Application of small peptide in preventing and treating plant diseases and promoting plant growth

Abstract

The invention provides application of small peptides in preventing and treating plant diseases and promoting plant growth. The structural formula of the small peptide is shown in the specification. Experimental results show that the small peptide shown in the formula A has the effects of improving the disease resistance of plants and regulating the growth of the plants. The small peptide not only can be used as a plant immune resistance inducer at low concentration to improve the immunocompetence of plants for resisting pathogen invasion, but also can promote plant growth, and has a good application prospect. R1-R2-beta-Ala-Trp-Gly-NH2 Formula A

Description

Technical Field

[0001] The present invention belongs to the field of agriculture, and particularly relates to the application of small peptides in preventing and controlling plant diseases and promoting plant growth. Background Art

[0002] In global agricultural production, plant diseases are one of the main factors leading to crop yield reduction and crop failure. In order to reduce the safety problems such as soil degradation and water pollution caused by chemical pesticides, finding a green, pollution-free and residue-free prevention and control method is an important development strategy in the agriculture of our country and even the world today (Li Linlin, Li Tianlai, etc. Regulation of calcium on the resistance of tomato seedlings to Botrytis cinerea induced by SA [J]. Acta Horticulturae Sinica, 2012, 39(2): 273-280.). Most of the basic principles of the drugs for controlling plant diseases in the past targeted pathogens and aimed to quickly and comprehensively kill the targets, ignoring the resistance of the host plants damaged by the pathogens to these foreign organisms. In 2007, German scientists published an article in "Science" pointing out that plants in nature have special immune sensors that can recognize the invasion of microorganisms such as bacteria and viruses (Shen QH, Saijo Y, et al. Nuclear Activity of MLA Immune Receptors Links Isolate-Specific and Basal Disease-Resistance Responses [J]. Science, 2007, 315(5815): 1098-103.). These substances can be called elicitors. An elicitor is a general term for a class of special compounds that can activate the host plant to produce a defense response, and can be divided into oligosaccharides, proteins, polypeptides (Wang Hegui, Sun Xiaotang, etc. Research progress of biogenic protein elicitors [J]. Guihaia, 2016, 36(04): 413-8.), etc. People utilize the induced immune resistance of plants to develop inducers or elicitors into plant immune elicitors and make full use of their functions to prevent and control plant diseases.

[0003] Insect neuropeptides are a class of small-molecule active polypeptides synthesized, secreted periodically by specific nerve cells in the insect brain and transmitted through nerves or body fluids, regulating every developmental process of insects such as growth and development, molting and metamorphosis, diapause, metabolism, and reproduction. Based on this important physiological function, they are considered potential pest control agents. The inventor found through structural optimization in the early stage that some insect neuropeptide analogs have good insecticidal effects on pests. However, there has been no report on the research of applying insect neuropeptides to the fields of plant immunity and plant growth regulation. Summary of the Invention

[0004] The purpose of the present invention is to provide a new use of a class of small peptides.

[0005] The small peptide provided by the present invention has a general structural formula as shown in Formula A:

[0006] R1-R2-β-Ala-Trp-Gly—NH2

[0007] Formula A

[0008] Among them, R1 is selected from any one of cinnamic acid and 4-nitrocinnamic acid;

[0009] R2 is an amino acid;

[0010] The carboxyl group contained in R1 and the amino group of the amino acid represented by R2 form an amide bond;

[0011] Preferably, R2 is selected from any one of glycine, L-aspartic acid, D-aspartic acid, D-tert-leucine, L-tert-leucine, D-homophenylalanine, L-homophenylalanine, D-2-aminobutyric acid, L-2-aminobutyric acid, D-4-trifluoromethylphenylalanine, L-4-trifluoromethylphenylalanine, D-valine, L-valine, D-cyclohexylalanine, L-cyclohexylalanine, L-phenylalanine, 2-amino-5-cyclohexyloxy-5-oxopentanoic acid, and O-benzyl-L-serine.

[0012] The small peptide shown in Formula A above is obtained by the preparation method in the reference (Zhang C L, Li X L, Song D L, Ling Y, Zhou Y L, Yang X L*, Synthesis, aphicidal activity and conformation of novel insect kinin analogues as potential eco-friendly insecticides. Pest Management Science, 2020, 76(10): 3432-3439.).

[0013] A new use of the small peptide provided by the present invention is the application of the small peptide shown in Formula A in preventing and controlling plant diseases.

