Application of PPTase protein and its encoding gene in the preparation of drugs for preventing and controlling plant scab

Abstract

The present invention provides a potential drug target protein PPTase and its encoding gene for use in the preparation of drugs for preventing and treating plant fusarium rust. ppt1 After the gene was removed, the production of GA3, GA4, and GA7 in gibberellins decreased significantly; at the same time, the growth ability of the knockout bacteria on PDA, MM, SM and other media decreased significantly. Therefore, inhibitor drugs designed with proteins as targets at this site can prevent and control diseases caused by gibberellin-producing fungi in plant fungi, providing a new way to screen fungal drugs for the prevention and control of plant fusarium rust.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to the application of PPTase protein and its encoding gene in the preparation of drugs for the prevention and control of plant scab. Background Technology

[0002] When rice, wheat, corn, and other plants are infected with pathogenic fungi such as Fusarium graminearum, Fusarium graminearum, and Fusarium moniliforme, they will exhibit rampant growth. Diseased plants are often more than 50% taller than normal plants, and the seed setting rate is greatly reduced. They are often accompanied by yellowing and withering symptoms, hence the name "bad seedling disease" or "Fusarium head blight".

[0003] The primary reason for the excessive height and uncontrolled growth of plants is generally considered to be the secretion of a plant growth hormone called gibberellin by pathogens affecting this type of crop. Gibberellins, chemically classified as diterpenoid acids, are a widely distributed class of plant hormones. Currently, bioactive gibberellins GA1, GA3, GA4, and GA7 have been identified. They act on cash crops including wheat, corn, and sugarcane, causing a series of problems such as yield reduction and death, seriously jeopardizing the development of my country's agricultural economy.

[0004] Studies have found that the mechanism by which crops such as rice and corn exhibit excessive height growth is that excessive gibberellin causes an abnormal increase in auxin within the plant, leading to accelerated cell division and excessive elongation, ultimately resulting in bakanae disease in rice and other crops. Fusarium head blight significantly impacts and hinders the development of my country's agricultural economy, making the effective prevention and control of Fusarium head blight infection in crops a focal point in this field.

[0005] In 2020, wheat scab was listed as a Class I crop disease and pest in the "List of Class A Crop Diseases and Pests" by the Ministry of Agriculture and Rural Affairs of China. Currently, the main control measure for scab relies on chemical fungicides sprayed during the flowering period. However, whether using a single fungicide or a combination of fungicides, pathogens easily develop resistance after a period of use, significantly reducing the effectiveness of chemical fungicides. In addition, to avoid the adverse effects of diseases and pests on crop yields, scientists are also conducting research on the breeding of biological control plant varieties. However, due to the long development cycle of antimicrobial varieties and the safety concerns of genetically modified crops, the pesticide market urgently needs to develop targeted drugs with high specificity that are less likely to induce resistance in pathogens.

[0006] The number of proteins that can serve as drug targets within the eukaryotic cells of pathogens is extremely small, and the number of fungicide molecular targets with high drug sensitivity and selectivity is even smaller. As a result, many major crops lack effective drug control, so the pesticide market urgently needs to explore new targets with control effects. Summary of the Invention

[0007] The purpose of this invention is to provide the application of PPTase protein and its encoding gene in the preparation of drugs for the prevention and control of plant scab.

[0008] The technical solution adopted in this invention is:

[0009] Application of PPTase (4'-phosphopantoylthioethylaminotransferase) protein as a target protein for fungal drugs in the preparation of drugs to control Fusarium head blight.

[0010] Specifically, the application involves designing inhibitory drugs that target the fungal drug target protein. For example, designing inhibitors targeting the specific protein site. ppt1 CRISPR / ZNF system with gene mutations, expression repression ppt1 Methods such as siRNA of genes can reduce or eliminate PPTase expression; or drug screening can be used to obtain inhibitors of PPTase protein activity, ultimately reducing the yield of GA3, GA4 and GA7 in gibberellins. This can further reduce a series of problems caused by gibberellins acting on economic crops such as wheat, corn and sugarcane, including yield reduction and death.

[0011] Preferably, the amino acid sequence of the fungal drug target protein is shown in SEQ ID No. 1.

