CN111713510A

CN111713510A - Application of gallnut extract in preparation of plant-derived resistance inducer and resistance inducer

Info

Publication number: CN111713510A
Application number: CN202010513626.8A
Authority: CN
Inventors: 冯俊涛; 郭文慧; 闫合; 王勇; 蔡崇林; 马志卿
Original assignee: Northwest A&F University
Current assignee: Northwest A&F University
Priority date: 2020-06-08
Filing date: 2020-06-08
Publication date: 2020-09-29
Anticipated expiration: 2040-06-08
Also published as: CN111713510B

Abstract

The invention discloses application of a Chinese gall extract in preparation of a plant-derived resistance inducer and the resistance inducer. The gallnut extract is a plant extract obtained by extracting gallnuts with an organic solvent, the mass percentage content of the gallnut extract in the plant resistance inducer is 10-50%, and the gallnut extract is prepared into soluble solution, powder and microemulsion which are used for preventing or relieving various plant fungi, bacteria and virus diseases such as Tobacco Mosaic Virus (TMV), kiwi fruit canker (Pseudomonas syringae PV) and rape sclerotinia sclerotiorum (S serotina (Lib.) de Bary) on crops such as Tobacco, kiwi fruit, rape and the like. The resistance inducer can activate the systematicness of tobacco to obtain resistance, and has good prevention effect on various diseases.

Description

Application of gallnut extract in preparation of plant-derived resistance inducer and resistance inducer

Technical Field

The invention belongs to the field of plants and application thereof, and particularly relates to application of a gallnut extract in preparation of a plant-derived resistance inducer and the resistance inducer.

Background

Plants have developed a variety of immune mechanisms over long periods of evolution. Chemical substances capable of inducing plants to produce systemic resistance (also called Systemic Acquired Resistance (SAR)) are called plant disease resistance activators (elicers). The secondary metabolites contained in plants are important for ensuring the normal growth of plants in nature. Besides directly resisting pathogens, pests and even other plant invasion, some secondary metabolites can also participate in the activation of immune signal pathways in plants, induce the generation of plant disease resistance mechanisms and further enhance the self resistance of the plants. The natural substance of plant source is used for inducing plants to generate antibacterial or antiviral products to inhibit the infection of pathogens, so that the occurrence of diseases is reduced, and the method is an environment-friendly, safe and efficient pest control strategy.

Typical products of the existing immune elicitors which are researched more in China comprise natural gibberellin, brassinolide, amino-oligosaccharin, chitosan and the like. Wherein 54 amino-oligosaccharin registered products and 40 brassinolide registered products gradually become registration hotspots in the pesticide industry. Innovative products such as Dufulin, Vanilla acetal and sea island essence researched by Songbaoan team of academy of China are currently better immune elicitors, and have better effects and application prospects on disease resistance, stress resistance, yield increase and quality improvement of crops such as tea trees, fruit trees, rice, traditional Chinese medicines, tobacco and the like. The 6% oligosaccharide-catenin wettable powder (Altailing) is a formal registered plant immunity inducer protein biological pesticide developed and obtained by a Changqiu text researcher team, which is a subsidiary of plant protection research institute of Chinese agricultural academy of sciences, and can effectively improve plant immunity, control disease occurrence, promote plant root system growth, be safe and environment-friendly, have no residue, and have obvious effects of increasing yield and income. At present, the plant resistance inducer is widely applied to various varieties of various plants such as potatoes, tomatoes, soybeans, cucumbers, hot peppers, arabidopsis thaliana and the like to prevent and treat various diseases, and obtains good prevention effect. The resistance inducer transfers the focus from a pathogen to the plant, promotes the robust growth of the plant, improves the capability of the plant to resist the virus diseases and other various pathogens, avoids a series of hazards caused by pesticide use, and conforms to the development strategy of harmonious plant protection. Plant resistance elicitors are becoming hot spots in the research and development of pesticides.