[0014] The diseases include sheath blight of rice, gray mold of cucumber, gray mold of tomato, sclerotinia of soybean, and rust of soybean;

[0015] In the above application, the small peptide shown in Formula A serves as a plant immune inducer, plays an immune induction role, and stimulates the plant's own immunity to prevent and control plant diseases.

[0016] The present invention also provides the application of the small peptide shown in Formula A in promoting plant growth.

[0017] Specifically, the small peptide shown in formula A can promote the germination and emergence of plant seeds and the growth of plant plants.

[0018] The plants may be: rice, soybean, tomato and cucumber.

[0019] The present invention also provides a method for preventing and controlling plant diseases and / or promoting plant growth.

[0020] The method for preventing and controlling plant diseases and / or promoting the growth of plant plants provided by the present invention includes the following steps: spraying the plant plants with a preparation containing the small peptide shown in the above formula A is sufficient.

[0021] Among them, the preparation containing the small peptide shown in formula A contains the small peptide shown in formula A, a cosolvent, a surfactant and water.

[0022] The cosolvent may be one or more of methanol, propanol, butanol, pentanol, dimethyl sulfoxide (DMSO), and specifically, dimethyl sulfoxide (DMSO) may be preferably selected;

[0023] The surfactant may be styrylphenyl polyoxyethylene ether, styrylphenyl polyoxyethylene ether phosphate, alkylphenol formaldehyde resin polyoxyethylene ether, phenethylphenyl polyoxyethylene-polyoxypropylene ether, calcium dodecylbenzenesulfonate, Triton-x-100, etc., and specifically, Triton-x-100 may be preferably selected;

[0024] According to mass percentage, in the preparation, the concentration of the small peptide may be 10 -1 ~10 -6 mg / L; preferably 10 -2 ~10 -4 mg / L;

[0025] The spraying is foliar spraying, spraying once every 3 days, and spraying a total of 3 times.

[0026] The present invention also provides a method for promoting the germination and emergence of plant seeds.

[0027] The method for promoting the germination and emergence of plant seeds provided by the present invention includes the following steps: soaking the plant seeds with the small peptide shown in the above formula A is sufficient.

[0028] Experimental results show that the small peptide shown in formula A has both the function of enhancing the disease resistance of plants and regulating plant growth. Such small peptides not only act as plant immune elicitors at low concentrations, improving the immune ability of plants to resist pathogen invasion, but also can promote plant growth, and have good application prospects.

[0029] The beneficial effects of the present invention are as follows: The present invention provides a class of small peptides with the function of inducing plant immune disease resistance. The small peptides enhance the levels of defense enzymes and defense hormones in plants, activate the expression of disease resistance-related genes, and thus improve the plant's own immunity and enhance the ability of various plants to resist pathogenic bacteria. At the same time, it is also found that these small peptide mimics have the function of plant growth regulators.

[0030] The mimics modified from the terminal core pentapeptide of neuropeptide mimics of the present invention have the function of immune elicitors. Without the activity of in vitro bactericidal, they can improve the plant's own immunity, enhance the disease resistance ability, and have a good application prospect in the growth regulation of plants. Detailed implementation manners

[0031] The present invention will be further described in detail below in conjunction with the specific implementation manners. The provided embodiments are only for clarifying the present invention, rather than limiting the scope of the present invention. The following provided embodiments can be used as a guide for those of ordinary skill in the art to make further improvements, and do not limit the present invention in any way.

[0032] The experimental methods in the following embodiments are all conventional methods unless otherwise specified, and are carried out according to the techniques or conditions described in the literature in this field or according to the product instructions. The materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

[0033] The small peptides shown in Formula A are obtained by the preparation methods of References 1) Pest Management Science, 2020, 76(10): 3432-3439; 2) Pest Management Science, 2022, 78(7): 2952-2963.

[0034] The structural formulas of the compounds A1-A10 in the following embodiments are shown in Table 1:

[0035] Table 1 Structures of R1 and R2 corresponding to the structure of the small peptide of Formula A

[0036]

[0037] Example 1, In vitro bactericidal activity of small peptides

[0038] The structures of the small peptides selected in this example and the following examples are shown in Table 1.

