Application of Rice Transcription Factor OsbHLH091 and Its Encoding Gene in Disease Resistance Breeding

Abstract

The present invention discloses the application of the rice transcription factor OsbHLH091 and its encoding gene in disease-resistant breeding, belonging to the field of genetic engineering. The amino acid sequence of the rice transcription factor OsbHLH091 is shown in SEQ ID NO.2, and the encoding gene is shown in SEQ ID NO.1. The present invention proves through experiments that the encoding gene of the rice transcription factor OsbHLH091 is overexpressed in rice, and the transgenic rice plants have higher disease resistance than the recipient rice plants and the rice plants with OsbHLH091 knocked out, indicating that OsbHLH091 positively regulates rice resistance to rice blast. The OsbHLH091 protein and its encoding gene discovered by the present invention are of great significance for breeding disease-resistant rice varieties and can be applied to molecular design to breed rice with improved disease resistance.

Description

Technical Field

[0001] The present invention relates to the field of genetic engineering, in particular to the application of rice transcription factor OsbHLH091 and its encoding gene in disease-resistant breeding. Background Art

[0002] Fungal diseases such as rice blast cause massive crop yield losses, making the development of disease-resistant crops a major goal of the agricultural industry. Numerous genes associated with plant disease resistance have been reported, including both effector and regulatory genes, from crops such as rice, wheat, maize, and soybean, as well as from the model plant Arabidopsis thaliana. Several of these have been used as target genes in crop disease resistance genetic engineering, successfully resulting in the development of disease-resistant rice, wheat, maize, and soybean varieties. However, differences in genetic backgrounds hinder the application of some genes, necessitating the continued need for new relevant genes to meet breeding needs.

[0003] As one of the most important food crops, improving rice disease resistance is of great theoretical and practical significance. Therefore, finding new rice disease resistance genes remains the key to current disease resistance breeding. Summary of the Invention

[0004] The purpose of the present invention is to provide a rice transcription factor OsbHLH091 and its encoding gene for use in disease resistance breeding to solve the problems of the above-mentioned prior art. OsbHLH091 positively regulates rice resistance to rice blast.

[0005] To achieve the above object, the present invention provides the following solutions:

[0006] The present invention provides the use of rice transcription factor OsbHLH091 in any of the following:

[0007] (1) Application in controlling rice blast;

[0008] (2) Application in the preparation of products for regulating rice blast;

[0009] (3) Application in blast-resistant rice breeding;

[0010] (4) Application in breeding blast-resistant rice;

[0011] The amino acid sequence of the rice transcription factor OsbHLH091 is shown in SEQ ID NO. 2. However, the amino acid sequence of the rice transcription factor OsbHLH091 of the present invention is not limited thereto. The amino acid sequence of the rice transcription factor OsbHLH091 of the present invention may also be a protein derived from the amino acid sequence shown in SEQ ID NO. 2, obtained by substitution and / or deletion and / or addition of one or more amino acid residues, or a protein having 80% or greater identity with the protein of the sequence and having the same function. The protein or derivative may also be a fusion protein obtained by attaching a protein tag to the N-terminus and / or C-terminus of the protein or derivative.

[0012] In the aforementioned proteins or protein derivatives, a protein tag refers to a polypeptide or protein that is fused and expressed with the target protein using in vitro DNA recombination techniques to facilitate the expression, detection, tracing, and / or purification of the target protein. Protein tags can include GFP tags, Flag tags, His tags, MBP tags, HA tags, myc tags, GST tags, and / or SUMO tags.

[0013] In the above-mentioned proteins or derived proteins, 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, the identity of a pair of amino acid sequences can be calculated by searching in Advanced BLAST 2.1 using blastp as the program, setting the Expect value to 10, all filters to OFF, BLOSUM62 as the matrix, and setting the Gap existence cost, Per residue gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively. The identity value (%) can then be obtained.

[0014] In the above proteins or derived proteins, the above 80% or more identity may be at least 81%, 82%, 85%, 86%, 88%, 90%, 91%, 92%, 95%, 96%, 98%, 99% or 100% identity.