Gallnut (Rhus chinensis Mill.) is a certain aphid female of Homoptera Migophyridae (Pemphigae Mill.) parasitizing on Rhus chinensis Mill (Rhus chinensis Mill.) of Anacardiaceae (anacadiaceae), Populus pulus (Rhus chinensis Maxim.), Rhus pulus (Rhus punjabensis var. sinica Rehd. et Wils.) and tender leaves or leaf stalks of other plants of the same genus, stabbing to generate a cystic polyploid insect gall, and drying. The Chinese Galla chinensis is mainly produced in mountainous areas and hilly areas such as Qinling, Daba mountain, Wudangshan, Wushan, Wuling mountain, Emei mountain, Daro mountain, and Daliang mountain. The elevation is 250-1600 m in vertical distribution, and 500-600 m is more concentrated, so that the resources are rich. Commercial gallnut is of horny fold type, tripe type and gall flower type. The jianbao gallnut is mainly produced in Guizhou, Sichuan, Hubei Hunan, Yunnan and Guangxi provinces. Ganbao gallnut is mainly produced in Hubei, Shaanxi and Jiangxi provinces. Galla chinensis is used as a traditional Chinese medicinal material, is more researched in the medical field, and has the antibacterial and anti-inflammatory effects. Galla chinensis extract is reported to have inhibitory activity against Staphylococcus aureus (Staphyloccocusareus), Escherichia coli (Escherichia coli), Candida albicans (Monilia albicans), and the like. In the agricultural field, the gallnut extract has a good inhibition effect on Botrytis cinerea; we find for the first time that the Chinese gall extract can effectively induce plants to resist diseases and has further development and application values in agriculture.

Disclosure of Invention

The invention aims to provide application of a Chinese gall (Rhus chinensis Mill.) extract in preparing a plant source induced disease-resistant activator (hereinafter referred to as an inducer) and the inducer.

In order to achieve the purpose, the technical scheme adopted by the invention comprises the following steps:

application of Galla chinensis extract in preparing plant resistance inducer is provided.

Optionally, the gallnut extract is a plant extract obtained by extracting gallnuts with an organic solvent.

Optionally, the gallnut extract accounts for 10-50% of the plant resistance inducer by mass.

Optionally, the preparation method of the gallnut extract comprises the following steps:

crushing Chinese gall into powder with the particle size of 1-5 mm, and adding an extraction solvent at room temperature in a ratio of 1:10(w/v), wherein the extraction solvent is any one or a mixture of more than two of methanol, ethanol, ethyl acetate, chloroform and petroleum ether; standing at room temperature for 24 hr, and extracting in an ultrasonic cleaning instrument with 40 deg.C and 100Hz frequency for 60 min; filtering, extracting the residue for 2 times, mixing the three filtrates, and concentrating the filtrate under reduced pressure to obtain plant extract.

An inductive resistance agent takes a Chinese gall extract as an effective component, and the mass percentage content of the Chinese gall extract in the inductive resistance agent is 10-50%.

The resistance inducer is a gallnut extract microemulsion which is prepared from the following raw materials in percentage by weight:

10-50% of Chinese gall extract, organic solvent: 10% -40%, surfactant: 5% -25%, antifreeze: 4% -8%, antimicrobial agent: 0.1 to 1 percent of the total weight of the raw materials, and the balance of water, wherein the total weight percentage of the raw materials is 100 percent.

The resistance inducer is gallnut extract soluble solution, and the soluble solution is prepared from the following raw materials in percentage by weight:

10-50% of Chinese gall extract, organic solvent: 10% -40%, surfactant: 5% -30%, antifreeze: 1 to 5 percent of ethanol, and the balance of ethanol, wherein the sum of the weight percentages of the raw materials is 100 percent.

The resistance inducer is gallnut extract powder, and the powder is prepared from the following raw materials in percentage by weight:

10-50% of nutgall extract, and filler: 10% -50%, organic solvent: 10-30%, dispersant: 5% -10%, stabilizer: 1 to 5 percent, and the sum of the weight percentages of the raw materials is 100 percent.

The gallnut plant source resistance inducer prepared by the invention has the following advantages:

the Chinese gall plant disease-resistant activator has remarkable activity, and can be used for preventing or relieving various fungal, bacterial and viral diseases such as Tobacco Mosaic Virus (TMV), kiwifruit ulcer (Pseudomonas syringae) and Sclerotinia sclerotiorum (Sclerotinia sclerotiorum) on various crops; rich resources, simple preparation method and low cost.

Drawings

The accompanying drawings, which are included to provide a further understanding of the disclosure and are incorporated in and constitute a part of this specification, illustrate embodiments of the disclosure and together with the description serve to explain the disclosure without limiting the disclosure. In the drawings:

FIG. 1 shows the activity change of SOD after spraying Galla chinensis extract resistance inducer on tobacco leaves;

FIG. 2 shows the activity change of PAL after spraying Galla chinensis extract resistance inducer on tobacco leaves.