[0012] Phosphopantoylthioethylamine transferase (PPTase) catalyzes post-translational modification reactions of proteins such as fatty acid synthase, polyketide synthase, acyl carrier proteins of nonribosomal peptide synthase, and peptidyl carrier proteins. It transfers the phosphopantoylthioethylamine group on coenzyme A to a conserved serine residue in the above proteins, thereby synthesizing fatty acids, polyketides, and nonribosomal peptides.

[0013] This invention also relates to ppt1 Application of genes in the preparation of drugs to control Fusarium head blight in plants.

[0014] Specifically, the application is as follows: [The application is for the following]. ppt1 Gene design enables ppt1 Inhibitor drugs that suppress or reduce gene expression.

[0015] Preferably, the nucleotide sequence of the gene encoding the fungal drug target protein is shown in SEQ ID No. 2.

[0016] The plants mentioned include: rice, wheat, corn, cucumber, etc.

[0017] This invention is based on the study of factors affecting the overall metabolic flux of gibberellin. ppt1 The study focused on the knockout of the PPTase gene, which is associated with post-translational modifications of fungal lysine metabolism, polyketides, and non-ribosomal peptides. Experiments showed that knocking out the PPTase gene in *Fusarium oxysporum*... ppt1 After gene knockout, the yield of GA3 in gibberellin decreased by 70%-95%, the yield of GA4 decreased by 50%-80%, and the yield of GA7 also decreased by 80%-95%; at the same time, the growth ability of the knockout bacteria on PDA, MM, SM and other media decreased significantly.

[0018] The aforementioned ppt1 Genes and PPTase proteins are widely present in fungi such as *Fusarium graminearum*, *Fusarium moniliforme*, and *Fusarium fusiforme*, which are known to cause Fusarium head blight in plants. Therefore, inhibitors targeting these sites could be designed to control diseases caused by gibberellin-producing fungi in plants.

[0019] The plants mentioned include: rice, wheat, corn, cucumber, etc. The plant pathogens that cause Fusarium head blight include: Fusarium graminearum, Fusarium moniliforme, and Fusarium fusiforme.

[0020] Compared with the prior art, the beneficial effects of the present invention are mainly reflected in:

[0021] This invention provides the application of a potential drug target protein, PPTase, and its encoding gene in the preparation of drugs for controlling Fusarium head blight in plants. Experiments have shown that knocking out PPTase in *Fusarium oxysporum*... ppt1 Following gene deletion, the production of GA3, GA4, and GA7 in gibberellins all decreased significantly; simultaneously, the growth capacity of the knockout bacteria on PDA, MM, and SM media was also greatly reduced. Therefore, inhibitors targeting this site could be designed to control diseases caused by gibberellin-producing fungi in plants, providing a new approach for screening fungal drugs resistant to Fusarium head blight. Attached Figure Description

[0022] Figure 1 This is a schematic diagram of the construction of Pcas-ppt1 containing Cas9 protein in an embodiment of the present invention;

[0023] Figure 2 It contains a hygR-containing antibody and a donor fragment with 700bp homologous arms above and below it;

[0024] Figure 3 For two ppt1 PCR pairs of knockout bacteria ppt1 Upstream and downstream validation gel plots; in the plot, up and down indicate validation of... ppt1 The original fragments at upstream and downstream positions of the gene are knocked out and replaced with donor DNA fragments;

[0025] Figure 4 To determine the yields of GA3, GA4, and GA7 by high performance liquid chromatography. Detailed Implementation

[0026] The present invention will be further described in detail below with reference to specific examples, but the present invention is not limited to the following embodiments:

[0027] Example 1:

[0028] Fumigranobacter fusiforme ppt1 Gene knockout:

[0029] To verify the biological function of PPTase protein, CRISPR / Cas9 technology was used to study *Fujikura scab*. ppt1 Gene knockout was performed. The Pcas-ppt1 vector and donor fragment were introduced into the bacterial strain, and mutant strains were obtained through colony PCR verification and screening. The Pcas-ppt1 vector contains DNA elements such as targeting gRNA, Cas9 protein expression cassette, and hygromycin resistance selection tags (e.g., Figure 1 As shown). The donor fragment is preferably constructed using a multi-fragment fusion method (e.g. Figure 2 (As shown).