[0039] Weigh an appropriate amount of small peptide analogs and dissolve them in DMSO, dilute them with 0.05% Triton solution to a mother solution with a concentration of 100 mg / L, and dilute them into 50 mL of liquid PDA medium with a concentration of 50 mg / L. Pour 50 mL of liquid culture medium evenly into 3 culture dishes with a diameter of 9 cm in the clean bench, use a sterilized puncher to punch out activated fungal cakes with a diameter of 5 mm, and use an inoculation needle to transfer the cakes to the center of the PDA culture medium. Seal the inoculated culture dish with a sealing film and culture it in a dark incubator at 25°C. The positive control of Rhizoctonia solani uses the same concentration of tebuconazole, the positive control of Sclerotinia sclerotiorum uses the same concentration of sclerotium net, the positive control of Magnaporthe oryzae and Botrytis cinerea uses the same concentration of boscalid, and the drug-free treatment is set as the control. Observe the growth of mycelium. When the CK group grows to almost fill the culture dish (about 70-80% of the culture dish), the cross method is used to calculate the colony diameter and calculate the antibacterial rate. The calculation formula is as follows:

[0040]

[0041] Table 2 In vitro antibacterial activity of small peptides against pathogenic fungi

[0042]

[0043] The results of in vitro antibacterial activity determination in Table 2 show that the inhibition rates of the small peptides against Rhizoctonia solani, Sclerotinia sclerotiorum and Botrytis cinerea at a concentration of 50 mg / L were all lower than 10%, while the positive control fungicides had significant antibacterial effects against different pathogens, indicating that compared with the fungicides, the small peptides had no obvious bactericidal effect on the above plant pathogenic fungi and did not have an in vitro antibacterial effect.

[0044] Example 2: Anti-rice disease activity of small peptides

[0045] The pot test method was used: rice seeds with full grains and uniform size were selected, the surface was disinfected with 75% alcohol, rinsed with distilled water and dried, and germinated at 25°C to about 5mm. In a 28°C greenhouse, the germinated seeds were sown in sterilized nutrient soil and vermiculite mixed in a 2:1 ratio, with 5 seeds per pot, 10 pots per group, and repeated 3 times. Appropriate amount of water was added every morning and evening, and the induction treatment was started when the growth reached about 40 days. An appropriate amount of small peptide mimetic was weighed and dissolved in DMSO to form a mother solution, and the mother solution was diluted to 10 with 0.05% Triton solution. -4 mg / L, spray evenly on rice plants, spray 200μL per plant, select clean water as the control, induce once every 3 days, for a total of 3 times, and inoculate the disease 24h after the last induction.

[0046] Inoculation method for rice sheath blight (Rhizoctonia solani): Through the mycelial disc inoculation method, the rice sheath blight pathogen is cultured in a Petri dish with potato agar medium. When the mycelium covers the Petri dish, holes are punched at the edge of the colony to make mycelial discs with a diameter of 5 mm for inoculation. During inoculation, the first leaf sheath of the rice is separated, and a sterile toothpick is used to insert the mycelial disc into the separated leaf sheath. After inoculation, the leaf sheath is restored. One mycelial disc is inoculated per plant. The relative humidity is maintained at 80%, and after continuing to culture in the greenhouse for one week, the disease incidence of each group of rice is recorded. The positive control uses Flg22 (amino acid sequence: QRLSTGSRINSAKDDAAGLQIA), which is a conserved 22-amino acid sequence isolated from Pseudomonas aeruginosa and located at the N-terminus of bacterial flagellin. When applied externally to plants, it can induce disease resistance in plants.

[0047] The disease grading standard for rice sheath blight refers to the "Standard Operating Procedures for Pesticide Bioactivity Testing - Fungicide Volume":

[0048] Grade 0: The first leaf sheath shows no disease.

[0049] Grade 1: The lesion area on the first leaf sheath accounts for less than 5% of the total leaf sheath area.

[0050] Grade 3: The lesion area on the first leaf sheath accounts for 6% - 10% of the total leaf sheath area.

[0051] Grade 5: The lesion area on the first leaf sheath accounts for 11% - 25% of the total leaf sheath area.

[0052] Grade 7: The lesion area on the first leaf sheath accounts for 26% - 50% of the total leaf sheath area.

[0053] Grade 9: The lesion area on the first leaf sheath accounts for more than 50% of the total leaf sheath area.

[0054] The control effect is calculated based on the disease index, and the calculation formula is as follows:

[0055] Disease index (%) = Σ (number of diseased leaves at each level × relative level value) / (total number of leaves surveyed × highest disease level) × 100 Control effect (%) = (control disease index - treatment disease index) / control disease index × 100

[0056] Table 3 Induced resistance control effect of rice sheath blight after small peptide treatment

[0057]

[0058]

[0059] Table 3 shows that after 10 -4After treatment with the small peptide compound at mg / L, the incidence of sheath blight of rice can be reduced, as manifested by a decrease in the disease index.

[0060] Example 3. Antifungal activity of the small peptide of the present invention against cucumber diseases

[0061] Using the pot experiment method: Select cucumber seedlings at the two-leaf stage with consistent growth and similar health status, group them with 1 plant per pot and 10 pots per group, and repeat 3 times. Configure the small peptide into a solution with a concentration of 10 -4 mg / L according to the method of Example 1, spray it evenly on the rice plants, 200 μL per plant, select clear water as the control, induce once every 3 days, for a total of 3 inductions, and inoculate with the disease 24 h after the last induction.

[0062] Inoculation method for cucumber gray mold (Botrytis cinerea): Through the disk inoculation method, culture Botrytis cinerea of cucumber on a potato agar medium in a petri dish. When the mycelium covers the petri dish, punch holes at the edge of the colony to make disks with a diameter of 5 mm for inoculation. When inoculating, use a sterile toothpick to stick the fungal cake to the back of the cucumber leaf avoiding the veins, 1 fungal cake per leaf, keep the relative humidity at 80%, continue to culture for 4 days, and then record the disease incidence of each treatment according to the disease grading standard of cucumber gray mold, and calculate the disease index and disease prevention effect.

[0063] The disease grading standard for cucumber gray mold refers to:

[0064] Grade 0: No symptoms;

[0065] Grade 1: A small number of lesions appear on the inoculated leaf;

[0066] Grade 2: The lesion area accounts for 1 / 3 of the leaf area;

[0067] Grade 3: The lesion area accounts for 1 / 3 - 1 / 2 of the leaf area;

[0068] Grade 4: The lesion area accounts for 1 / 2 - 2 / 3 of the leaf area;

[0069] Grade 5: The lesion area accounts for more than 2 / 3 of the leaf area.

[0070] Disease index (%) = (number of diseased leaves at each level × disease grade) / (total number of leaves surveyed × highest disease grade) × 100.

[0071] Control effect (%) = (control disease index - treatment disease index) / control disease index × 100.

[0072] Table 4. Induced resistance and control effect of the small peptide against cucumber gray mold

[0073]

[0074]

[0075] Table 4 shows that after treatment with the small peptide, the incidence of Botrytis cinerea on cucumbers is reduced, thus confirming that the small peptide has a certain preventive effect on Botrytis cinerea of cucumbers.

[0076] Example 4. Anti-soybean disease activity of the small peptide of the present invention

[0077] Using the pot experiment method: Select soybean seeds of the same size and plump grains, soak them in clean water for 6 h for germination, and then sow them into seedling pots, with 5 seeds in each pot, 3 pots in each group, and 3 replicates. Select healthy soybean plants with the same growth trend and size. According to the method of Example 1, configure the small peptide into a solution with a concentration of 10 -4 mg / L, and evenly spray it onto the soybean plants, spraying 200 μL per plant. Select clean water as the control, induce once every 3 days, for a total of 3 inductions, and perform disease inoculation 24 h after the last induction.

[0078] Inoculation method for Soybean sclerotium: By the method of inoculating with a fungus disk, cultivate the soybean sclerotium pathogen in a petri dish with a potato agar medium. When the mycelium covers the petri dish, punch holes at the edge of the colony to make fungus disks with a diameter of 5 mm for inoculation. When inoculating, stick the fungus cake to the back of the soybean leaf avoiding the leaf veins, stick 1 fungus cake on each leaf, keep the relative humidity at 80%, continue to culture for 2 - 3 days, and then record the disease incidence of each treatment according to the disease grading standard of Soybean sclerotium, and calculate the disease index and disease prevention effect.

[0079] Inoculation method for Phakopsora pachyrhizi: Inoculate the prepared spore suspension of 5×10 5 per mL onto the soybean leaves, let it dry naturally, and then place it in a humidity chamber at a temperature of 25°C and a humidity of 100% for dark humidity cultivation for 24 h. After 24 h, place it in a culture room at about 25°C for normal cultivation, with a light-dark ratio of 12 hL / 12 hD and an air relative humidity of 60% - 80%. Continue to culture for 10 days, and then record the disease incidence of each treatment according to the disease grading standard of Phakopsora pachyrhizi, and calculate the disease index and disease prevention effect.

[0080] Reference for the disease grading standard of Soybean sclerotium:

[0081] Grade 0: No symptoms;

[0082] Grade 1: The diseased area accounts for less than 5% of the total leaf area;

[0083] Grade 2: The diseased area accounts for 5% - 10% of the total leaf area;

[0084] Grade 3: The diseased area accounts for 11% - 30% of the total leaf area;

[0085] Level 4: The diseased area accounts for 31% - 50% of the total leaf area;

[0086] Level 5: The diseased area accounts for more than 50% of the total leaf area.

[0087] The grading standard for soybean rust disease refers to GB / T 17980.89 - 2004:

[0088] Level 0: Disease - free;

[0089] Level 1: The diseased area accounts for less than 5% of the whole leaf area;

[0090] Level 3: The diseased area accounts for 6% - 25% of the whole leaf area;

[0091] Level 5: The diseased area accounts for 26% - 50% of the whole leaf area;

[0092] Level 7: The diseased area accounts for 51% - 75% of the whole leaf area;

[0093] Level 9: The diseased area accounts for more than 76% of the whole leaf area.

[0094] Disease index (%) = Σ (number of diseased leaves at each level × relative level value) / (total number of leaves surveyed × highest disease level) × 100.

[0095] Control effect (%) = (control disease index - treatment disease index) / control disease index × 100.

[0096] Table 5 Disease index and induced resistance effect of soybean plants after small peptide treatment

[0097]

[0098] Table 5 shows that after treatment with 10 -4 mg / L small peptide, the incidence degrees of soybean sclerotinia and soybean rust both decreased to a certain extent, manifested as a decrease in the disease index, indicating that small peptide has good induced disease - resistance effects on both soybean sclerotinia and soybean rust.

[0099] Example 5. Growth - regulating function of the small peptide of the present invention on rice

[0100] In this example, the effects of small peptide on rice seed germination through seed - soaking treatment and the growth of rice plants grown from the sown seeds were measured. The specific operations are as follows: 11 treatments were set in the experiment, namely CK treated with clear water, and the concentrations were 10 -6 mg / L, 10 -4 mg / L, 10 -2Solutions of PMDDⅢ-Y07 and PMDDⅡ-Y01 at 0 mg / L, 1 mg / L, and 100 mg / L. Select seeds that are uniform and plump. After soaking the seeds in the above three treatment solutions for 24 h respectively, place the seeds in a petri dish with moist filter paper and germinate them at 37°C. For each treatment, use 3 petri dishes, with 100 seeds in each dish. After 7 days, measure the seed germination rate. Seed germination rate = (number of germinated seeds / total number of seeds) × 100%. Count the bud length and root length of the germinated seeds. Conduct the experiment in 3 replicates.

[0101] Transplant the seeds treated with the above 11 soaking solutions into seedling pots. The cultivation conditions are 12 h of light at 28°C and 12 h of dark cultivation at 22°C. After the rice seedlings emerge from the soil, spray water once in the morning and once in the evening every day to keep them moist. After 21 days, for each treatment, count the height, root length, stem base width, and fresh weight of 20 rice seedlings. Conduct the experiment in 3 replicates and take the average value.

[0102] Table 6 Effects of soaking rice seeds with different concentrations of small peptides on seed germination

[0103]

[0104] It shows that treatments with different concentrations of small peptides all have a promoting effect on the emergence of rice seeds, especially the best effect is achieved at a concentration of 10 -6 mg / L.

[0105] Table 7 Quality of rice seedlings treated with different concentrations of small peptides

[0106]

[0107]

[0108] Table 7 shows that treatments with different concentrations of small peptides all have a certain promoting effect on rice plants. The best effect concentrations are 10 -2 mg / L and 10 -6 mg / L respectively.

[0109] Example 6. Growth regulation function of small peptides on soybeans

[0110] Select high-quality and undamaged soybean seeds. After disinfection, soak the seeds in a solvent and the corresponding compound solution for 6 hours at 25°C in the dark. After taking them out, rinse them three times with distilled water and dry them. For each treatment, use a seedling tray, and cultivate 10 seeds in each replicate (total replicates = 3). Spread the seeds on wet filter paper and lay them along the same direction on the hypocotyl. Cover the seeds with filter paper and place them in the dark at 25°C and a relative humidity of 75% for 4 days. After 4 days, transfer them to flower pots. There are 10 seeds in each flower pot, and there are 3 biological replicates for each treatment. These flower pots are kept at 25 - 26°C and a relative humidity of 75% in an incubator for 7 days. Measure the stem length and root length 7 days after transplantation.

[0111] Table 8 Growth indexes of soybean seedlings treated with small peptides at different concentrations for 11 days

[0112]

[0113]

[0114] Table 8 shows that soaking soybean seeds with small peptides can promote the growth of soybean bud length.

[0115] Example 7. Effects of small peptides on phytohormones in soybeans

[0116] The standard operating procedure for quantitatively detecting auxin (IAA), abscisic acid (ABA), salicylic acid (SA), and jasmonic acid (JA) using ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) is as follows: First, take 50 mg of freeze-dried and ground leaf samples, add 2 mL of 10% methanol solution (v / v), and incubate in an ice bath for 45 minutes. Subsequently, use an IKA Ultra-Turrax disperser to perform 3 intermittent homogenizations (20 seconds each time) under ice bath conditions. Place the samples in a 4°C environment and centrifuge at 4000 rpm for 45 minutes using a horizontal rotor centrifuge. Take the supernatant and transfer it to a new centrifuge tube. Adjust the pH to 2.5 with acetic acid under ice bath conditions, and add 2 mL of diethyl ether for two liquid-liquid extractions. After centrifuging at 4000 rpm for 3 minutes at 4°C again, collect the organic phase into a new tube and concentrate and dry it under vacuum. Before detection, reconstitute with a methanol / water solution (10:90) containing 0.01% formic acid and add isotope internal standard solutions: abscisic acid-d6 (ABA-d6), salicylic acid-d5 (SA-d5), indoleacetic acid-d4 (IAA-d4), dehydrojasmonic acid (dhJA), and jasmonoyl-isoleucine-c6 (JA-Ile-c6), with a final concentration of 100 ng / mL each. Establish an external standard curve using standard products with a gradient concentration of 0.1 - 150 ppb for accurate quantification. Chromatographic separation is performed using a Phenomenex UPLC Kinetex 2.6μm EVO C18 chromatographic column (2.1×50 mm), and mass spectrometry detection is performed using a Waters Acquity UPLC system coupled with an Xevo TQS triple quadrupole mass spectrometer. Among them, IAA and ABA are detected using full-MS full scan in the negative ion mode combined with MS / MS secondary confirmation. Quantitative analysis is based on MS1 spectral data, and the MS2 spectrum is used for target compound structure verification.

[0117] Table 9 Percentage changes in phytohormone concentrations in soybean seedling stems and roots

[0118]

[0119] Table 9 reveals that the growth-promoting effect of small peptides on soybeans is closely related to the regulation of key phytohormones: it simultaneously increases the IAA content in both roots and stems, raises the SA level in stems, and shows a differential distribution pattern with a decrease in JA in stems and an increase in JA in roots. This hormone balance mechanism synergistically optimizes plant growth and stress resistance.

[0120] Example 8. Growth regulation function of the small peptides of the present invention on tomatoes

[0121] Select high-quality and undamaged tomato seeds. After growing for 14 days, spray small peptides at a concentration of 10 -4 mg / L, spray once every 3 days, for a total of 3 sprays. Measure the growth situation 7 days after the last spray. The results are shown in Table 7.

[0122] Table 10 Effects of small peptide treatment on tomato growth

[0123]

[0124]

[0125] The results in Table 10 show that after the tomato plants were treated with the small peptide solution by spraying, most small peptides had a certain promoting effect on the growth and development of tomatoes.

[0126] Example 9. Changes in defense enzyme activities after treating rice with small peptides

[0127] After the rice plants were induced by spraying with a small peptide solution at a concentration of 10 -4 mg / L once, samples were taken 48 h later and quickly placed in liquid nitrogen and stored frozen at -80 °C for later measurement. A treatment without the drug was set as the control, and each treatment was repeated 3 times.

[0128] When measuring the defense enzyme activities, take leaf tissues, wash the surface dirt with distilled water, dry with absorbent paper, take an appropriate amount into a mortar, cut into pieces, add an appropriate amount of liquid nitrogen, quickly grind into powder, weigh the powder, and add 9 times the volume of phosphate buffer (phosphate buffer: 0.1 mol / L, pH 7 - 7.4) according to the ratio of weight (g): volume (mL) = 1:9. Vortex and mix for 3 minutes, centrifuge at more than 3500 revolutions per minute for 10 minutes, take the clear and bright 10% homogenate supernatant (crude extract), and then dilute it with phosphate buffer to different concentrations for the experiment. The activities of different enzymes such as SOD (superoxide dismutase), CAT (catalase), POD (peroxidase), PPO (polyphenol oxidase), and PAL (phenylalanine ammonia-lyase) were detected by the absorbance method according to the reagents and steps provided by each kit. The test results are expressed in enzyme activity units per gram of fresh weight (U / g·FW). The kits were selected from Nanjing Jiancheng.

[0129] Table 11 Effects of small peptide treatment on changes in defense enzyme activities in rice

[0130]

[0131]

[0132] Table 11 results show that small peptides can effectively induce the activation of various defense enzymes in rice plants, thus having a certain impact on enhancing the disease resistance activity of rice.

[0133] The above has detailed the present invention. For those skilled in the art, without departing from the gist and scope of the present invention and without unnecessary experiments, the present invention can be implemented within a relatively wide range under equivalent parameters, concentrations and conditions. Although specific embodiments of the present invention are given, it should be understood that the present invention can be further improved. In short, according to the principle of the present invention, this application intends to cover any changes, uses or improvements to the present invention, including changes made using conventional techniques known in the art that depart from the scope disclosed in this application.

Claims

1. Use of the small peptide represented by formula A in preventing and controlling plant diseases, R1-R2-β-Ala-Trp-Gly-NH2 Formula A In formula A, R1 is selected from any one of cinnamic acid and 4-nitrocinnamic acid; R2 is an amino acid; R2 is selected from any one of glycine, L-aspartic acid, D-aspartic acid, D-tert-leucine, L-tert-leucine, D-homophenylalanine, L-homophenylalanine, D-2-aminobutyric acid, L-2-aminobutyric acid, D-4-trifluoromethylphenylalanine, L-4-trifluoromethylphenylalanine, D-valine, L-valine, D-cyclohexylalanine, L-cyclohexylalanine, L-phenylalanine, 2-amino-5-cyclohexyloxy-5-oxopentanoic acid, and O-benzyl-L-serine; The carboxyl group contained in R1 and the amino group of the amino acid represented by R2 form an amide bond.

2. The use according to claim 1, wherein the diseases include sheath blight of rice, gray mold of tomato, gray mold of cucumber, sclerotinia of soybean, and rust of soybean.

3. The application according to claim 1, characterized in that In the said use, the small peptide represented by formula A serves as a plant immune inducer, plays an immune induction role, and stimulates the plant's own immunity to prevent and control plant diseases.

4. Use of the small peptide represented by formula A in claim 1 in promoting plant growth.

5. The application according to claim 4, characterized in that, In the said use, the small peptide represented by formula A promotes the germination and emergence of plant seeds and the growth of plant plants.

6. The application according to claim 4, characterized in that, The said plants are: rice, soybean, tomato, and cucumber.

7. A method for preventing and controlling plant diseases and / or promoting plant growth, comprising the following steps: spraying the plant plants with a preparation containing the small peptide represented by formula A in claim 1.

8. The method according to claim 7, wherein The preparation containing the small peptide represented by formula A contains the small peptide represented by formula A, a cosolvent, a surfactant, and water; The said cosolvent is one or more of methanol, propanol, butanol, pentanol, and dimethyl sulfoxide; The said surfactant is one or more of styrylphenyl polyoxyethylene ether, styrylphenyl polyoxyethylene ether phosphate, alkylphenol formaldehyde resin polyoxyethylene ether, phenethylphenyl polyoxyethylene-polyoxypropylene ether, calcium dodecylbenzenesulfonate, and Triton-x-100; In terms of mass percentage, in the preparation, the concentration of the small peptide is 10 -1 ~10 -6 mg / L; The said spraying is foliar spraying, spraying once every 3 days, for a total of 3 times.

9. A method for promoting the germination and emergence of plant seeds, comprising the following steps: soaking the plant seeds with the small peptide represented by formula A in claim 1.

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