Detailed Description

The following detailed description of specific embodiments of the present disclosure is provided in connection with the accompanying drawings. It should be understood that the detailed description and specific examples, while indicating the present disclosure, are given by way of illustration and explanation only, not limitation.

Gallnut (Rhus chinensis Mill.) mentioned in the present invention is a kind of aphid female insects belonging to the family Homoptera Migophyridae (Pemphigae Mill.) parasitizing on Rhus chinensis Mill (Rhus chinensis Mill.) of the family Anacardiaceae (Anacardiaceae), Populus pulus pulchrus (Rhus potaninii Maxim.), Rhus chinensis Var. sinica Rehd. Wils) and young leaves or petioles of other plants of the same genus, stabbing to generate a cystic polypide gall, and drying to obtain the product. The Chinese Galla chinensis is mainly produced in mountainous areas and hilly areas such as Qinling, Daba mountain, Wudangshan, Wushan, Wuling mountain, Emei mountain, Daro mountain, and Daliang mountain. The elevation is 250-1600 m in vertical distribution, and 500-600 m is more concentrated, so that the resources are rich. Commercial gallnut is of horny fold type, tripe type and gall flower type. The jianbao gallnut is mainly produced in Guizhou, Sichuan, Hubei Hunan, Yunnan and Guangxi provinces. Ganbao gallnut is mainly produced in Hubei, Shaanxi and Jiangxi provinces.

The gallnut extract resistance inducer is prepared by taking a gallnut extract as an active substance and adding a certain proportion of auxiliaries into the active substance to carry out dosage form processing, and can be prepared into environment-friendly preparations such as micro-emulsion, soluble solution or powder. The tests of the inventor prove that the Chinese gall micro-emulsion with 10 percent of Chinese gall extract, the Chinese gall micro-emulsion with 50 percent of Chinese gall extract, the Chinese gall soluble solution with 10 percent of Chinese gall extract, the Chinese gall soluble solution with 50 percent of Chinese gall extract powder, the Chinese gall extract powder with 10 percent of Chinese gall extract powder and the Chinese gall extract powder with 50 percent of Chinese gall extract powder have good prevention effect on Tobacco Mosaic Virus (TMV), Chinese gooseberry ulcer (Pseudomonas syringae) and sclerotinia sclerotiorum (sclerotinia sclerotiorum).

Preparing a gallnut extract:

crushing Chinese gall into powder with the particle size of 1-5 mm, and adding an extraction solvent at room temperature in a ratio of 1:10(w/v), wherein the extraction solvent is any one or more of methanol, ethanol, ethyl acetate, chloroform and petroleum ether. Standing at room temperature for 24 hr, and extracting in an ultrasonic cleaner with frequency of 100Hz at 40 deg.C for 60 min. After filtration, the residue was extracted 2 times and the three filtrates were combined. Concentrating the filtrate under reduced pressure to obtain plant extract as effective component of Galla chinensis plant source resistance inducer (the extraction solvent in the embodiment of the invention is ethanol).

A plant-derived resistance inducer comprises a Chinese gall extract as an effective component, wherein the Chinese gall extract accounts for 10-50% of the mass of the resistance inducer.

A plant source resistance inducer is a Chinese gall extract microemulsion, a Chinese gall extract soluble solution and a Chinese gall extract powder, wherein:

the Chinese gall extract microemulsion is prepared from the following raw materials in percentage by weight:

10-50% of Chinese gall extract, organic solvent: 10% -40%, surfactant: 5% -25%, antifreeze: 4% -8%, antimicrobial agent: 0.1 to 1 percent of water, and the balance of water, wherein the sum of the weight percentages of the raw materials is 100 percent; (ii) a

The gallnut extract soluble solution is prepared from the following raw materials in percentage by weight:

10-50% of Chinese gall extract, organic solvent: 10% -40%, surfactant: 5% -30%, antifreeze: 1 to 5 percent of ethanol, and the balance of ethanol, wherein the sum of the weight percentages of the raw materials is 100 percent;

the Chinese gall extract powder is prepared from the following raw materials in percentage by weight:

The filler is one or a mixture of more than two of sodium sulfate, ammonium sulfate, attapulgite, kaolin, white carbon soil, argil and light calcium carbonate;

the organic solvent is one or a mixture of more than two of methanol, ethanol, acetone, ethyl acetate, isopropanol and N, N-dimethylformamide;

the surfactant is one or a mixture of more than two of alkyl benzene sulfonate, carboxylate, sulfate, alkyl polyoxyethylene ether, alkylphenol polyoxyethylene ether, polyaryl polyoxyethylene ether and tween;

the antifreezing agent is one of glycol, glycerol, urea and glycerol;

the antimicrobial agent of the present invention is one of 2-hydroxybiphenyl, sorbic acid and benzoic acid.