[0030] The *Fujikura fusarium* species were obtained from GenBank accession number HE614113.1. ppt1 The gene sequence shows high homology and similar activity across different fungi. ppt1 After selecting sites to design gRNAs, the plasmid pUC-fFuCas9-HTB was used. NLS -hph was digested with the restriction endonuclease EcoRI, and then the synthesized gRNA fragment (containing a 20 bp PAM sequence) was ligated to the digested plasmid using a one-step cloning kit. The recombinant plasmid was then transformed into DH5α E. coli for storage and later use.

[0031] The amino acid sequence of the PPTase protein (derived from *Fujikura fusiforme*) is shown in SEQ ID NO.1:

[0032] MSRAQSSPTVIQWVIDTRPLWPSALKTKDLTSAASRALSLLTEEEQSSVLRYYHVRDAKLALASALLKRYAISRFCHVPWFLAKTTRDARTKPVFVLPSGDEPLIFNVSHQAGLAVLLAVHDPPKGLAVGVDVVCPSERRDRDLSS LEEDGWASFVDIHADVFGAGEVSALKSMNPVPTVQERDRALRYFYALWCLREAYVKMTGDALLASWLKDLEMHNFAPPEDMKEAQEVRLRGKKVEGVDMRLMPLLEEYMVSTAVRNGDNGESVELGEFQSLDLEEILAFGEQASKP.

[0033] The gene for the drug target PPTase protein (derived from *Fujikura fusiforme*) has the nucleotide sequence shown in SEQ ID NO. 2:

[0034] ATGAGTAGGGCCCAATCCTCACCTACGGTCATCCAATGGGTGATCGACACTCGTCCACTTTGGCCCTCCGCTCTCAAGACCAAAGACCTCACATCCGCTGTAAGTGCTCTGGTCATCCACTTTAACCTGATACTCATGTCATACCTTAGGCTTCACGTGCACTCTCACTTCTCACCGAGGAGGAACAATCTTCTGTCCTGAGATACTACCACGTCCGCGATGCCAAGCTCGCTCTCGCTTCGGCTCTTCTTAAGCGCTACGCAATCTCTCGCTTCTGTCACGTCCCCTGGTTCCTGGCCAAGACTACGCGAGATGCCCGTACTAAGCCCGTGTTTGTTTTGCCCAGCGGGGATGAGCCGCTGATCTTTAACGTCTCGCATCAGGCCGGTCTCGCGGTCCTTCTGGCCGTCCATGATCCGCCAAAGGGCCTTGCCGTGGGAGTTGATGTTGTCTGTCCGTCAGAGCGACGAGATCGAGATCTGAGTAGCTTAGAAGAAGATGGGTGGGCGAGTTTCGTCGACATTCATGCCGATGTCTTTGGAGCGGGAGAAGTCTCAGCGCTTAAGAGCATGAACCCTGTTCCTACCGTCCAAGAACGAGATCGTGCATTGCGCTACTTTTATGCGCTATGGTGTCTCCGTGAAGCCTATGTCAAGATGACAGGGGACGCTCTTCTGGCAAGCTGGTTGAAAGACCTAGAGATGCACAACTTTGCCCCGCCAGAAGACATGAAGGAAGCCCAAGAGGTTCGACTGAGAGGCAAGAAGGTTGAGGGCGTGGACATGAGACTGATGCCGCTACTTGAGGAATACATGGTCTCCACAGCTGTAAGAAACGGTGATAATGGGGAGAGTGTCGAGCTCGGAGAGTTTCAAAGTCTTGATCTGGAGGAAATATTGGCATTTGGTGAACAGGCATCCAAACCATGATCAGAGAACGTTTTAT。

[0035] Construction of the donor fragment containing the hygR resistance screening tag is as follows:

[0036] Amplification was achieved using three pairs of upstream and downstream primers: F1, R1; F2, R2; and F3, R3. ppt1 Upstream homologous regions of genes, hygR resistance genes and ppt1 Downstream homologous arm region of the gene.

