Tomato endogenous immune peptide and application thereof in plant immune excitation

Abstract

The invention provides a tomato endogenous immune peptide and application thereof in plant immune excitation, and belongs to the technical field of plant immunity. The tomato endogenous immune peptide TIP12 (SEQ ID NO.1) is found in tomatoes for the first time, and the tomato endogenous immune peptide TIP12 (SEQ ID NO.1) can activate the immune response of plants, so that the systematic resistance of the plants to pathogens is triggered. Moreover, it is found that TIP12 participates in regulation and control of various immune-related pathways, has a wide functional effect in the plant field, and shows a broad-spectrum effect in the aspect of enhancing multi-species immunity. Therefore, the tomato endogenous immune peptide TIP12 provided by the invention can be used as a widely effective plant immune excitant, and has good safety and application prospects.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of plant immunity, and particularly relates to a tomato endogenous immune peptide and application thereof in plant immune stimulation. BACKGROUND

[0002] Plants face biotic stresses such as microbial pathogens and insect pests during growth, and have evolved a complex immune system. Unlike animals, plants lack the ability to move and specialized adaptive immune cells, and rely on innate immune mechanisms present in every cell to defend against pathogens. Plant immune responses are divided into two levels: pathogen-associated molecular pattern (PAMP) triggered immunity (PTI) and effector triggered immunity (ETI), which together form the basis of defense. PTI is the first line of defense against pathogens, triggered by PAMPs, which are common molecular characteristics of pathogens such as bacterial flagellin, lipopolysaccharide and fungal chitin. After PRRs on the plasma membrane of plant cells recognize PAMPs, defense responses are initiated, including reactive oxygen species burst, MAPK pathway activation, callose deposition and stomatal closure. These responses not only protect plants from threats, but also regulate growth, development and stress balance, and intercellular signaling is crucial in this process. Small secreted peptides (SSPs) as signaling molecules are key to the plant immune regulation network.

[0003] Small secreted peptides (SSPs) are produced by specific processing of small precursor proteins and are involved in plant development, stress response and immune response regulation. They are composed of a signal peptide, a variable region and a conserved domain. They enter the secretory pathway with the help of the signal peptide, are processed by proteolytic cleavage and post-translational modification to form mature peptides, are released into the extracellular space, are detected by adjacent cell PRRs, and play a regulatory role similar to PAMPs. However, due to the preference of gene annotation and protein analysis tools for large proteins, small peptides are often underestimated. There are more than 1000 genes with secreted peptide characteristics in the genome of Arabidopsis thaliana, and the functions of most of them are unknown. There are fewer SSP cases in other species, indicating that plant small secreted peptide gene diversity needs to be explored, and its role in immunity needs to be further studied.

[0004] Tomato is a widely cultivated economic crop around the world, and is vulnerable to pathogens such as bacteria, fungi, viruses and nematodes, leading to reduced yield and fruit quality. With in-depth research, tomato small secreted peptides have attracted attention. For example, proteomics identified the CAPE of the PR-1 family of tomato, which was induced by injury and jasmonic acid methyl ester, and had significant disease resistance and slight deterrent effect on herbivorous animals; comparative genomics found the SlSolP peptide family that can induce systemic resistance in tomato. However, many potential small secreted peptides in tomato have not been explored, and as a representative of Solanaceae and with complete genome sequencing, it provides resources for discovering new small secreted peptides, and its role needs to be studied.

[0005] Developing new small secreted peptides is of great significance to improve the disease resistance of tomatoes. It can provide new molecular insights into plant immune mechanisms and is expected to become a key tool for molecular breeding. Identifying and verifying more small secreted peptides can help to understand plant immune regulation in depth and lay a theoretical and technical foundation for cultivating new varieties of disease-resistant tomatoes. These small secreted peptides can also be used as potential resources for biological control to reduce the use of chemical pesticides and promote sustainable agricultural development. Therefore, it is of great prospect and value to further explore the role of small secreted peptides in tomato plant immunity. SUMMARY

[0006] In view of the problems of the prior art, the purpose of the present application is to provide a tomato endogenous immune peptide and its use in plant immune stimulation.

[0007] The present application provides a tomato endogenous immune peptide, which is a polypeptide having at least 90% sequence similarity with the amino acid sequence set forth in SEQ ID NO. 1.

[0008] Further, the amino acid sequence of the polypeptide is as set forth in SEQ ID NO. 1.