The dispersant is one or a mixture of more than two of alkyl sulfonate, alkyl phenol or fatty alcohol ethylene oxide adduct sulfonate and phosphate of alkyl phenol or fatty alcohol ethylene oxide adduct;

the stabilizer is one or a mixture of more than two of organic acid, fatty alcohol, diphenyl phosphate, alkyl sulfonate, diisopropyl phosphate and isopropyl phosphate;

the preparation method of the gallnut extract microemulsion comprises the following steps:

fully dissolving and mixing the nutgall extract, the organic solvent, the surfactant and the antimicrobial agent in a mixing kettle according to a proportion; mixing an antifreezing agent and water in proportion, slowly adding into a mixing kettle, and uniformly stirring and dissolving; and (3) adjusting the temperature in the mixing kettle to 50-60 ℃, stirring for 30min, and cooling to obtain the gallnut extract microemulsion.

The preparation method of the gallnut extract soluble solution comprises the following steps:

the gallnut extract, the organic solvent, the surfactant, the antifreezing agent and the ethanol are mixed in a mixing kettle according to a proportion, fully stirred for 30min at a rotating speed of 60-80 r/min, and stirred uniformly to obtain the gallnut extract soluble liquid.

The preparation method of the gallnut extract powder comprises the following steps:

dissolving Galla chinensis extract in organic solvent; uniformly mixing the filler, the dispersant and the stabilizer in proportion, and sieving by a 100-mesh sieve; and (3) fully and uniformly spraying the dissolved gallnut solution on the mixture, and uniformly mixing to obtain the gallnut extract powder.

For better understanding of the essence of the invention, the technical contents of the invention will be described in detail with examples, but the invention is not limited to these examples.

Example 1: determination of anti-TMV activity of tobacco induced by gallnut extract

Two varieties of the heart-leaf tobacco and the common tobacco (K326) are selected for induced disease resistance activity verification. TMV infected heart leaf tobacco can form virus withered spots, the symptom infected by common tobacco is flower leaves, and different symptoms are counted by different methods. And respectively spraying 0.2, 1 and 5mg/mL gallnut extract solutions to the lower three leaves of the tobacco with consistent growth vigor in the 6-7 leaf stage. TMV was inoculated to the non-sprayed upper leaves after 48 h. The blank control is treated with clear water, and the positive control is BTH solution. Each plant was inoculated with 2-3 leaves, each treatment included 10 tobacco plants, and the entire experiment was repeated 3 times. And after 3d, counting the number of the heart-leaf tobacco withered spots and the disease index of the common tobacco, and calculating the inhibition rate formula as above and the prevention and treatment effect formula as below.

Disease index ∑ (number of disease stages × number of diseased plants)/(highest number of disease stages × total number of treated plants) × 100

The control effect is (control disease index-treatment disease index)/control disease index is multiplied by 100 percent

The results are shown in Table 1.

Table 1: induced disease resistance effect of gallnut extract on TMV

Note: data are mean ± sem, significant differences between 3 activities were tested by Duncan Multiple Range Test (DMRT) in SPSS software, and different letters indicated significant differences at the 0.05 level between data.

As can be seen from Table 1, the Galla chinensis extract showed good effect of inducing resistance to TMV at a concentration of 1mg/mL on 2 varieties of tobacco, which is superior to the control medicament BTH, indicating that the topical application of Galla chinensis extract can induce the non-application part of tobacco to generate disease resistance, and achieve resistance to TMV. The disease resistance of the common tobacco is slightly better than that of the heart-leaf tobacco, probably because the heart-leaf tobacco is a resistant variety, the induced disease resistance of the Chinese gall extract is more obvious on the common tobacco. The result shows that the control effect of the gallnut extract treated by the concentration of 1mg/mL and 5mg/mL has no obvious difference, while the control effect of the gallnut extract treated by the concentration of 0.2mg/mL is weaker, which indicates that the control effect of the gallnut extract can achieve better control effect when the control effect is 1 mg/mL. From the perspective of saving resources and reducing phytotoxicity, 1mg/mL is a reasonable concentration of drug delivery.