[0015] The present invention also provides the use of the gene encoding the rice transcription factor OsbHLH091 in any of the following:

[0016] (1) Application in controlling rice blast;

[0017] (2) Application in the preparation of products for regulating rice blast;

[0018] (3) Application in blast-resistant rice breeding;

[0019] (4) Application in breeding blast-resistant rice;

[0020] The nucleotide sequence of the coding gene is shown in SEQ ID NO. 1. The nucleotide sequence of the coding gene is a cDNA or DNA molecule sequence.

[0021] The present invention also provides the use of a recombinant vector containing a gene encoding rice transcription factor OsbHLH091 in any of the following:

[0022] (1) Application in controlling rice blast;

[0023] (2) Application in the preparation of products for regulating rice blast;

[0024] (3) Application in blast-resistant rice breeding;

[0025] (4) Application in breeding blast-resistant rice;

[0026] The amino acid sequence of the rice transcription factor OsbHLH091 is shown in SEQ ID NO.2, and the nucleotide sequence of the encoding gene is shown in SEQ ID NO.1.

[0027] The present invention also provides the use of a host bacterium containing a recombinant vector in any of the following:

[0028] (1) Application in controlling rice blast;

[0029] (2) Application in the preparation of products for regulating rice blast;

[0030] (3) Application in blast-resistant rice breeding;

[0031] (4) Application in breeding blast-resistant rice;

[0032] The recombinant vector is a vector that integrates the coding gene of rice transcription factor OsbHLH091 into the genome, and the nucleotide sequence of the coding gene is shown in SEQ ID NO.1.

[0033] Carrier of the present invention is a plant expression vector, which includes a binary agrobacterium vector and a vector that can be used for plant microprojectile bombardment. Such as pAHC25, pWMB123, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa or pCAMBIA1391-Xb (CAMBIA company) etc. The plant expression vector can also include the 3' non-translated region of the foreign gene, i.e., a DNA fragment that includes a polyadenylic acid signal and any other DNA fragment that participates in mRNA processing or gene expression. The polyadenylic acid signal can guide polyadenylic acid to join the 3' end of the mRNA precursor, and the non-translated region that transcribes as agrobacterium crown gall induction (Ti) plasmid gene (such as nopaline synthase gene Nos) or plant gene (such as rice starch synthase gene) 3' end all has similar function. When constructing a plant expression vector using the gene of the present invention, enhancers, including translation enhancers or transcription enhancers, may also be used. These enhancer regions may be the ATG start codon or the start codon of an adjacent region, etc., but must be identical to the reading frame of the coding sequence to ensure correct translation of the entire sequence.

[0034] The host bacteria of the present invention include yeast, bacteria, algae and fungi.

[0035] Optionally, the method of regulating rice blast disease is that rice transcription factor OsbHLH091 positively regulates rice blast disease.

[0036] The present invention also provides a method for regulating rice blast, comprising any one of the following methods:

[0037] (1) reducing the resistance of rice to rice blast by knocking out the OsbHLH091 gene in rice;

[0038] (2) improving the resistance of rice to rice blast by overexpressing the OsbHLH091 gene in rice;

[0039] The nucleotide sequence of the OsbHLH091 gene is shown in SEQ ID NO.1.

[0040] The present invention also provides a method for cultivating rice blast-resistant rice, comprising the steps of overexpressing the OsbHLH091 gene in rice, increasing the expression level of the OsbHLH091, and obtaining rice blast-resistant rice; wherein the nucleotide sequence of the OsbHLH091 gene is shown in SEQ ID NO.1.

[0041] The present invention discloses the following technical effects:

[0042] During their research on rice immune responses, the inventors identified a member of the bHLH transcription factor family, which they named OsbHLH091. Experimental results show that overexpressing OsbHLH091 in rice resulted in transgenic rice exhibiting higher disease resistance than the recipient rice, while knocking out OsbHLH091 in rice resulted in transgenic rice exhibiting lower disease resistance than the recipient rice, indicating that overexpression of OsbHLH091 can enhance rice disease resistance. This study demonstrates that OsbHLH091 can positively regulate rice disease resistance, and that the OsbHLH091 protein and its encoding gene are of great significance for the development of disease-resistant rice varieties. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0044] Figure 1 Schematic diagram of the gene structure of OsbHLH091 and the sgRNA designed for the knockout target site;

[0045] Figure 2 This is the gene sequence identification result of OsbHLH091 knockout plant;

[0046] Figure 3 Schematic diagram of the overexpression vector structure of OsbHLH091;

[0047] Figure 4 The expression level of OsbHLH091 gene in OsbHLH091 overexpressing plants is detected.