[0009] The present application also provides a small secreted peptide, which is a peptide capable of activating the receptor of the above-mentioned tomato endogenous immune peptide.

[0010] The present application also provides the use of the above-mentioned tomato endogenous immune peptide or small secreted peptide as a plant immune stimulant.

[0011] Further, the plant immune stimulant is used to trigger the basic immune response of the plant.

[0012] The present application also provides a plant immune stimulant, which is prepared by adding the above-mentioned tomato endogenous immune peptide or small secreted peptide as an active ingredient into a pesticide-acceptable adjuvant.

[0013] The present application also provides a method for constructing disease-resistant or insect-resistant crops, comprising inserting a gene for expressing the above-mentioned tomato endogenous immune peptide or small secreted peptide into the genome of the crop.

[0014] In the present application, the "sequence similarity" is an index to measure the degree of similarity in structure and evolution of two biological sequences (such as protein or nucleic acid sequence). It is not only based on the number of completely identical amino acid residues or nucleotides in two sequences, but also considers the possibility of partial matching, that is, different but biologically conservative substitution relationship of residues can also contribute to a certain similarity score. For protein (or polypeptide) sequence, scoring matrix (such as PAM or BLOSUM matrix) is usually used to calculate similarity, which assigns a score to each pair of possible amino acid substitution, reflecting their substitution probability in the evolution process and the expected effect on function. By aligning two sequences and accumulating scores, a global or local similarity value can be obtained. A large number of previous studies have shown that small peptides with homologous sequences and activating the same receptor have the same or similar biological functions. For example, AtPIP1 (RLASGPSPRGRGH) and AtPIP2 (VKHSGPSPSGPGH) of PIP family in Arabidopsis thaliana both activate plant immune function through receptor RLK7 (Hou et al., 2014, PLoS Pathogens, 183, 399-413.). In addition, Arabidopsis CLE40 (RQVPTGSDPLHH) and CLV3 (RTVPSGPDPLHH) as homologous sequences have similar functions, which promote stem cell differentiation by binding to receptor kinase ACR4 ARABIDOPSIS CRINKLY 4) / CLV1 (Stahl et al., 2013, Current Biology, 23, 362-371; Stahl et al., 2009, Current Biology, 19, 909-914); In addition, rice FON4 (FLORAL ORGAN NUMBER 4) is a homologous gene of Arabidopsis CLV3, which can regulate rice meristem development and affect its yield traits (Ren et al., 2019, Plant Biotechnology Journal, 17, 1007-1009). Similarly, CLV3 homologous sequences in corn, tomato and other plants also play an important role in yield trait regulation (Liu et al., 2021, Nature Plants, 7, 287-294; Wang et al., 2021, Nature Plants, 7, 419-427). Therefore, sequences of the same family or activating the same receptor across species should be considered as equivalent sequences and should be within the protection scope of the present patent.

[0015] The present application has the following beneficial effects:

[0016] (1) The present application identified a new family of small secreted peptides in the tomato genome using bioinformatics tools, which was named as tomato endogenous immune peptides (TIPs).

[0017] (2) The present application found that a specific TIP12 can induce PAMP-triggered PTI response, thereby enhancing the resistance of tomato to gray mold.

[0018] (3) And, the present application further verified the intrinsic immune function of TIP12 in tomato using virus-induced gene silencing technology, and found that TIP12 is involved in the regulation of multiple immune-related pathways.

[0019] (4) In addition, the present application found that TIP12 showed a wide range of functional effects in the plant kingdom, not only active in Solanaceae (such as pepper and tobacco), but also active in other plant families (including Cucurbitaceae) and Brassicaceae (Brassicaceae).

[0020] (5) TIP12 showed a broad spectrum of effects in enhancing immunity in multiple species. This finding not only provides a new perspective for understanding plant immune mechanisms, but also lays a foundation and potential target for the development of innovative strategies for plant disease management.

[0021] Obviously, according to the above content of the present application, according to the ordinary technical knowledge and common practice in the art, other various forms of modification, replacement or change can be made without departing from the above technical idea of the present application.