Example 2: galla chinensis extract causes activity change of tobacco defense enzyme SOD

Spraying Galla chinensis extract solution 0.2, 1 and 5mg/mL onto leaf, collecting leaf on days 1,3,5,7, and 9, respectively, and detecting activity changes of defense enzymes SOD and PAL.

0.5g of tobacco leaf was placed in a precooled mortar, 2.0mL of precooled PBS (50mmol/L, pH8.8 phosphate buffer) was added and ground into slurry in an ice bath, and buffer was added to make the final volume 5.0 mL. Centrifuging at 4 deg.C for 15min at 10,000r/min, collecting supernatant, and measuring Volume (VT) to obtain SOD crude extract.

2) And (3) activity determination: taking 7 test tubes, 3 test tubes and 4 control tubes (3 maximum photoreduction tubes and 1 reference tube) of 10mL test tubes, and adding the solutions in sequence according to the table 1.

TABLE 1 order and amount of reagents added to each test tube

After the total volume of the reaction system is 3.0mL and mixed uniformly, 1 control tube is placed in the dark, and other tubes (the other 3 control tubes are used as maximum photoreduction tubes) react for 20min under 4000lx sunlight (the conditions of all the tubes are required to be consistent, the reaction temperature is controlled to be 25-35 ℃, and the reaction time can be properly adjusted according to the enzyme activity). After the reaction was completed, the reaction was stopped by covering the reaction vessel with a black cloth. The absorbance values of the other tubes were measured at 560nm wavelength, respectively, with reference to the control tubes not illuminated. And (3) calculating enzyme activity:

wherein V1 is the amount of the sample mL in the determination process.

The results of the SOD activity measurement are shown in FIG. 1. FIG. 1 illustrates that the treatment of tobacco with Galla chinensis extract at different concentrations can cause a significant increase in SOD activity in tobacco. Among them, the activity of SOD was significantly increased by high concentration treatment. The SOD activity after 24 hours under the treatment of 5mg/mL gallnut extract reaches 2.31 times of that of the control group. The results show that the Chinese gall extract can activate the activity of defense enzyme SOD in the tobacco body and increase the disease resistance of the tobacco.

Example 3: galla chinensis extract causes activity change of tobacco defense enzyme PAL

1) Enzyme solution extraction, namely taking 1.25g of tobacco leaves, adding precooled 5mL of enzyme extract (0.1mol/L of Tris-H with pH8.8)₂SO₄Buffer) and 0.5g of polyvinylpyrrolidine (PVP) (to remove phenolic poisons and prevent color interference), homogenized in a mortar or tissue triturator. Filtering with 4 layers of gauze, centrifuging the filtrate at 4 deg.C and low temperature for 30min at 10,000r/min, collecting supernatant, measuring volume and recording.

2) The activity of the enzyme is measured by taking 1mL of 0.2mol/L phenylalanine solution and 2mL of 0.1mol/L Tris-H₂SO₄Buffer (pH 8.8) (3 mL of buffer was directly taken without substrate phenylalanine in the control tube) and incubated for 3min in a 30 ℃ water bath. 0.5mL of enzyme solution to be detected is added into each test tube, no enzyme solution is added into a blank tube, the initial value is measured at the wavelength of 209nm immediately after shaking up, the blank tube is used for zero setting, and the time is accurately counted. Placing each test tube in 30 deg.C water bath, reacting for 30min, and measuring A again₂₀₉。

3) And (3) calculating enzyme activity: the amount of enzyme required for an increase in optical density of 0.01 at 290nm per hour was 1 unit (U) in terms of the fresh weight of the plant.

Wherein delta A is the absorbance difference of the front and back 2 times of measurement; w is the sample fresh weight (g); t is the reaction time (30 min).

The results of the PAL activity assay are shown in FIG. 2. FIG. 2 illustrates that treatment of tobacco with varying concentrations of Galla rhois extract resulted in a significant increase in PAL activity in tobacco. Within 48 hours after treatment, the activity of the enzyme increased rapidly; at 48 hours, the enzyme activity reached a maximum; after 48 hours, the activity of the enzyme decreased rapidly. Wherein PAL activity after 48 hours was 4.88 times that of the control group by treatment with 5mg/mL of Galla rhois extract. The results show that the gallnut extract can activate the activity of the defense enzyme PAL in the tobacco body and increase the disease resistance of the tobacco