[0048] Figure 5Figure 2 is a diagram of the phenotypic changes of wild-type rice Z11 and OsbHLH091 knockout and overexpression plants after puncture inoculation with fungi; A is a representative picture of lesions on the leaves of Zhonghua 11 (ZH11), knockout plants OsbHLH091#KOs, and overexpression plants OsbHLH091#OEs after puncture inoculation with rice blast fungi; B is a representative picture of lesions on the leaves of Zhonghua 11 (ZH11), knockout plants OsbHLH091#KOs, and overexpression plants OsbHLH091#OEs after puncture inoculation with rice blast fungi Statistical results of spot length; C is a representative picture of lesions on the leaves of Zhonghua 11 (ZH11), knockout plant OsbHLH091#KOs, and overexpression plant OsbHLH091#OEs after spray inoculation with rice blast fungus; D is the statistical result of the number of lesions on the leaves of Zhonghua 11 (ZH11), knockout plant OsbHLH091#KOs, and overexpression plant OsbHLH091#OEs after puncture inoculation with rice blast fungus; asterisks above the bar graphs indicate significant differences (P<0.05). DETAILED DESCRIPTION

[0049] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0050] It should be understood that the terms described herein are intended only to describe particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges herein, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value within any stated value or stated range, and each smaller range between any other stated value or intermediate value within the stated range, is also encompassed within the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0051] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of any conflict with any incorporated document, the contents of this specification shall prevail.

[0052] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments described herein without departing from the scope or spirit of the invention. Other embodiments will be apparent to those skilled in the art from the description of the invention. The description and examples are intended to be illustrative only.

[0053] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0054] During research on rice immune responses, the applicant identified a member of the bHLH transcription factor family, named OsbHLH091. bHLH transcription factors play an important role in plant development and biotic stress processes, and therefore previous research on OsbHLH091 has focused on its regulation of plant development. The applicant's research found that compared with controls, lesions in OsbHLH091 knockout plants became longer, while lesions in OsbHLH091 overexpressing plants became shorter and fewer, indicating that OsbHLH091 positively regulates plant disease resistance and could be used as a target gene in molecular breeding to enhance plant disease resistance. The disease resistance function of OsbHLH091 is described in detail below using examples.

[0055] Unless otherwise specified, the experimental methods in the examples of the present invention are conventional methods and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials and reagents used in the examples of the present invention are all commercially available unless otherwise specified.

[0056] Some of the experimental materials and reagents involved in the following examples:

[0057] (1) The rice variety Zhonghua 11 (O. Sativa L. spp. japonica, var. zhonghua11, AA genome, ZH11) belongs to the japonica subspecies and is described in the following literature: Ni Yuchong, A New Rice Variety Cultivated by Flower Culture—Zhonghua 11, Agricultural Science and Technology Bulletin, 1989, 07, 35; provided by Professor Chen Xuewei, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences.

[0058] (2) The OsbHLH091 overexpression and knockout transgenic rice of the present invention was completed by Boyuan Biotechnology Company.

[0059] (3) The plant binary expression vector pTCRISPR was provided by Associate Professor Tang Yongyan of the State Key Laboratory of Plant Genetics, Sichuan Agricultural University. The plant binary expression vector pCAMBIA2300 was provided by Professor Wu Xianjun of the Rice Research Institute, Sichuan Agricultural University.

[0060] (4) Total RNA extraction kit: TRIzol purchased from Invitrogen, USA, catalog number 15596026.

[0061] Reverse transcription kit: HiScript III RT SuperMix for qPCR (+gDNA wiper) purchased from Vanzyme, China, catalog number R323-01.

[0062] Fluorescence quantitative kit: AceQ qPCR SYBR Green MasterMix purchased from Vanzyme, China, with the catalog number Q111-02.

[0063] Homologous recombination kit: ClonExpress II One Step Cloning Kit purchased from Vanzyme, China, with the catalog number C112-01.

[0064] Example 1 Gene OsbHLH091 Induced in Rice Immune Response and Its Cloning

[0065] During the study of rice immune responses, we discovered that the expression of OsbHLH091 was induced by rice blast fungus. We used polymerase chain reaction to verify the expression of OsbHLH091 at various time points after rice blast infection. The specific method was as follows:

[0066] ZH11 was used as the research material. The experimental group was sprayed with 5×10 5 ml -1 A solution of 10-8-14 rice blast fungus spores and 0.1% Tween 20 was used. A control group was sprayed with a 0.1% Tween 20 solution. Rice leaves were harvested 12 hours later. The samples were sent to Novogene for ribosomal profiling and sequencing. Analysis confirmed that the translation of this gene is induced by the blast fungus. Comparison revealed that this gene encodes a bHLH transcription factor. Primers were designed based on the reference gene sequence of the rice genome:

[0067] OsbHLH091F (SEQ ID NO.3): 5'-ATGGACGGCGGCAGGGTCGG-3';

[0068] OsbHLH091R (SEQ ID NO. 4): 5'-GAACTCATTTTCATGTGGCCT-3'.

[0069] Using ZH11 total cDNA as a template, the above primers amplified a DNA band of about 1.2Kb. After sequencing, it was confirmed to be the OsbHLH091 gene belonging to the bHLH family. After alignment of the LOC_Os08g41320 sequence in the rice genome reference sequence and the clone sequence amplified from the ZH11 total cDNA, it was shown that the clone sequence obtained from ZH11 was identical to the LOC_Os08g41320 sequence. The amino acid sequence of the OsbHLH091 protein is shown in SEQ ID NO.2. The CDS sequence of the coding chain of the gene OsbHLH091 encoding the OsbHLH091 protein is shown in SEQ ID NO.1 (see Figure 1).

[0070] Example 2 Construction of OsbHLH091 Plant Knockout Vector

[0071] 1. OsbHLH091 knockout vector pTCRISPR-sgRNA OsbHLH091 Construction

[0072] Using the genome of rice ZH11 as a template, the knockout target sequence was designed to be: 5'-AGAATTGTACCCTCTCAGCG-3'. Figure 1 .

[0073] And synthesize the following primers:

[0074] F (SEQ ID NO. 5): 5'-TGTG AGAATTGTACCCTCTCAGCG G-3';

[0075] R (SEQ ID NO. 6): 5'-AAAAC CGCTGAGAGGGTACAATTCT -3'.

[0076] After denaturation at 95°C, anneal to form a double-stranded DNA fragment. Digest the pTCRISPR vector with BsaI to recover the linearized vector. Mix the double-stranded DNA fragment and the linearized vector using the following system and perform a T4 DNA ligase ligation reaction. Incubate at room temperature for 30 minutes.

[0077] Table 1 Connection system

[0078] Reagents Dosage Double-stranded DNA fragments 0.5-4 μL Linearized vector 0.5-4 μL T4ligase 2μL <![CDATA[ddH2O]]> XμL Total volume 10 μL

[0079] The same effect can be achieved within the range shown in Table 1. This experiment specifically uses a 10 μL ligation system: 0.5 μL DNA double-stranded fragment, 0.5 μL linearized vector, 2 μL T4 ligase, and ddH2O to 10 μL. Follow the ligation system reaction to obtain pTCRISPR-sgRNA OsbHLH091 Knockout vector.

[0080] 2. Add the above pTCRISPR-sgRNA OsbHLH091 The knockout vector was transformed into DH5α competent medium. After selection with kanamycin, single clones were picked for colony PCR identification. Plasmids were extracted from positive colonies and sequenced to confirm the correct pTCRISPR-sgRNA. OsbHLH091 If correct, genetic transformation can be carried out.

[0081] 3. OsbHLH091 knockout transgenic rice was developed by Boyuan Biotechnology Co., Ltd. Sequencing and screening of genetically stable knockout plants OsbHLH091#KO1 and OsbHLH091#KO2 revealed insertions or deletions of bases at the target sequence in the knockout plants. Figure 2 .

