TaGly gene for negatively regulating resistance to wheat scab and application thereof

Abstract

The application discloses a TaGly gene for negatively regulating resistance to wheat scab and application thereof, and belongs to the technical field of genetic engineering and molecular breeding. The nucleotide sequence of the TaGly gene is shown as SEQ ID NO. 1, and the amino acid sequence of the encoded protein is shown as SEQ ID NO. 3. The application obtains a transgenic plant by constructing an overexpression vector to transform wheat. Experimental results show that overexpression of the TaGly gene can significantly increase the susceptibility of wheat to scab, indicating that the gene is a susceptibility gene for negatively regulating resistance to wheat scab. Therefore, inhibition or elimination of the TaGly gene by a method such as gene knockout, gene silencing or marker-assisted selection can effectively improve the resistance of wheat to scab. The application provides a new gene resource and technical strategy for molecular breeding of wheat resistant to scab, and has important significance for cultivating wheat varieties resistant to diseases.

Description

Technical Field

[0001] This invention belongs to the fields of genetic engineering and molecular breeding technology, specifically relating to a TaGly gene that negatively regulates wheat scab resistance and its application. Background Technology

[0002] Wheat scab (Fusarium head blight, FHB) is caused by Fusarium graminearum (… Fusarium graminearum This is a global wheat disease caused by fungi such as deoxynivalenol (DON), which seriously threatens global food security. The disease not only leads to a significant drop in wheat yield and shriveled grains, but more seriously, the pathogen accumulates fungal toxins such as DON during infection, contaminating grains and posing a major risk to human and animal health.

[0003] Currently, the control of wheat scab mainly relies on chemical fungicides and agricultural cultivation practices. However, chemical control not only increases production costs but also easily leads to drug resistance in pathogens and causes environmental problems. Therefore, discovering and creating wheat germplasm resources resistant to scab, and breeding and promoting resistant varieties are the most economical, effective, and environmentally friendly control strategies. However, wheat scab resistance is a typical quantitative trait, controlled by multiple genes with complex genetic mechanisms. In addition, the wheat genome is large, highly resistant germplasm resources are scarce, and conventional breeding progress is slow. At present, the number of highly resistant varieties widely used in production is very limited, and the main sources of resistance are still concentrated in a few germplasms represented by "Sumai 3". The resistance gene resources are severely homogenized, and there is a huge risk of resistance failure.

[0004] In recent years, with the development of molecular biology techniques, an increasing number of genes related to Fusarium head blight resistance have been discovered and their functions verified. These genes can be divided into two categories based on their mechanisms of action: one is positively regulated resistance genes, whose high expression or enhanced function can improve plant resistance; the other is negatively regulated resistance genes (also known as susceptibility genes), which pathogens often use to suppress host defenses and promote their own infection. Therefore, knocking out or silencing these genes can effectively enhance plant resistance. In the plant-pathogen interaction, pathogens secrete effectors that hijack the host's susceptibility genes, weakening the immune response. Discovering and analyzing the function of susceptibility genes provides novel molecular targets for creating durable resistant varieties through gene editing. For example, Chinese patent application CN121826030A shows that knocking out the TaCatB gene (encoding catalase) in wheat can significantly enhance Fusarium head blight resistance, confirming that knocking out susceptibility genes is an effective strategy for improving Fusarium head blight resistance.

[0005] Glycosyltransferases (GTs) are a large superfamily of genes in plants that catalyze glycosylation reactions, participating in cell wall synthesis, hormone homeostasis, and secondary metabolite modification, playing crucial roles in plant growth, development, and stress responses. Existing research has shown that the expression levels of some UDP-glycosyltransferase (UGT) genes change significantly after *Fusarium graminearum* infection of wheat. For example, the TaUGT7 gene has been reported to be upregulated by the pathogen and DON toxin, playing a positive regulatory role in resistance to Fusarium head blight. However, the functional mechanisms of many other wheat UGT genes, which are also induced by the pathogen but whose DON detoxification activities are not yet clear, remain unclear. Particularly in model plants such as *Arabidopsis thaliana* and rice, some homologous GT genes have been shown to negatively regulate immune responses by modifying defense signaling molecules such as salicylic acid (SA). However, whether these pathogen-induced GT genes also play a similar negative regulatory role in wheat Fusarium head blight resistance has not been reported in the current technology.

[0006] Therefore, discovering and identifying novel susceptible genes in wheat that participate in the regulation of resistance to Fusarium head blight and elucidating their molecular mechanisms of negative regulation of resistance is of great scientific significance and application value for enriching the theoretical basis of wheat disease resistance breeding, expanding the available gene resources, and breaking the bottleneck of single resistance sources. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention provides a TaGly gene that negatively regulates wheat resistance to Fusarium head blight and its application. For the first time, the TaGly gene was isolated and identified from wheat, and its biological function as a negative regulator in wheat resistance to Fusarium head blight was clarified. By constructing an overexpression vector and introducing it into the wheat variety Fielder, stable transgenic plants were obtained. Experimental results showed that transgenic wheat overexpressing the TaGly gene, after inoculation with Fusarium graminearum, exhibited a significantly increased rate of diseased spikelets and a markedly enhanced susceptibility to Fusarium head blight, thus confirming that the TaGly gene can negatively regulate wheat resistance to Fusarium head blight.

[0008] This invention is achieved through the following technical solution:

[0009] A TaGly gene that negatively regulates wheat scab resistance, the nucleotide sequence of which is as follows (A1) or (A2):

[0010] (A1) The DNA sequence shown in SEQ ID NO.1;

[0011] (A2) The coding region is the DNA sequence shown in SEQ ID NO.2.

[0012] A TaGly protein encoded by the aforementioned TaGly gene, wherein the amino acid sequence of the protein is any one of the following (B1) to (B4):

[0013] (B1) The amino acid sequence as shown in SEQ ID NO.3;

[0014] (B2) A derivative protein having the same function by substitution, deletion and / or addition of one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3;

[0015] (B3) is a protein that has more than 80% homology with the amino acid sequence defined in (B1) or (B2) and has the same function;

[0016] (B4) A fusion protein obtained by attaching a tag to the N-terminus and / or C-terminus of any of the proteins defined in (B1) to (B3).

[0017] A recombinant vector containing the aforementioned TaGly gene.

[0018] Preferably, the nucleotide sequence of the recombinant vector is as shown in SEQ ID NO.4, which is obtained by inserting the nucleotide sequence shown in SEQ ID NO.1 between the HindIII and BamHI multiple cloning sites of the pCambia3300 backbone vector.

[0019] A host cell containing the above-mentioned recombinant vector.

[0020] The application of the aforementioned TaGly gene, the aforementioned TaGly protein, the aforementioned recombinant vector, or the aforementioned host cell in the negative regulation of wheat scab resistance.

[0021] The aforementioned TaGly gene is used as a negative regulatory target in improving wheat scab resistance.

[0022] A breeding method for improving wheat resistance to Fusarium head blight includes the following steps:

[0023] The expression of the TaGly gene mentioned above in wheat can be suppressed or knocked out, or the susceptible allelic variant of the TaGly gene can be eliminated, through gene editing, RNA interference, or marker-assisted selection.

[0024] Preferably, the gene editing uses the CRISPR / Cas9 system to target and knock out the TaGly gene.

[0025] A method for identifying wheat scab resistance includes the following steps:

[0026] The wheat's resistance to Fusarium head blight is determined by detecting the expression level or allelic variation type of the TaGly gene mentioned above: wheat is susceptible when the TaGly gene is highly expressed or when there is a susceptible allelic variation; wheat is resistant when the TaGly gene is knocked out or when there is a resistant allelic variation.

[0027] The beneficial effects of this invention are as follows:

[0028] (1) This invention provides a novel gene TaGly that negatively regulates wheat resistance to Fusarium head blight and its encoded protein, which enriches the gene resources of the wheat resistance to Fusarium head blight regulatory network and provides a new theoretical basis for understanding the molecular mechanism of the interaction between wheat and Fusarium graminearum.