[0022] The above content of the present application will be further described in detail through the specific embodiments below. However, this should not be understood as limiting the scope of the above subject matter of the present application to the following embodiments. Any technology realized based on the above content of the present application belongs to the scope of the present application. BRIEF DESCRIPTION OF DRAWINGS

[0023] Figure 1 TIP gene family analysis: (A) Characteristics of TIP family members; (B) Conservation structure analysis of TIP family members; (C) Distribution of TIP family members in phylogenetic tree and (D) chromosome; (E) Gene structure of TIP family members; (F) Distribution of cis-acting elements in TlP genes; (G) Motif distribution among TIP members.

[0024] Figure 2To investigate the immune response induced by TIP12 in tomatoes: (A) TIP12-induced ROS generation; (B) Western blot detection of TIP12-activated MAPK signaling in tomato leaves; (C) Visualization and (D) quantification of TIP12-triggered callose deposition in tomato leaves; (E) Photographing symptoms of gray mold infection in tomato leaves treated with TIP12; (F) Statistical analysis of lesion area in tomato leaves (n≥3), *** indicates P<0.001 as statistically significant.

[0025] Figure 3 To investigate the effects of silencing ProTIP12 on tomato gray mold infection: (A) gene expression levels in plants under different treatments; (B) leaf phenotypes; (C) statistics of gray mold infection; (D) leaves after trypan blue staining; (E) counted lesion area (n≥3), *** indicates that P<0.001 is statistically significant.

[0026] Figure 4 Transcriptome remodeling after TIP12 treatment: (A) Transcriptome sequencing analysis; (B) Identification and (C) analysis of upregulated and downregulated genes; (D) GO classification and (E) KEGG pathway analysis of differentially expressed genes; (F) Validation of defense-related genes. *** indicates that P < 0.001 is statistically significant.

[0027] Figure 5 TIP12 activation of immune responses in chili peppers and tobacco: TIP12 induces ROS bursts in (A) chili peppers and (B) tobacco; TIP12 activates the MAPK signaling pathway in (C) chili peppers and (D) tobacco; TIP12 induces corpus callosum deposition in (E) chili peppers and (F) tobacco; Quantitative analysis of corpus callosum deposition in chili peppers (G) and tobacco (H); Photographs of gray mold spots on (I) chili peppers and (J) tobacco leaves; Statistical analysis of lesion area in (K) chili peppers and (L) tobacco (n≥3), *** indicates P<0.001 is statistically significant.

[0028] Figure 6 TIP12 activation of immune responses in cucumber and rapeseed: TIP12 induces an increase in ROS in (A) cucumber and (B) rapeseed; TIP12 activates the MAPK signaling pathway in (C) cucumber and (D) rapeseed; TIP12 induces corpus callosum deposition in (E) cucumber and (F) rapeseed; Quantitative analysis of corpus callosum deposition in (G) cucumber and (H) rapeseed; Photographs of gray mold spots on leaves of (I) cucumber and (J) rapeseed; Statistical analysis of leaf damage area (n≥3) in (K) cucumber and (L) rapeseed. *** indicates P<0.001 as statistically significant. Detailed Implementation

[0029] In the present application examples and experimental examples, the raw materials and equipment not specifically described are known products, which are obtained by purchasing commercially available products.

[0030] Example 1, Discovery of tomato endogenous immune peptide TIP12 and its use

[0031] I. Materials and methods

[0032] 1. Plant growth conditions

[0033] Tomato, tobacco, pepper, cucumber and rape were grown in a mixture of nutrient soil, vermiculite and perlite (3:1:1). The environmental temperature was 25°C, and the light intensity was 400 μM·m -2 ·s -1 . Each tree species was subjected to a photoperiod of 16 h light and 8 h darkness, maintaining a constant relative humidity of 80%.

[0034] 2. Synthetic peptides

[0035] All peptides were 95% pure and were synthesized by GenScript Company (Nanjing, China), including TIP12 (SEQ ID NO. 1: NGDAIVEYNVVVLHY) and the positive control flg22 (SEQ ID NO. 2: QRLSTGSRINSAKDDAAGLQIA).

[0036] 3. Reactive oxygen species production

[0037] The lumino-based analysis method was used to quantify the reactive oxygen species in the plant leaves. The third or fourth pair of true leaves of four-week-old soil-grown plants were cut into leaf discs with a diameter of 3 mm. These leaves were then immersed in 30 mL of water and incubated in a 96-well plate for 12 hours. Then, a solution containing 200 μM of luminol and 2 μg-mL -1 peroxidase, and 1 μL of peptide (final concentration of 100 nM-10 μM) was added to each well. Measurements were taken every minute for a total of 60 minutes per well.