[0037] F1: GTTCCATCCACTCAGATCCGG (SEQ ID NO.3)

[0038] R1: CAGGCTTTTTCATTTACTGCAACTATTGCCTTGGAAT (SEQ ID NO.4)

[0039] F2: GCAGTAAATGAAAAAAGCCTGAACTCACCG (SEQ ID NO.5)

[0040] R2: TTCCTATTCCTTTGCCCTCGGACG (SEQ ID NO.6)

[0041] F3: CGAGGGCAAAGGAATAGGAATAATGAGAAACGCAAGCGC (SEQ ID NO.7)

[0042] R3: AGAAGTACAAGGTCTTCATCCCTGA (SEQ ID NO.8)

[0043] The total PCR reaction volume is 50 μL. Fusarium fujikuroi 978 # 2 uL of whole genome template, 25 uL of 2×PhantaMix Buffer, 2 uL each of forward and reverse primers, 1 uL of dNTP Mix, 1 uL of Phanta Mix Super-Fidelity, and ddH2O to make up to 50 uL (the reagents used in the PCR reaction were purchased from Novizan Biosciences Co., Ltd.).

[0044] The PCR reaction conditions were: 95°C for 10 min, (95°C for 30 s, 60°C for 30 s, 72°C for 1 min, 30 cycles) followed by a 72°C extension for 10 min.

[0045] The three obtained fragments were purified using an agarose gel extraction kit and used as templates. Fusion PCR was performed on the three fragments using F1 and R3 primers to obtain the donor fragment. Fusion PCR reaction conditions: 95°C for 10 min, (95°C for 30 s, 60°C for 30 s, 72°C for 2 min 10 s, 30 cycles) followed by a 72°C extension for 10 min.

[0046] Protoplast preparation method:

[0047] Use a sterile inoculation spatula to scoop 1 cm of the eggplant flask slant culture medium. 2 The bacterial blocks of different sizes were transferred to 25 mL of YEPD medium (YEPD medium: yeast extract 30 g / L, peptone 10 g / L, glucose 20 g / L) and cultured at 250 rpm and 28°C for 2 days. Transfer 1 mL of bacterial culture from YEPD medium to an EP tube (Eppendorf), centrifuge at 12000 rpm for 3 min, and discard the supernatant. Add 1 mL of sterile water (ultrapure water sterilized) to the EP tube and mix thoroughly. Centrifuge at 12000 rpm for 3 min, and discard the supernatant. Add 1 mL of sterile 0.8M NaCl solution to the EP tube and mix thoroughly. If bacterial balls are present, remove them with a pipette. Centrifuge at 12000 rpm for 3 min, and discard the supernatant. Add 1 mL of sterile cell wall digestion solution (prepared with 1% Driselase, 2% Yatalase, 1% Snailase, and 0.8M NaCl solution) to the EP tube and mix thoroughly. Incubate in a 150 rpm, 30°C water bath for 60-90 min. The specific time depends on the bacterial concentration and the number of bacterial balls. Centrifuge the digested bacterial culture at 4500 rpm for 3 min, and transfer the supernatant to a container. Add an equal volume of 0.8M NaCl to a sterile EP tube, mix thoroughly, centrifuge at 4500 rpm for 3 min, and discard the supernatant; add 1 mL of 0.8M NaCl to the EP tube, mix thoroughly, centrifuge at 4500 rpm for 3 min, and discard the supernatant; wash once with 0.8M NaCl, discard the supernatant, add 20 μL of STC solution, and store at 4℃ for later use.

[0048] Protoplast transformation methods:

[0049] The constructed Pcas-ppt1 plasmid and the donor fragment containing hygR and 700bp upper and lower homologous arms were transformed and integrated using a PEG-CaCl2-mediated chemical transformation method. Add 100 μL of protoplast suspension, 10 μg of the prepared Pcas-ppt1 plasmid, and 10 μg of donor fragment to a transformation tube, mix gently, and incubate on ice for 20 min. Add 50 μL of 60% PEG6000, incubate on ice for 20 min, then add 500 μL of 60% PEG6000 and 1 mL of STC solution and mix thoroughly. Add this transformation system to 10 mL of MYG semi-solid medium (MYG medium: maltose 5 g / L, yeast extract 5 g / L, glucose 10 g / L, sucrose 171 g / L, agar powder 15 g / L) cooled to room temperature and containing 100 μg / mL hygromycin resistance agar powder. Shake well and pour onto MYG plates with the same hygromycin resistance concentration. Incubate at 28°C for 4-5 days to obtain transformants Δ. ppt1 -14 and Δ ppt1 -18.

[0050] Verification of Fumigia fujikuro ppt1 Gene-deleted strains:

[0051] First, through growth defect verification, the knockout bacteria were cultured on SM medium (SM medium: glucose 20 g / L, KH2PO4 1 g / L, MgSO4·7H2O 0.5 g / L, KCl 2 g / L, FeSO4·7H2O 10 mg / L, agar powder 20 g / L, NaNO3 3 g / L) which does not contain any amino acids. The knockout bacteria could not grow on this medium and exhibited a lysine-deficient phenotype, while the wild-type bacteria could grow under the same conditions.

[0052] Secondly, the gene fragment deletion was verified by PCR. Primers F4 and R4, and F5 and R5 were designed as two pairs of primers for upstream and downstream verification. The resulting transformants were transferred to PDA plates (PDA medium: potato 200g / L, 20g / L, agar powder 15g / L) and cultured at 28℃ for 4-5 days. Genomic DNA was then extracted as templates for PCR verification. The results are shown in the figure. Figure 3 It is evident that *Fujikura gibberellinii* ppt1 The gene has been knocked out.

[0053] F4: CTAAGCACAGTAGCAATGCCTC (SEQ ID NO.9)

[0054] R4: CCACTATCGGCGAGTACTTCTAC (SEQ ID NO.10)

[0055] F5:TCTCCGACCTGATGCAGCTC (SEQ ID NO.11)

[0056] R5: GCTGCAACATACCTCCTGTTGTC (SEQ ID NO.12)

[0057] The total PCR reaction volume was 25 μL: 2 μL of genomic template extracted from each transformed single colony, 12.5 μL of 2 × Rapid TaqMaster Mix, 1 μL each of forward and reverse primers, and ddH2O to make up to 25 μL (the reagents used in the PCR reaction were purchased from Novizan Biosciences Co., Ltd.). The PCR reaction conditions were: 95°C for 3 min, (95°C for 15 s, 60°C for 15 s, 72°C for 40 s, 30 cycles), followed by a 72°C extension for 10 min.

[0058] Example 2:

[0059] Wild-type fungi ( Fusarium fujikuroi 978 # (Provided by Zhejiang Qianjiang Biochemical Co., Ltd.) Two plants served as the control group. ppt1 Gene knockout bacteria (Δ ppt1 -14 and Δ ppt1 -18) is the experimental group, and the growth status of the strain is evaluated.

[0060] The growth status of the strain was evaluated using three culture media: potato dextrose agar (PDA), two synthetic media (MM medium (20 g / L glucose, 1 g / L KH₂PO₄, 0.5 g / L MgSO₄·7H₂O, 2 g / L KCl, 10 mg / L FeSO₄·7H₂O, 20 g / L agar powder, 3 g / L NaNO₃), and SM medium). An additional 10 mM lysine was added to the synthetic medium.

[0061] Colony diameters were measured after the control and knockout strains were grown at 28°C for 7 days. The results are shown in Table 1. (PPTase gene deletion strains) Δppt1 Its growth capacity is greatly reduced.

[0062] Table 1: Colony growth

[0063]

[0064] Example 3: Gibberellin metabolism

[0065] The purpose of this invention is to identify the role of PPTase proteins from filamentous fungi in gibberellin synthesis, thereby demonstrating the feasibility of using these proteins as targets for agricultural antibiotics to control Fusarium head blight, and providing technical and theoretical support for drug development for Fusarium head blight. This embodiment evaluates the role of PPTase proteins in gibberellin synthesis by assessing the metabolic synthesis levels of three major gibberellins: GA3, GA4, and GA7.