[0082] Example 3 Construction of OsbHLH091 plant expression vector

[0083] 1. Construction of OsbHLH091 overexpression vector pCAMBIA2300-OsbHLH091

[0084] The cDNA obtained by reverse transcription of total RNA from rice ZH11 was used as the template, and the cloning primers were:

[0085] F (SEQ ID NO. 7): 5'- GGACAGGGTACCCGGGGATCC ATGGACGGCGGCAGGGTCG G-3';

[0086] R (SEQ ID NO. 8): 5'- GGTACTAGTGTCGACTCTAGA GAACTCCATTTTCATGTGGC CT-3'.

[0087] The full-length cDNA of OsbHLH091 was amplified by PCR, and the pCAMBIA2300-35S-eGFP-OCS vector was digested with BamHI and XbaI to recover the linearized vector. The full-length cDNA of OsbHLH091 recovered by PCR and the linearized vector were mixed and homologous recombination was performed as follows.

[0088] Table 2 Homologous recombination reaction system

[0089] Reagents Dosage Double-stranded DNA fragments 0.5-5μL Linearized vector 0.5-5μL 5×CEIIBuffer 4μL ExnaseII 2μL <![CDATA[ddH2O]]> XμL Total volume 20 μL

[0090] The same effect can be achieved within the range shown in Table 2. This experiment specifically used a 20 μL homologous recombination reaction system: 1 μL double-stranded DNA fragment, 1 μL linearized vector, 4 μL 5×CE II Buffer, 2 μL Exnase II, and ddH2O to 20 μL. Recombination was performed according to the homologous recombination reaction system to obtain the pCAMBIA2300-OsbHLH091 recombinant vector, see Figure 3 .

[0091] 2. The pCAMBIA2300-OsbHLH091 recombinant vector was transformed into DH5α competent cells. After kanamycin selection, single clones were picked for colony PCR identification. Plasmids were extracted from positive colonies and sequenced to confirm that pCAMBIA2300-OsbHLH091 was correct, and genetic transformation could be carried out.

[0092] 3. OsbHLH091 overexpression transgenic rice was completed by Boyuan Biotechnology Company. The expression of the target gene was detected and genetically stable overexpression plants OsbHLH091#OE1 and OsbHLH091#OE2 were screened. The results showed that the expression of the target gene in the overexpression plants was significantly increased compared with the wild-type target gene. Figure 4 .

[0093] Example 4 OsbHLH091 is involved in regulating plant disease resistance

[0094] In plants, research on OsbHLH091 has primarily focused on developmental and abiotic stress regulation. The applicants discovered for the first time that the bHLH protein OsbHLH091 is involved in regulating jasmonic acid (JA) biosynthesis in rice, acting as a regulator of the JA pathway. Jasmonic acid, known as a defense hormone, plays a crucial role in regulating plant disease resistance. Therefore, they tested the effect of OsbHLH091 on rice disease resistance.

[0095] Treatment of seedling inoculation during the puncture stage:

[0096] (1) Test materials

[0097] Three-week-old Zhonghua 11 (ZH11), the genetically stable OsbHLH091 knockout lines OsbHLH091#KO1 and OsbHLH091#KO2 with ZH11 background, and the genetically stable OsbHLH091 overexpression lines OsbHLH091#OE1 and OsbHLH091#OE2 with ZH11 background.

[0098] (2) Experimental methods

[0099] Select plump seeds and place them in a conical flask filled with tap water. Germination is carried out in a dark incubator at 37°C, with water changed daily. After 2 days, select white seeds and place them in a 96-well seedling plate. The 96-well seedling plate is placed on a float and grown in Hoagland nutrient solution. After 21 days, select the second-to-last rice leaf of consistent growth and size, puncture it, and inoculate 5 μL of a 5×10 5 ml -1 The blast fungus spores of Zhong10-8-14 were inoculated and the length of the lesions was observed and counted after 5 days. The rice leaves were obtained by the same method and the spray inoculation concentration was 5×10 5 ml -1The blast fungus spores of Zhong10-8-14 were used, and the number of lesions was counted after 5 days.