[0038] 4. Callose deposition

[0039] The detection method of callose deposition was slightly modified according to the literature (Clay et al., 2009, Science, 323, 95-101). Four-week-old tomato leaves were injected with the polypeptides (H2O as control) and treated overnight (about 12 hours). The leaves were fixed in FAA solution (10% formaldehyde, 5% acetic acid, 50% ethanol) for 24 h, transferred to absolute ethanol for decolorization for 6 h, and then incubated in 50% ethanol for 30 min. The ethanol was removed, 67 mM K2HPO4(pH 12) was added, and incubated for 30 min. The K2HPO4solution was removed, and staining solution (0.01% aniline blue in 67 mM K2HPO4, pH 12) was added and incubated for 1 h. Callose deposition was observed by ultraviolet light microscopy and photographed by an optical microscope Olympus BX-53 (Olympus, Tokyo, Japan). Callose deposition was quantified.

[0040] 5. Protein extraction and MAPKs phosphorylation detection

[0041] The leaves of 4-week-old soil-grown plants were soaked in sterile water overnight, then induced with 1 mM polypeptides for 15 min, and then the seedlings were quickly frozen in liquid nitrogen and ground into fine powder, and total proteins were extracted therefrom using a protein extraction solution (Tris-HCl (pH 6.8), 25% (V / V) glycerol, 2% (V / V) SDS, 0.001% (W / V) bromophenol blue, 5% (V / V) mercaptoethanol). The active MPK6 and MPK3 were detected by immunoblotting using anti-phospho p44 / p42 MAPK antibody (Huabio, China). The amount of total protein loaded in the electrophoresis process was compared to a consistent level by using ponceau red staining.

[0042] 6. Pathogen infection

[0043] Botrytis cinerea was cultured in potato dextrose agar medium (PDA) and incubated vertically in the dark at 28°C for 30 days. The cultured fungus plate was added with 5% sterilized glucose solution, and the mycelium on the surface of the plate was scraped off gently and poured into a 50 ml centrifuge tube, shaken thoroughly, filtered with a water-absorbing cotton, and the obtained filtrate was a spore suspension. Tomato leaves were treated with 30 mL of 1 mM peptide solution and left overnight. The leaves of similar size were cut and placed in a culture dish with sterile water soaked filter paper, and inoculated with 106mL of Botrytis cinerea spores. After being placed in an artificial climate box for 5 days, photographs were taken for observation. The degree of disease of the infected plant samples was determined and quantified. -1

[0044] 7. Transcriptome analysis

[0045] ​Tomato plants at 4 weeks old were immediately frozen in liquid nitrogen and stored at -80 °C for RNA extraction after 1 mM peptide treatment for 15 min. Total RNA was extracted from frozen leaves of 4-week-old tomato using the RNAprep Pure Plant Plus Kit (TIANGEN, Beijing, China) according to the manufacturer’s instructions. RNA-seq and bioinformatics analysis were performed by Majorbio Cp. Ltd (Shanghai, China). Sample sequencing libraries were constructed using the TruSeq RNA Sample Preparation Kit (Illumina, San Diego, CA, USA) and sequenced on the Illumina Hiseq X platform. The reference genome database was extracted from the TAIR database (https: / / www.arabidopsis.org / index.jsp) and gene annotations were obtained from the genome. Bowtie2 (2.2.6) was used to build the reference genome index, and HISAT (2.0.5) was used to map clean reads to the reference genome. HTSeq (0.9.1) was applied to gene expression and quantification levels to calculate the number of fragments per kilobase per million fragments (FPKM). DEG was identified by DESeq (1.20.0) with the screening conditions: expression difference fold |log2FoldChange| > 1 and p-value significant < 0.05. Subsequently, all differential genes were subjected to cluster analysis using the “Pheatmap” library in the R statistical package (1.0.8). Gene ontology (GO) annotation was analyzed using Blast2GO, and KEGG annotation was analyzed using KAAS. Each point had 3 biological replicates for RNA-seq analysis.

[0046] 8. Gene expression analysis

[0047] RNA-seq samples were used for quantitative RT-PCR validation. Total RNA was extracted from plant tissues by FastPure Plant Total RNA Isolation Kit (Vazyme, Nanjing, China) according to the manufacturer’s method. A total of 1 pg total RNA preparation was subjected to reverse transcription using HiScript III 1st Strand cDNA Synthesis Kit (Vazyme, Nanjing, China). The resulting cDNA was amplified using Taq Pro Universal SYBR qPCR Master Mix (Vazyme, Nanjing, China) and gene-specific primers (Table 1). SlActin was used as a reference sequence.

[0048] Table 1 RT-qPCR primers for TIP12-induced gene expression

[0049]

[0050] 9. Bioinformatics analysis

[0051] Homology search was performed using NCBI BLAST+v2.13 and iterative comparison using HMMER 3.0. Signal peptide prediction was performed by Signal 6.0. Transmembrane domains were predicted using DeepTMHMM. Phylogenetic trees were reconstructed by Muscle alignment and evolutionary history was inferred using the neighbor-joining (NJ) method. A bootstrap consensus tree was constructed using 1000 replicates. The inferred phylogenetic trees were visualized using iTOL 541. Phylogenetic analysis of PIP sequences was performed using MEGA 642. Motif prediction was performed using MEME43. Cis-acting elements were predicted using PlantCARE44. Amino acid conservation analysis was performed by Weblogo.

[0052] 10. Transcriptomic data ID

[0053] Raw reads of transcriptomic data are available in the NCBI BioProject database (https: / / www.ncbi.nlm.nih.gov / sra / PRJNA1232014), BioProject ID: PRJNA1232014.

[0054] 11. Data analysis

[0055] Statistical analysis of the data obtained was performed using IBM SPSS Statistics version 26, using the Analysis of Variance (ANOVA) procedure to test the significant effect of all the study variables. Duncan’s Multiple Range Test (DMRT) was used for mean separation, with p<0.05 as the significance test. Graphs were prepared using TBtools, Excel 2019 and Origin 2019b.

[0056] II. Experimental results

[0057] 1. Identification of SSPs in tomato

[0058] A set of peptides highly conserved in their C-terminal end, named Tomato Intrinsic Peptides (TIPs), was identified in the tomato plant genome (Table 2). These peptides are characterized by being composed of a signal peptide, a variable region and a highly conserved C-terminal region ( Figure 1 A). There is a significant conservation of the HY motif ( Figure 1 B) in the C-terminal region. Clustering and phylogenetic analysis showed that these peptide members are widely distributed in the phylogenetic tree ( Figure 1 C), mainly in chromosomes 1, 2, 6, 7 and 10 ( Figure 1D). Detailed examination of the gene structure of these members revealed that they all lack introns except TIP10, and almost all of them lack UTRs except TIP2 and TIP10 Figure 1 E). Further investigation of cis-acting elements revealed that these peptide members are rich in various cis-elements associated with light response, cold response, defense and stress, and hormone response elements. Notably, TIP2, TIP5, TIP9, TIP10 and TIP12 exhibit abundant elements associated with defense and stress response Figure 1 F). In addition, through MEME analysis, the study identified 10 core conserved motifs in the protein sequence Figure 1 G).

[0059] Table 2 Information about TIP small peptide genes

[0060]

[0061]

[0062] 2. Verification of the immune function of TIP12

[0063] The present application synthesized TIP12 and found that exogenous application of TIP12 can induce ROS burst and activate the MAPK signaling pathway in tomato plants Figure 2 A and 2B). Treatment with TIP12 also led to enhanced callose deposition Figure 2 C and 2D). In the test of infection with Botrytis cinerea, plants treated with TIP12 showed stronger resistance to pathogen invasion Figure 2 E and 2F).

[0064] The above experimental results show that TIP12 is a potential immune-regulating peptide that can enhance the immune response of plants and enhance the defense mechanism against diseases.

[0065] 3. Immune response of VIGS-mediated TIP12 precursor gene knockout tomato plants

[0066] To verify the immune effect of endogenous TIP12, the present application used VIGS technology to specifically silence the ProTIP12 gene, and qRT-PCR analysis showed that the expression of the ProTIP12 gene was significantly reduced Figure 3 A). The resistance of plants with silenced ProTIP12 gene to infection with Botrytis cinerea was significantly reduced Figure 3 B and 3C). Further cell death detection of infected plants by Evans blue staining showed that the cell death rate of plants with silenced ProTIP12 gene increased Figure 3D and 3E), indicating that the loss of ProTIP12 gene impaired the defense ability of the plants, thus increasing the cell mortality during infection.

[0067] The above experimental results show that the ProTIP12 gene plays a key role in the plant's resistance to pathogen invasion, and the reduction of its expression directly affects the immune response effect of the plant, confirming the important role of ProTIP12 in the defense library of tomato.

[0068] 4. TIP12-mediated transcriptome remodeling

[0069] To reveal the molecular mechanism of TIP12 enhancing tomato resistance to pathogen infection, the present application performs transcriptome sequencing analysis, cluster analysis, RT-qPCR verification, GO classification and KEGG pathway analysis on tomato plants treated with exogenous TIP12. The results show that TIP12, as a peptide signal, can enhance the resistance of tomato to pathogen infection by regulating various molecular pathways such as photosynthesis, hormone signal and immune response. Figure 4 A-F) show that TIP12, as a peptide signal, can enhance the resistance of tomato to pathogen infection by regulating various molecular pathways such as photosynthesis, hormone signal and immune response.

[0070] 5. Broad-spectrum resistance of TIP12 to other crops

[0071] The present application further studies the immune response of TIP12 to other Solanaceae plants. The results show that treatment with exogenous TIP12 leads to significant ROS burst and MAPK activation in pepper and tobacco ( Figure 5 A-D), indicating that TIP12 can effectively trigger their immune systems. In addition, TIP12 treatment also leads to significant accumulation of callose deposition in these plants ( Figure 5 E-H), indicating the potential role of TIP12 in promoting cell wall thickening and enhancing physical barriers of plants. Pathogen challenge tests show that TIP12 treatment can enhance the resistance of pepper and tobacco to Botrytis cinerea ( Figure 5 I-L), indicating that TIP12 has broad-spectrum disease resistance efficacy in other Solanaceae plants. It is shown that TIP12 has been proven to have broad disease resistance potential in Solanaceae plants, strengthening its possibility as a new type of plant immune activator for disease management. The present application further explores the immune function of TIP12 in other plants. In cucumber and rape, TIP12 treatment leads to ROS burst and MAPK activation ( Figure 6 A-D). TIP12 also promotes the accumulation of callose and the closure of stomata in tomato ( Figure 6 E-H), indicating that TIP12 can activate the immunity of cucumber and rape. In addition, the resistance of cucumber and rape to Botrytis cinerea is significantly enhanced after the application of TIP12. Figure 6I-L). Therefore, TIP12 has a broad-spectrum disease resistance function in Solanaceae and other plant species, and provides an important resource for developing biological pesticides with broad-spectrum effects on multiple plants.

[0072] The above experimental results show that TIP12 is a widely effective plant immune elicitor.

[0073] As can be seen from the above examples, the present application provides a tomato endogenous immune peptide and its use in plant immune elicitation. The present application first discovers a tomato endogenous immune peptide TIP12 in tomato, which can activate the immune response of plants, thereby triggering the systemic resistance of plants to pathogens. Moreover, it is found that TIP12 is involved in the regulation of multiple immune-related pathways and shows a wide functional effect in the plant kingdom, and shows a broad-spectrum effect in enhancing the immunity of multiple species. Therefore, the tomato endogenous immune peptide TIP12 provided by the present application can be used as a widely effective plant immune elicitor, and has good safety and application prospect.

Claims

1. A tomato endogenous immune peptide, characterized in that, It is a polypeptide with at least 90% sequence similarity to the amino acid sequence described in SEQ ID NO.

1.

2. The tomato endogenous immune peptide according to claim 1, characterized in that, The amino acid sequence of the polypeptide is as described in SEQ ID NO.

1.

3. A small secretory peptide, characterized in that, It is a peptide that can activate the receptor for the tomato endogenous immune peptide described in claim 1 or 2.

4. Use of the tomato endogenous immune peptide of claim 1 or 2 or the small secretory peptide of claim 3 as a plant immune stimulant.

5. The use according to claim 4, characterized in that, The plant immune stimulant is used to trigger the plant's basic immune response.

6. A plant immune stimulant, characterized in that, It is made by adding pesticide-acceptable excipients to the tomato endogenous immune peptide as described in claim 1 or 2 or the small secretory peptide as described in claim 3 as the active ingredient.

7. A method for constructing disease-resistant or insect-resistant crops, characterized in that, This includes inserting a gene for expressing the tomato endogenous immune peptide of claim 1 or 2 or the small secretory peptide of claim 3 into the crop genome.

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