[0066] The culture media used in the experiment included seed propagation medium (corn starch 20 g / L, sucrose 15 g / L, peanut powder 15 g / L, KH2PO4 1 g / L, MgSO4 1 g / L, soybean meal 3 g / L) and fermentation medium (corn starch 75 g / L, rice flour 87.5 g / L, soybean meal 5 g / L, peanut powder 5 g / L, KH2PO4 0.5 g / L, K2SO4 0.5 g / L, MgSO4·7H2O 0.11 g / L). The composition of the culture media was consistent with that of industrial fermentation media, simulating natural conditions. The carbon and nitrogen source components in the culture media were all derived from plants, which is similar to natural conditions.

[0067] The method for determining gibberellin metabolism is to use Gibberellin bacteria... ppt1 Gene knockout strains and wild-type 978 # The strain will grow to approximately 2 cm in diameter after 4 days of incubation at 28°C on a PDA plate. 2 Transformant agar blocks were inoculated into seed culture medium for expansion. After culturing at 28℃ and 250 rpm for 2 days, the culture was transferred to fermentation medium with an inoculation amount of 6.25%. After culturing at 28℃ and 250 rpm for 7 days, the supernatant of the bacterial culture was collected, and after reasonable dilution, the yields of GA3, GA4 and GA7 were determined by high performance liquid chromatography.

[0068] The gibberellin metabolism level of the wild-type strain was set to 100%, and the results are shown in Table 2. ppt1 The gene knockout strain showed a significant reduction in gibberellin production, which was significantly different from the control strain.

[0069] Table 2: ppt1 Effect of gene deletion strains on gibberellin production

[0070]