Example 4: preparation of 10% gallnut extract microemulsion

Weighing 1kg of nutgall extract concentrate, dissolving the nutgall extract concentrate in 1kg of ethyl acetate, adding 1kg of calcium dodecyl benzene sulfonate, 0.5kg of styrene acid polyoxyethylene ether and 0.1kg of benzoic acid, and fully stirring the mixture in a mixing kettle at the temperature of about 40 ℃ at 60-80 r/min for 30 min; mixing 0.1kg of urea and 6.3kg of deionized water until the urea and the deionized water are fully dissolved, slowly adding the mixture into a mixing kettle, and starting stirring; and (3) adjusting the temperature of the mixing kettle to 50-60 ℃, stirring for 30min, and cooling to obtain 10kg of 10% gallnut microemulsion. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 5: preparation of 50% gallnut extract microemulsion

Weighing 5kg of nutgall extract concentrate, dissolving the nutgall extract concentrate in 3kg of ethyl acetate, adding 1kg of calcium dodecyl benzene sulfonate, 0.5kg of styrene acid polyoxyethylene ether and 0.1kg of benzoic acid, and fully stirring the mixture in a mixing kettle at the temperature of about 40 ℃ at 60-80 r/min for 30 min; mixing 0.1kg of urea and 0.3kg of deionized water until the urea and the deionized water are fully dissolved, slowly adding the urea and the deionized water into a mixing kettle, and starting stirring; and (3) adjusting the temperature of the mixing kettle to 50-70 ℃, stirring for 30min, and cooling to obtain 10kg of 50% gallnut microemulsion. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 6: preparation of 10% gallnut extract soluble agent

Weighing 1kg of nutgall extract concentrate, dissolving the nutgall extract concentrate in 1kg of ethyl acetate, adding 0.5kg of alkylphenol polyoxyethylene, 1kg of calcium dodecyl benzene sulfonate, 0.5kg of ethylene glycol and ethanol to supplement the mixture to 10kg, mixing the mixture uniformly, fully stirring the mixture for 30min at the stirring speed of 60-80 r/min, and stirring the mixture uniformly to obtain 10kg of 10% nutgall soluble agent. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 7: preparation of 50% gallnut extract soluble agent

Weighing 5kg of nutgall extract concentrate, dissolving the nutgall extract concentrate in 2.5kg of ethyl acetate, adding 0.5kg of alkylphenol polyoxyethylene ether, 1kg of calcium dodecyl benzene sulfonate, 0.5kg of ethylene glycol and 10kg of ethanol, fully stirring for 30min at a stirring speed of 60-80 r/min after uniformly mixing, and preparing 10kg of 50% nutgall soluble solution after uniformly stirring. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 8: preparation of 10% gallnut extract powder

Weighing 1kg of Galla chinensis extract concentrate, and dissolving in 2.5kg of ethyl acetate; uniformly mixing 5kg of sodium sulfate and 1.5kg of calcium dodecyl benzene sulfonate, and sieving by using a 100-mesh sieve; and (3) fully and uniformly spraying the dissolved gallnut solution on the mixture, and uniformly mixing to obtain 10kg of 10% gallnut powder. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 9: preparation of 50% gallnut powder

Weighing Galla chinensis extract concentrate 5kg, and dissolving in ethyl acetate 3.5 kg; 1kg of sodium sulfate and 0.5kg of calcium dodecyl benzene sulfonate are uniformly mixed and sieved by a 100-mesh sieve; and (3) fully and uniformly spraying the dissolved gallnut solution on the mixture, and uniformly mixing to obtain 10kg of 50% gallnut powder. The stability, appearance and the like of the preparation meet the requirements of commercial preparations.

Example 10: galla chinensis inducer for field prevention of tobacco mosaic virus

The cell test is random arrangement, repeated for 3 times, and the cell area is equal to 60m²The selection of the test field requires uniform fertility and consistent crop planting and management level, and protective rows are arranged between each treatment room and around the test area. Spraying 10% and 50% Galla chinensis resistance inducer 100 times and 500 times respectively, and adding 50% benzothiadiazole water dispersible granule (BTH)5000 times solution as control agentSpraying on the leaf surface in a constant amount, and setting clear water for comparison; all test agents must be diluted twice. Spraying the tobacco seedlings from 4-5 leaf periods of common tobacco seedlings, and then spraying the tobacco seedlings for 1 time every 4 days for 3 times. And (3) carrying out friction inoculation on the whole leaf after spraying for 2d for the last time, inoculating 3 leaves for each plant, repeating the steps for three times after each 10 plants are treated, investigating the disease index of each treatment after 10d of virus inoculation, and calculating the control effect.