[0029] (2) This invention clarifies the application value of the TaGly gene as a target for disease susceptibility (negative regulation). By inhibiting or eliminating the disease susceptibility allelic variant of this gene through gene knockout, gene silencing, or marker-assisted selection, the resistance of wheat to Fusarium head blight can be effectively improved, providing a new technical strategy and important gene resource for molecular breeding of wheat resistant to Fusarium head blight.

[0030] (3) This invention overcomes the limitations of the existing technology that mainly relies on a single resistance source (such as Fhb1), and provides a new solution to break the bottleneck of single resistance source in wheat scab resistance breeding, which helps to cultivate new wheat varieties with broad-spectrum and long-lasting resistance.

[0031] (4) The TaGly gene and its recombinant vector and host cell provided by the present invention can be used to prepare transgenic wheat with improved resistance to Fusarium head blight, and can also be used as a molecular marker for resistance identification and screening of wheat germplasm resources and breeding materials, which has important theoretical and applied value. Attached Figure Description

[0032] Figure 1 Nucleic acid sequence alignment of TaGly gene in Weining 7840 and Clark;

[0033] Figure 2 This is a simplified diagram of the construction of the recombinant vector pCambia3300-TaGly in Example 1;

[0034] Figure 3 This is the positive identification result of the transgenic wheat overexpressing wild-type Fielder and TaGly genes in Example 2;

[0035] Figure 4 The image shows the Fusarium head blight phenotype of wild-type Fielder and TaGly gene overexpressing transgenic wheat after inoculation with Fusarium graminearum in Example 3;

[0036] Figure 5The results show the statistical results of the disease spikelet rate of transgenic wheat overexpressing wild-type Fielder and TaGly genes in Example 3. Detailed Implementation

[0037] The present invention will now be described in further detail with reference to the accompanying drawings and specific embodiments.

[0038] Unless otherwise specified, the technical means used in the following embodiments are all conventional means well known to those skilled in the art, and the experimental methods without specific conditions are all conventional methods in the art.

[0039] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0040] The TaGly gene described in this invention is located within a susceptible QTL region on chromosome 2DS of Sumai 3. Ning 7840, a derivative line of Sumai 3, exhibits multiple allelic variations that can cause amino acid changes compared to the nucleic acid sequence of this gene in the Clark background (sequence alignment is as follows). Figure 1 As shown, N7840 represents Ning 7840, and the red arrow indicates the nucleic acid mutation site. Therefore, this invention extracts the DNA of Ning 7840, amplifies the DNA sequence of the TaGly gene, and performs vector construction, genetic transformation, and Fusarium head blight phenotype identification. The full-length nucleotide sequence of the TaGly gene is shown in SEQ ID NO.1 (1869 bp in length), containing a complete coding region. The coding region sequence is shown in SEQ ID NO.2 (1401 bp in length, including a stop codon), encoding 466 amino acids. The corresponding protein amino acid sequence is shown in SEQ ID NO.3.

[0041] Example 1: Cloning and Overexpression Vector Construction of the Wheat TaGly Gene

[0042] 1. Extraction of wheat genomic DNA

[0043] Young leaves of wheat variety Ning 7840 were ground into powder in a mortar using liquid nitrogen. The powder was transferred to a 1.5 mL centrifuge tube, and 650 μL of preheated CTAB extraction buffer (100 mM Tris-HCl pH=8.0, 4 mol / L NaCl, 20 mmol / L EDTA pH=8.0, 2% CTAB, with 2% β-mercaptoethanol added before use) was added. The mixture was then incubated in a 65℃ water bath for 30 min, with gentle shaking several times during incubation. After cooling on ice for 5 min, 400 μL of chloroform was added, and the mixture was gently mixed. The mixture was allowed to stand at room temperature for 10 min, and then centrifuged at 12000 rpm for 15 min. The supernatant was collected. Two volumes of ice-cold ethanol were added, and the mixture was incubated at -20℃ for 30 min. The supernatant was discarded after centrifugation. The precipitate was washed twice with 70% ethanol (1 mL), dried, and dissolved in 100 μL of sterile water to obtain genomic DNA.

[0044] 2. TaGly gene amplification

[0045] Using Ning 7840 genomic DNA as a template, PCR amplification was performed using the following primers (synthesized by Nanjing GenScript Biotech):

[0046] Forward primer (SEQ ID NO.5): 5'-CCCAAGCTTATGGAGAGCACGGGCCA-3';

[0047] Reverse primer (SEQ ID NO.6): 5'-CGGGATCCCATTGTTGTAAATTCAAATTGACAATACTTG-3'.

[0048] PCR reaction system (50 μL): 25 μL 2×KOD Buffer, 4 μL dNTPs, 0.5 μL KOD enzyme, 1.5 μL each of forward and reverse primers, 4 μL template, and ddH2O to 50 μL. Reaction program: 94℃ pre-denaturation for 3 min; 94℃ denaturation for 30 s, 58℃ annealing for 30 s, 68℃ extension for 3 min, for a total of 30 cycles; final extension at 68℃ for 10 min; product stored at 4℃. The amplified product was detected by 1% agarose gel electrophoresis, and a specific band appeared at approximately 1869 bp. This band corresponds to the full-length genome sequence of the TaGly gene (SEQ ID NO.1), and its coding region (CDS) sequence is shown in SEQ ID NO.2.

[0049] 3. Agarose gel purification of PCR products

[0050] Prepare a 2% agarose gel using TAE electrophoresis buffer and add 1% (v / v) of 0.5 mg / mL EB. Add all PCR products to 5 µL of 10× loading buffer and load the gel. Use Trans2k DNA Marker as the molecular weight standard and perform electrophoresis at 130 V for 15 min. Under UV light using a gel imaging system (Bio-RAD), cut a gel block matching the size of the target fragment and place it in a 2.0 mL centrifuge tube. Recover the target fragment according to the gel extraction kit (Sangon Biotech, China) instructions to obtain the purified PCR product.

[0051] 4. Ligate the cloning vector pEASY-Blunt3

[0052] The purified PCR product was ligated into the pEASY-Blunt3 cloning vector. Ligation mixture (5 μL): 4 μL recovered fragment, 1 μL pEASY-Blunt3 vector. Incubate at 25°C for 30 min.

[0053] 5. Transformation of E. coli competent cells

[0054] Thaw the Fast-T1 competent cell suspension on ice. Mix 50 μL of competent cells with the ligation product described above, aspirate and mix thoroughly, then incubate on ice for 30 min. Next, heat shock the cells in a metal bath at 42°C for 1.5 min, followed by an immediate ice bath for 3–5 min. Add 1 mL of LB liquid medium, aspirate and mix thoroughly, then incubate on a shaker at 37°C and 150 rpm for 1 h. After incubation, centrifuge at 5000 rpm for 5 min, discard the supernatant, and gently aspirate and mix the precipitate with the remaining liquid using a pipette. Spread the mixture onto LB solid medium containing ampicillin (Amp). Once the plate surface is free of obvious water stains, seal the plate and invert it for overnight incubation at 37°C.

[0055] 6. Screening and sequencing verification of positive clones

[0056] Ten single colonies were randomly selected from LB agar medium for colony PCR identification. The PCR reaction mixture (10 μL) consisted of 5 μL of 2×Taq Master Mix, 0.25 μL of forward primer (SEQ ID NO.5), 0.25 μL of reverse primer (SEQ ID NO.6), and 4 μL of ddH2O, using a single colony as a template. The PCR program was as follows: 95℃ pre-denaturation for 3 min; 95℃ denaturation for 30 s, 58℃ annealing for 30 s, 72℃ extension for 3 min, for a total of 28 cycles; final extension at 72℃ for 5 min. The products were detected by 1% agarose gel electrophoresis, and positive clones were screened.

[0057] Positive clones were inoculated into LB liquid medium containing ampicillin and cultured overnight at 37°C and 180 rpm. The bacterial culture was then sent to a sequencing company (Genewiz, Suzhou) for sequencing. The sequencing results were compared with the target sequence using DNAMAN software for verification. For correctly sequenced strains, plasmids were extracted according to the instructions of a plasmid DNA mini-extraction kit (Sangon Biotech, China) to obtain recombinant cloning plasmids containing the sequence shown in SEQ ID NO. 1.

[0058] 7. Construction of overexpression vectors

[0059] The recombinant cloning plasmid was double-digested with restriction endonucleases HindIII and BamHI. The digestion mixture (50 μL) consisted of: 1 μL HindIII, 1 μL BamHI, 2.5 μL 10×QCut Buffer, 10 μL plasmid, and ddH2O to a final volume of 50 μL. The reaction was carried out at 37°C for 2 h. The digestion products were separated by agarose gel electrophoresis, and the TaGly gene fragment was recovered according to the gel recovery kit (Sangon Biotech, China).

[0060] Simultaneously, the plant expression vector pCambia3300 (backbone vector) was digested with the same restriction endonucleases HindIII and BamHI, and the linearized vector was recovered. The target gene fragment and the linearized vector were mixed at a ratio of 6:3, and the ligation system (10 μL) consisted of: 1 μL 10×T4 Buffer, 1 μL T4 Ligase, 6 μL target gene, and 3 μL target vector. Ligation was carried out at 25°C for 2 h.

[0061] The ligation product was transformed into *E. coli* Fast-T1 competent cells and plated on LB agar plates containing ampicillin, then incubated overnight at 37°C. Twenty single colonies were randomly selected from each LB agar plate for colony PCR identification. Positive clones were screened, and plasmids were extracted for restriction enzyme digestion and sequencing verification to obtain the recombinant vector pCambia3300-TaGly, whose full nucleotide sequence is shown in SEQ ID NO.4 (12736 bp in length).

[0062] The constructed carrier diagram is as follows Figure 2 As shown, the TaGly gene in this vector is driven by the strong CaMV 35S (Tobacco Mosaic Virus 35S) promoter and contains the BAR gene (which can confer herbicide resistance to plants) as a plant selection marker.

[0063] 8. Agrobacterium-mediated transformation

[0064] The verified recombinant vector was transformed into Agrobacterium EHA105 competent cells using the freeze-thaw method. The cells were then plated on YEB plates containing the corresponding antibiotics and cultured at 28°C for 2 days. Single clones were picked for PCR identification, and positive clones were used for subsequent wheat genetic transformation.

[0065] Example 2: Creation and positive identification of TaGly overexpression transgenic wheat

[0066] 1. Wheat genetic transformation

[0067] Using the wheat variety Fielder as the recipient material, transgenic plants were created using Agrobacterium-mediated embryo genetic transformation. The recombinant vector pCambia3300-TaGly constructed in Example 1 was transformed into Agrobacterium EHA105 competent cells via electroporation. Positive clones were obtained by PCR identification and used for transformation. Immature wheat seeds, 12-14 days after pollination, were surface-sterilized with 70% ethanol for 1 min, sterilized with 20% sodium hypochlorite solution for 15 min, and rinsed 3-4 times with sterile water. Under aseptic conditions, immature embryos with a diameter of approximately 1.5-2.0 mm were extracted and placed in 2 mL plastic centrifuge tubes containing 1.8 mL of suspension. Approximately 150 immature embryos were processed within 30 min. The suspension was removed, and 1.0 mL of Agrobacterium suspension was added. The mixture was incubated at room temperature for 5 min, then the embryos were gently resuspended and poured onto a co-culture medium. Excess Agrobacterium suspension on the surface was removed with a pipette, and the mixture was co-cultured in the dark at 23°C for 2 days. After co-culture, the immature embryos were transferred to recovery medium and cultured in the dark at 28°C for 8 days. They were then transferred to differentiation medium containing glufosinate resistance and cultured at 25°C under 5000 lx light for 3 weeks for resistant callus screening and differentiation. The differentiated seedlings were transferred to rooting medium containing glufosinate and cultured at 25°C under 5000 lx light until rooting. Well-rooted seedlings were then transferred to small pots for further growth. Once the plants reached a certain stage, they were transplanted into a greenhouse, and the offspring seeds were harvested after 3-4 months. Four independent transformant lines were obtained, numbered 1#, 2#, 3#, and 4#.

[0068] 2. Positive identification of genetically modified organisms

[0069] Young leaves from the four transformed lines and the wild-type Fielder were collected, and genomic DNA was extracted using the CTAB method. Using the extracted genomic DNA as a template, PCR amplification and detection were performed using the following primers:

[0070] Forward primer (SEQ ID NO.7): 5'-TTAGCCCTGCCTTCATACGAT-3';

[0071] Reverse primer (SEQ ID NO.8): 5'-AGACGAACAAGACTCACCTTTGG-3'.

[0072] PCR reaction mixture (10 μL): 5 μL 2×Taq Master Mix, 0.25 μL each of forward and reverse primers, 1 μL template DNA, and ddH2O to a final volume of 10 μL. Reaction program: 95℃ pre-denaturation for 3 min; 95℃ denaturation for 30 s, 58℃ annealing for 30 s, 72℃ extension for 1 min, for a total of 35 cycles; final extension at 72℃ for 5 min. Amplified products were detected by 2% agarose gel electrophoresis.

[0073] The results are as follows Figure 3 As shown, the four transgenic lines 1#, 2#, 3#, and 4# all amplified bands of the expected size, while the wild-type Fielder showed no band, proving that the TaGly gene has been successfully integrated into the genomes of the four transgenic lines.

[0074] Example 3: Identification of Fusarium head blight resistance in TaGly overexpressing transgenic wheat

[0075] 1. Preparation of Fusarium graminearum spore liquid

[0076] The Fusarium graminearum used in this embodiment ( Fusarium graminearum Wild-type strain PH-1 (also known as NRRL 31084) was isolated from wheat grains in a wheat field in Michigan, USA. It is a whole-genome sequenced strain and can be publicly obtained through strain collection centers such as the American Type Culture Collection (ATCC) (accession number ATCC MYA-4620) or the American NRRL (accession number NRRL 31084).

[0077] Fusarium graminearum strain PH-1 was inoculated onto PDA solid medium and activated by incubation at 25°C for 5 days. Four mycelial blocks were then collected using a sterile punch and transferred to 50 mL of mung bean soup liquid medium, and incubated at 25°C with shaking at 150 rpm for 3–5 days. Conidia were collected, and 1 µL of the spore suspension was dropped onto a hemocytometer. The spores were observed and counted under a microscope to determine the spore concentration. The spore concentration was adjusted to 1 × 10⁻⁶ with sterile water. 5 Quantity / mL, for later use.

[0078] 2. Inoculation and Phenotypic Identification

[0079] During the wheat flowering stage, the double-floret drip inoculation method was used. Wheat ears from identical transgenic lines (OE#1, OE#2, OE#3, OE#4) and the recipient variety Fielder (control) were selected, with at least 7 ears inoculated from each line. Using a microsyringe, 10 μL of the above spore solution was injected between the inner and outer glumes of the bilateral florets of the fifth spikelet in the middle of the ear, and marked accordingly. After inoculation, the ears were sealed in bags to maintain moisture for 3 days, and disease incidence was assessed 14 days later. The percentage of symptom spikelets (PSS) was calculated using the following formula:

[0080] Diseased spikelet rate = (Number of diseased spikelets / Total number of spikelets) × 100%

[0081] 3. Results Analysis

[0082] Phenotypic identification and statistical results of Fusarium head blight are as follows: Figure 4 , 5 As shown, the disease incidence rate of wild-type Fielder wheat 14 days after inoculation with Fusarium graminearum was approximately 41%. The disease incidence rates of the four TaGly overexpressing transgenic lines were: OE#1 line 68%, OE#2 line 57%, OE#3 line 78%, and OE#4 line 62%, all significantly higher than the wild-type control (P < 0.05). These results indicate that overexpression of the TaGly gene can significantly increase wheat susceptibility to Fusarium head blight, confirming that the TaGly gene negatively regulates wheat resistance to Fusarium head blight.