[0071] This invention demonstrates that the absence of PPTase leads to a significant decrease in the ability of strains to synthesize secondary metabolites (e.g., Figure 4 As shown in the figure, it effectively inhibited the synthesis of this metabolite. sequence list <110> Zhejiang University of Technology <120> Application of PPTase protein and its encoding gene in the preparation of drugs for the prevention and control of Fusarium head blight in plants. <160> 12 <170> SIPOSequenceListing 1.0 <210> 1 <211> 292 <212> PRT <213> Gibberella fujikuroi <400> 1 Met Ser Arg Ala Gln Ser Ser Pro Thr Val Ile Gln Trp Val Ile Asp 1 5 10 15 Thr Arg Pro Leu Trp Pro Ser Ala Leu Lys Thr Lys Asp Leu Thr Ser 20 25 30 Ala Ala Ser Arg Ala Leu Ser Leu Leu Thr Glu Glu Glu Gln Ser Ser 35 40 45 Val Leu Arg Tyr Tyr His Val Arg Asp Wing Lys Leu Wing Leu Wing Ser 50 55 60 Ala Leu Leu Lys Arg Tyr Ala Ile Ser Arg Phe Cys His Val Pro Trp 65 70 75 80 Phe Leu Ala Lys Thr Thr Arg Asp Ala Arg Thr Lys Pro Val Phe Val 85 90 95 Leu Pro Ser Gly Asp Glu Pro Leu Ile Phe Asn Val Ser His Gln Ala 100 105 110 Gly Leu Ala Val Leu Leu Ala Val His Asp Pro Pro Lys Gly Leu Ala 115 120 125 Val Gly Val Asp Val Val Cys Pro Ser Glu Arg Arg Asp Arg Asp Leu 130 135 140 Ser Ser Leu Glu Glu Asp Gly Trp Ala Ser Phe Val Asp Ile His Ala 145 150 155 160 Asp Val Phe Gly Ala Gly Glu Val Ser Ala Leu Lys Ser Met Asn Pro 165 170 175 Val Pro Thr Val Gln Glu Arg Asp Arg Ala Leu Arg Tyr Phe Tyr Ala 180 185 190 Leu Trp Cys Leu Arg Glu Ala Tyr Val Lys With Thr Gly Asp Ala Leu 195 200 205 Leu Ala Ser Trp Leu Lys Asp Leu Glu With His Asn Phe Ala Pro Pro 210 215 220 Glu Asp With Lys Glu Ala Gln Glu Val Arg Leu Arg Gly Lys Lys Val 225 230 235 240 Glu Gly Val Asp Met Arg Leu Met Pro Leu Leu Glu Glu Tyr Met Val 245 250 255 Ser Thr Ala Val Arg Asn Gly Asp Asn Gly Glu Ser Val Glu Leu Gly 260 265 270 Glu Phe Gln Ser Leu Asp Leu Glu Glu Ile Leu Ala Phe Gly Glu Gln 275 280 285 Here’s Dear Lys Pro 290 <210> 2 <211> 945 <212> DNA <213> Gibberella fujikuroi <400> 2 atgagtaggg cccaatcctc acctacggtc atccaatggg tgatcgacac tcgtccactt 60 tggccctccg ctctcaagac caaagacctc acatccgctg taagtgctct ggtcatccac 120 tttaacctga tactcatgtc ataccttagg cttcacgtgc actctcactt ctcaccgagg 180 aggaacaatc ttctgtcctg agatactacc acgtccgcga tgccaagctc gctctcgctt 240 cggctcttc tagcgctac gcaatctctc gcttctgtca cgtcccctgg ttcctggcca 300 agactacgcg agatgcccgt actaagcccg tgtttgtttt gcccagcggg gatgagcgc 360 tgatctttaa cgtctcgcat caggccggtc tcgcggtcct tctggccgtc catgatcgc 420 caaagggcct tgccgtggga gttgatgttg tctgtccgtc agagcgacga gatcgagatc 480 tgagtagctt agaagaagat gggtgggcga gttcgtcga cattcatgcc gatgtctttg 540 gagcgggaga agtctcagcg cttaagagca tgaaccctgt tcctaccgtc caagaacgag 600 atcgtgcatt gcgctacttt tatgcgctat ggtgtctccg tgaagcctat gtcaagatga 660 caggggacgc tcttctggca agctggttga aagacctaga gatgcacaac tttgccccgc 720 cagaagacat gaaggaagcc caagaggttc gactgagagg caagaaggtt gagggcgtgg 780 acatgagact gatgccgcta cttgaggaat acatggtctc cacagctgta agaaacggtg 840 ataatgggga gagtgtcgag ctcggagagt ttcaaagtct tgatctggag gaaatattgg 900 catttggtga acaggcatcc aaaccatgat cagagaacgt tttat 945 <210> 3 <211> 21 <212> DNA <213> Unknown <400> 3 gttccatcca ctcagatccg 21 <210> 4 <211> 37 <212> DNA <213> Unknown​​​​​​​​​​​​​​​​​​​​​<212> DNA <213> Unknown <400> 6 ttcctattcc tttgccctcg gacg 24 <210> 7 <211> 39 <212> DNA <213> Unknown <400> 7 cgagggcaaa ggaataggaa taatgagaaa cgcaagcgc 39 <210> 8 <211> 25 <212> DNA <213> Unknown <400> 8 agaagtacaa ggtcttcatc cctga 25 <210> 9 <211> 22 <212> DNA <213> Unknown <400> 9 ctaagcacag tagcaatgcc tc 22 <210> 10 <211> 23 <212> DNA <213> Unknown <400> 10 ccactatcgg cgagtacttc tac 23 <210> 11 <211> 20 <212> DNA <213> Unknown <400> 11 tctccgacct gatgcagctc 20 <210> 12 <211> 23 <212> DNA <213> Unknown <400> 12 gctgcaacat acctcctgtt gtc 23

Claims

1. Application of PPTase protein as a fungal drug target protein in the preparation of a drug for preventing and treating plant fusarium fusca caused by Fusarium fusca, characterized in that: The amino acid sequence of the fungal drug target protein is shown in SEQ ID No. 1, and the application is to design an inhibitor drug with protein as the target for the fungal drug target protein.

2. ppt1 Application of the gene in the preparation of a drug for preventing and treating plant fusarium fusarium caused by fusarium fusarium, the ppt1 The gene nucleotide sequence is shown in SEQ ID No. 2, and the application is: for the ppt1 Genetic design ppt1 Drugs that inhibit the loss or reduction of gene expression.

3. The use according to claim 1 or 2, characterized in that The plants include: rice, wheat, corn and cucumber.

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