Disease grading standard according to the tobacco industry standard of the people's republic of China-the tobacco mosaic disease severity grading survey method (YC/T39-1996):

level 0: the whole plant is disease-free;

level 1: the heart and leaf vein is clear or the flower and leaf are slight, or the leaf and leaf of the upper 1/3 leaves are not deformed, and the plant is not obviously dwarfed;

and 2, stage: 1/3 to 1/2 leaf lobes, or a few leaf lobes, are deformed; or the main pulse is blackened, the plant is dwarfed to be higher than 2/3 with normal plant height;

and 3, level: 1/2-2/3, or the main side pulse is changed to black, the plant is dwarfed to the normal plant height of 1/2-2/3;

4, level: the whole leaf has severe deformity or necrosis, and the diseased plant is dwarfed to the normal plant height of l/3 to 1/2. For refining the investigation result, on the basis of the above severity grading, the grading standard is refined, i + grade is added between 1 and 2 grades, 2+ grade is added between 2 and 3 grades, 3+ grade is added between 3 and 4 grades, and the grades are marked as 1.5, 2.5 and 3.5:

1+ level: the heart leaves have bright veins or slight flower leaves, or the upper 1/3 leaf leaves are slightly shrunken, and plants are not obviously dwarfed;

2+ level: l/3 to 1/2 leaves and leaves are deformed or the main pulse is blackened, and the plant is dwarfed to be more than 2/3 of a normal plant;

3+ level: 1/2-2/3 leaf mosaic, or deformed or primary side vein necrotic, or plant dwarfing to 1/2 of normal plant height.

And calculating the disease index according to the severity, and measuring the effect of different treatments by using the prevention and treatment effect.

Disease index ═ Σ [ (number of infected plants × graded representative value of severity)/(number of total investigated plants × highest representative value of severity) ] × 100

Control effect%

The results are shown in Table 3;

table 3: experiment of field plot pesticide effect of 3 gallnut resistance inducer for preventing tobacco virus diseases

From the table, 10% and 50% of the gallnut resistance inducer has good prevention effect on tobacco mosaic virus, and the inhibition rate of 50% of the gallnut powder on the tobacco mosaic virus reaches 66.78% at the maximum under the condition of 500-fold dilution, and is superior to the control effect of 58.86% of benzothiadiazole.

Example 11: galla chinensis inducer for field prevention of kiwifruit ulcer

The cell test is random arrangement, repeated for 3 times, and the cell area is equal to 60m²The selection of the test field requires uniform fertility and consistent crop planting and management level, and protective rows are arranged between each treatment room and around the test area. Respectively carrying out constant spraying on the leaf surfaces by using 100 times and 500 times of solutions of 10% and 50% gallnut resistance inducer, carrying out constant spraying on the leaf surfaces by using 5000 times of solution of 50% benzothiadiazole water dispersible granule as a control medicament, and setting clear water for control; all test agents must be diluted twice. And selecting new leaves with the same size after the young shoots of the kiwi fruit tree emerge, spraying for 1 time every 4 days, and spraying for 3 times in total. And spraying bacterial suspension after the quartz sand is rubbed and damaged after spraying for 2d for the last time, repeating the steps for three times after 10 leaves are treated, investigating the inhibition rate of the scab and the inhibition rate of the scab expansion after inoculating bacteria for 10d, and calculating the control effect.

Inhibition (%) - (control number of scorched spots-number of treated scorched spots)/control number of scorched spots × 100

Inhibition of lesion extension (%) (control lesion area-treated lesion area)/control lesion area × 100

The results are shown in Table 4;

table 4: galla chinensis inducer for field prevention of kiwifruit ulcer

From the above table, the gallnut resistance inducer has a good prevention effect on the kiwifruit ulcer. The 50% gallnut microemulsion has the highest inhibition rate of 51.70% on lesion spots and 46.43% inhibition rate of lesion spot expansion when diluted by 500 times, and is superior to the inhibition rate of 51.09% and 43.98% of the control medicament benzothiadiazole.