[0100] The results showed that there was no significant difference in the growth of ZH11, OsbHLH091#KO, and OsbHLH091#OE under hydroponic conditions. The experiment was repeated three times with 20 plants of each type each time. Statistics showed that after puncture inoculation, the length and number of lesions of OsbHLH091#KO plants under the ZH11 background were significantly increased compared with ZH11, indicating that they were more sensitive to pathogens (see Figure 5 ), while overexpression of OsbHLH091 in ZH11 significantly reduced the length and number of lesions, and significantly increased the disease resistance of the plants (see Figure 5 This indicates that OsbHLH091 positively regulates plant disease resistance and its encoding gene OsbHLH091 can be used as a target gene in molecular breeding to improve plant disease resistance.

[0101] SEQ ID NO.1 is as follows:

[0102]

[0103] SEQ ID NO.2 is as follows:

[0104] MDGGRVGGDYISSLLSSSSPRLDFGVPVLDAIVAPGGGGGGGDCGLDKLCGDPGFAERAARLSSFNNGGGGVGQRYGGAGAGLFGMPPPAPGDFAGGGSREASVSDPASSAMKDAAAANAKKRKSTAAAAAAAKGKGKEPPVGEEKESDGKRCKTGNGEKESSVKPKAEQAGSDSSVEDGGGGGQKQGKGKNAKPVEPPKDYVH VRARRGQATDSHSLAERVRRERISQRMKVLQDLVPGCNKVIGKALMLDEIINYVQSLQRQVEFLSMKLATVNPLDFSNLPTLLQKDMFQACGPSASSVFSL ESSNSAFRFAEQGDVFQQFAQNSMESQCTLNQLDLALSQATNAAQYAFQDGTAGANLQQRNFWEDDLQSVFHIENGQSQENGVSAPNFHGQQQAGHMKMEF.

[0105] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art should fall within the scope of protection defined by the claims of the present invention.

Claims

1. Use of rice transcription factor OsbHLH091 in any of the following: (1) Application in positive regulation of rice blast resistance; (2) Application in the preparation of products for positively regulating rice blast resistance; (3) Application in blast-resistant rice breeding; (4) Application in breeding blast-resistant rice; in, The amino acid sequence of the rice transcription factor OsbHLH091 is shown in SEQ ID NO.

2.

2. Use of the gene encoding the rice transcription factor OsbHLH091 in any of the following: (1) Application in positive regulation of rice blast resistance; (2) Application in the preparation of products for positively regulating rice blast resistance; (3) Application in blast-resistant rice breeding; (4) Application in breeding blast-resistant rice; in, The nucleotide sequence of the coding gene is shown in SEQ ID NO.

1.

3. Use of a recombinant vector containing a gene encoding rice transcription factor OsbHLH091 in any of the following: (1) Application in positive regulation of rice blast resistance; (2) Application in the preparation of products for positively regulating rice blast resistance; (3) Application in blast-resistant rice breeding; (4) Application in breeding blast-resistant rice; in, The amino acid sequence of the rice transcription factor OsbHLH091 is shown in SEQ ID NO.2, and the nucleotide sequence of the encoding gene is shown in SEQ ID NO.

1.

4. Use of host bacteria containing recombinant vectors in any of the following: (1) Application in positive regulation of rice blast resistance; (2) Application in the preparation of products for positively regulating rice blast resistance; (3) Application in blast-resistant rice breeding; (4) Application in breeding blast-resistant rice; in, The recombinant vector is a vector for integrating the coding gene of rice transcription factor OsbHLH091 into the genome, and the nucleotide sequence of the coding gene is shown in SEQ ID NO.

1.

5. A method for regulating rice blast resistance, characterized in that: The method comprises improving the resistance of rice to rice blast by overexpressing the OsbHLH091 gene in rice; The nucleotide sequence of the OsbHLH091 gene is shown in SEQ ID NO.

1.

6. A method for cultivating blast-resistant rice, characterized in that: The method comprises the steps of overexpressing the OsbHLH091 gene in rice, increasing the expression level of the OsbHLH091, and obtaining rice blast-resistant rice; wherein the nucleotide sequence of the OsbHLH091 gene is shown in SEQ ID NO.1.

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