating plants with altered stem diameter; The protein is as follows: A1), A2) or A3): A1) a protein having an amino acid sequence of SEQ ID No. 1; A2) a protein having an amino acid sequence as shown in SEQ ID No. 1 in the sequence listing, with substitutions and / or deletions and / or additions of amino acid residues, which has 98% or more identity with A1) and has the same function; A3) A fusion protein obtained by connecting a tag to the N-terminus and / or C-terminus of A1) or A2).

2. The use according to claim 1, characterized in that: The substance that regulates the protein content activity is any one of the following B1) to B7): B1) a nucleic acid molecule encoding the protein; B2) an expression cassette containing the nucleic acid molecule described in B1); B3) a recombinant vector containing the nucleic acid molecule described in B1), or a recombinant vector containing the expression cassette described in B2); B4) a recombinant microorganism containing the nucleic acid molecule described in B1), or a recombinant microorganism containing the expression cassette described in B2), or a recombinant microorganism containing the recombinant vector described in B3); B5) a transgenic plant cell line containing the nucleic acid molecule described in B1) or a transgenic plant cell line containing the expression cassette described in B2); B6) transgenic plant tissue containing the nucleic acid molecule described in B1) or transgenic plant tissue containing the expression cassette described in B2); B7) A transgenic plant organ containing the nucleic acid molecule described in B1) or a transgenic plant organ containing the expression cassette described in B2).

3. The use according to claim 2, characterized in that: B1) The nucleic acid molecule is a DNA molecule whose coding sequence is positions 386 to 1426 of SEQ ID No. 3 in the sequence listing.

4. The use according to any one of claims 1 to 3, characterized in that: The substance that regulates the activity of the protein content is a substance that increases the activity of the protein content, the substance that regulates the plant stem strength is to increase the plant stem strength, the substance that regulates the plant stem thickness is to increase the plant stem thickness, the plant stem strength is changed to increase the plant stem strength, and the plant stem thickness is changed to increase the plant stem thickness.

5. The use according to any one of claims 1 to 4, characterized in that: The plant is M1) or M2) or M3): M1) Monocots; M2) grasses; M3) Corn.

6. Any of the following methods: X1) Methods for increasing plant stem strength, including: Expressing the protein of claim 1 in a plant, or increasing the content or activity of the protein of claim 1 in a plant, to obtain a target plant with improved stem strength, thereby achieving improved plant stem strength; X2) A method for cultivating a plant with improved stalk strength, comprising: expressing the protein of claim 1 in the plant, or increasing the content or activity of the protein of claim 1 in the plant, to obtain the target plant with improved stalk strength; X3) A method for increasing the stem diameter of a plant, comprising: expressing the protein of claim 1 in a plant, or increasing the content or activity of the protein of claim 1 in a plant, to obtain a plant having increased stem diameter, thereby increasing the stem diameter of the plant; X4) A method for cultivating plants with increased stem thickness, comprising: expressing the protein of claim 1 in the plant, or increasing the content or activity of the protein of claim 1 in the plant, to obtain the target plant with increased stem thickness.

7. The method according to claim 6, wherein: The methods X1) to X4) are achieved by introducing a gene encoding the protein into the plant and expressing the gene encoding the protein.

8. The method according to claim 7, wherein: The coding gene is the nucleic acid molecule described in B1) of claim 2 or 3.

9. The method according to any one of claims 6 to 8, characterized in that: The plant is M1) or M2) or M3): M1) Monocots; M2) grasses; M3) Corn.

10. The protein according to claim 1, or the substance according to any one of claims 1 to 3 that regulates the activity of the protein.

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