[0083] Based on the functional verification of the TaGly gene's negative regulation of wheat resistance to Fusarium head blight, those skilled in the art can anticipate that knocking out or disrupting this gene using gene editing technologies such as CRISPR / Cas9 can effectively improve wheat resistance to Fusarium head blight. For example, an sgRNA targeting the sequence shown in SEQ ID NO.1 can be designed, a gene editing vector can be constructed, wheat can be transformed using Agrobacterium-mediated transformation, and homozygous knockout mutants can be obtained through sequencing identification. It is expected that the knockout mutants, after inoculation with Fusarium graminearum, will have a significantly lower rate of diseased spikelets compared to the wild type. Those skilled in the art, based on the teachings of this invention and combined with conventional techniques in the field, can achieve the above technical solutions and obtain the expected technical effects.

[0084] The embodiments described above are only some, not all, of the embodiments of the present invention. The detailed description of the embodiments of the present invention is not intended to limit the scope of the claimed invention, but merely to illustrate selected embodiments. The scope of protection of the present invention is determined by the scope claimed in the claims. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention.

Claims

1. A TaGly gene that negatively regulates wheat scab resistance, characterized in that, The nucleotide sequence of the gene is as follows (A1) or (A2): (A1) The DNA sequence shown in SEQ ID NO.1; (A2) The coding region is the DNA sequence shown in SEQ ID NO.

2.

2. A TaGly protein encoded by the TaGly gene as described in claim 1, characterized in that, The amino acid sequence of the protein is any one of the following (B1) to (B4): (B1) The amino acid sequence as shown in SEQ ID NO.3; (B2) A derivative protein having the same function by substitution, deletion and / or addition of one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3; (B3) is a protein that has more than 80% homology with the amino acid sequence defined in (B1) or (B2) and has the same function; (B4) A fusion protein obtained by attaching a tag to the N-terminus and / or C-terminus of any of the proteins defined in (B1) to (B3).

3. A recombinant vector containing the TaGly gene as described in claim 1.

4. The recombinant vector according to claim 3, characterized in that, The nucleotide sequence of the recombinant vector is shown in SEQ ID NO.4, which is obtained by inserting the nucleotide sequence shown in SEQ ID NO.1 between the HindIII and BamHI multiple cloning sites of the pCambia3300 backbone vector.

5. A host cell containing the recombinant vector as described in claim 3 or 4.

6. The application of the TaGly gene as described in claim 1, the TaGly protein as described in claim 2, the recombinant vector as described in claim 3 or 4, or the host cell as described in claim 5 in the negative regulation of wheat scab resistance.

7. The application of the TaGly gene as a negative regulatory target in improving wheat scab resistance, as described in claim 1.

8. A breeding method for improving wheat resistance to Fusarium head blight, characterized in that, Includes the following steps: The expression of the TaGly gene as described in claim 1 in wheat is suppressed or knocked out, or the susceptible allelic variant of the TaGly gene is eliminated, by means of gene editing, RNA interference, or marker-assisted selection.

9. A breeding method for improving wheat resistance to Fusarium head blight according to claim 8, characterized in that, The gene editing was performed using the CRISPR / Cas9 system to target and knock out the TaGly gene.

10. A method for identifying wheat scab resistance, characterized in that, Includes the following steps: The wheat's resistance to Fusarium head blight is determined by detecting the expression level or allelic variation type of the TaGly gene as described in claim 1: when the TaGly gene is highly expressed or there is a susceptible allelic variation, the wheat is susceptible; when the TaGly gene is knocked out or there is a resistant allelic variation, the wheat is resistant.

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