Example 12: galla chinensis resistance inducer for field control of sclerotinia rot of colza

The rape variety is Qin oil No. 10. The cell test is random arrangement, repeated for 3 times, and the cell area is equal to 60m²The selection of the test field requires uniform fertility and consistent crop planting and management level, and protective rows are arranged between each treatment room and around the test area. Respectively carrying out constant spraying on the leaf surfaces by using 100 times and 500 times of solutions of 10% and 50% gallnut resistance inducer, carrying out constant spraying on the leaf surfaces by using 5000 times of solution of 50% benzothiadiazole water dispersible granule as a control medicament, and setting clear water for control; all test agents must be diluted twice. Inoculating sclerotinia sclerotiorum in a PDA plate, and culturing at 25 ℃ for 3d for germ inoculation. In the 6-8 leaf period of the rape seedlings, selecting sunny days, spraying stem and leaf surfaces by adopting an industrial and agricultural 16-type manual sprayer for 2 times at the pesticide application interval of 7 days, wherein the liquid consumption is 10kg/60m². And inoculating sclerotinia sclerotiorum after spraying the pesticide for 2 days for the last time. The specific inoculation method of the germs comprises the following steps: carrying out moisture-keeping culture on sclerotinia sclerotiorum cakes with the diameter of 6cm at 20 ℃, inoculating hypha faces downwards to the base part of the leaf middle veins, spraying diazosulfide and clear water, and inoculating plants with pathogenic fungus hypha blocks as a control. The test is repeated 3 times by treating 10 strains, 1 hypha block, 1 leaf and 3 leaves, 1 strain and 30 inoculation points in total, and the disease condition is investigated and the control effect is calculated after inoculation for 5 days.

Grading standard of severity of leaf diseases:

level 0: disease free

Level 1: the disease is slightly attacked, and the area of disease spots accounts for 11-30% of the surface area of the main stem;

and 2, stage: moderate morbidity, wherein the lesion area accounts for 31-50% of the whole leaf area;

and 3, level: the disease is highly developed, and the lesion area accounts for 31-50% of the whole leaf area;

4, level: the disease is seriously attacked, and the area of the lesion spots accounts for more than 50 percent of the area of the whole leaf.

The disease index and prevention effect calculation formula is as follows:

disease index ═ Σ (number of diseased leaves at each stage × corresponding stage)/(total number of investigated plants × 4) × 100;

control effect is (1-medicament treatment disease index)/blank control disease index is multiplied by 100%.

Table 5: galla chinensis resistance inducer for field control of sclerotinia rot of colza

From the above table 5, it can be seen that the gallnut resistance-inducing agent has a good prevention effect on sclerotinia rot of colza, and the maximum prevention effect of the 50% gallnut microemulsion can reach 63.91% under the concentration of 500 times dilution, which is superior to the control agent benzothiadiazole prevention effect 60.82%.

Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

The preferred embodiments of the present disclosure are described in detail with reference to the accompanying drawings, however, the present disclosure is not limited to the specific details of the above embodiments, and various simple modifications may be made to the technical solution of the present disclosure within the technical idea of the present disclosure, and these simple modifications all belong to the protection scope of the present disclosure.

It should be noted that, in the foregoing embodiments, various features described in the above embodiments may be combined in any suitable manner, and in order to avoid unnecessary repetition, various combinations that are possible in the present disclosure are not described again.

In addition, any combination of various embodiments of the present disclosure may be made, and the same should be considered as the disclosure of the present disclosure, as long as it does not depart from the spirit of the present disclosure.

Claims

1. Application of Galla chinensis extract in preparing plant resistance inducer is provided.

2. The use of claim 1, wherein the gallnut extract is a plant extract obtained by extracting gallnuts with an organic solvent.

3. The use of claim 1 or 2, wherein the gallnut extract is contained in the plant resistance inducer in an amount of 10 to 50% by mass.

4. The use according to claim 1 or 2, wherein the preparation method of the gallnut extract comprises:

5. The resistance inducer is characterized by taking a Chinese gall extract as an effective component, wherein the mass percentage of the Chinese gall extract in the resistance inducer is 10-50%.

6. The resistance inducer according to claim 5, wherein the Galla chinensis extract is a plant extract obtained by extracting Galla chinensis with organic solvent.

7. The resistance inducer according to claim 5, wherein the preparation method of the Galla chinensis extract comprises:

8. The resistance inducer is characterized by being a gallnut extract microemulsion which is prepared from the following raw materials in percentage by weight:

9. The resistance inducer is characterized by being a gallnut extract soluble solution, wherein the soluble solution is prepared from the following raw materials in percentage by weight:

10. The resistance inducer is characterized by being gallnut extract powder, wherein the powder is prepared from the following raw materials in percentage by weight: