CN111771911B - Microbial preparation for preventing and treating tobacco mosaic virus and preparation method thereof - Google Patents
Microbial preparation for preventing and treating tobacco mosaic virus and preparation method thereof Download PDFInfo
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/22—Bacillus
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/50—Isolated enzymes; Isolated proteins
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
- A01N65/08—Magnoliopsida [dicotyledons]
- A01N65/38—Solanaceae [Potato family], e.g. nightshade, tomato, tobacco or chilli pepper
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- Dentistry (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Plant Pathology (AREA)
- Agronomy & Crop Science (AREA)
- Wood Science & Technology (AREA)
- Environmental Sciences (AREA)
- Pest Control & Pesticides (AREA)
- Virology (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
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Abstract
The invention provides a microbial preparation for preventing and treating tobacco mosaic virus, which comprises a microbial mixed microbial inoculum, a plant immunity inducer and a leaf extract, wherein the leaf extract is the leaf extract of a tobacco plant which generates immune reaction after being sprayed with the plant immunity inducer, the mass percentages of the microbial mixed microbial inoculum, the plant immunity inducer and the leaf extract are 5-10% of the microbial mixed microbial inoculum, 3-5% of the plant immunity inducer and 10-15% of the leaf extract, the balance is sterile water, and the plant immunity inducer is a coat protein solution of the tobacco mosaic virus; the plant immunity inducer plays a role of an immunity exciting factor, can trigger a plant immune system to generate stress immune response and induce the plant immune system to generate immune resistance against tobacco mosaic virus, and meanwhile, the leaf extracting solution contains a large amount of biochemical substances capable of limiting virus proliferation, and plays a certain virus prevention and control role when the immune response of a plant does not fully exert efficacy.
Description
Technical Field
The invention relates to the technical field of microbial preparations, in particular to a microbial preparation for preventing and treating tobacco mosaic virus and a preparation method thereof.
Background
Tobacco Mosaic Virus (TMV) is a single-stranded RNA virus that is extremely infectious and pathogenic to plants. The tobacco mosaic virus disease is widely distributed in various tobacco regions in China and is the main virus disease of tobacco.
In the past, chemical pesticides and transgenic breeding methods are mainly adopted for preventing and treating tobacco mosaic virus diseases. Chemical pesticides, however, cause increasingly significant viral resistance and high levels of pesticide residues; also, although transgenic technology can effectively control the occurrence and development of viral diseases, the biological safety of transgenic technology has attracted high attention from experts and governments at home and abroad.
Therefore, the technical personnel researches the microorganism control method, the principle is that the microorganism microbial inoculum is sprayed on the plant leaves, microorganism beneficial bacteria are attached to the tobacco leaves, biochemical substances which are beneficial to the plants and can improve the disease-resistant and immunity capabilities of the plants are released through normal life activities, and the biochemical substances are absorbed and utilized by the plants, so that a certain control effect is achieved.
However, the beneficial activity of the microbial flora is that the disease-resistant immunity of the whole organism of the plant is improved, but not the resistance of the plant is specially directed at the tobacco mosaic virus, so that after the plant is infected with the tobacco mosaic virus, a longer time is needed for carrying out an immune reaction, and even if the infection condition of the virus is limited successfully subsequently, the plant is damaged to a certain extent, so that the quality of the tobacco leaves is reduced.
Meanwhile, the microbial flora needs a long time for adapting to the growth environment and expanding the number of the flora, and rainfall or spraying liquid medicine to the leaves can also affect the attachment of the microbial flora on the leaves to a certain degree, so that the microbial control effect is ensured, the microbial control agent needs to be sprayed for multiple times, and the planting work intensity is high.
Disclosure of Invention
In view of the above, the invention provides a microbial preparation for preventing and treating tobacco mosaic virus, which has strong targeting pertinence, good disease-resistant immunity effect and long-lasting prevention and treatment effect, and a preparation method thereof.
The technical scheme of the invention is realized as follows: the invention provides a microbial preparation for preventing and treating tobacco mosaic virus, which comprises a mixed microbial inoculum, a plant immunity inducer and a leaf extracting solution, wherein the leaf extracting solution is a leaf extracting solution of a tobacco plant which generates immune reaction after being sprayed with the plant immunity inducer, and the mass percentages of the mixed microbial inoculum, the plant immunity inducer and the leaf extracting solution are 5-10% of the mixed microbial inoculum, 3-5% of the plant immunity inducer, 10-15% of the leaf extracting solution and the balance of sterile water; the microbial mixed microbial inoculum comprises bacillus amyloliquefaciens and lactobacillus plantarum, wherein the mass ratio of the bacillus amyloliquefaciens to the lactobacillus plantarum is (2-3): (2-3); the plant immunity inducer is tobacco mosaic virus capsid protein solution.
On the basis of the technical scheme, the cleaning solution preferably further comprises a surfactant, wherein the surfactant is tween 80, organic silicon, ethoxy modified polytrisiloxane or sodium sulfosuccinate.
More preferably, the mass of the surfactant accounts for 1-3% of the mass of the solution.
More preferably, the mass percent of the tween 80 is 3%, or the mass percent of the organic silicon is 2%, or the mass percent of the ethoxy modified polytriesiloxane is 1%, or the mass percent of the sodium sulfosuccinate is 1.5%.
In a second aspect, a method for preparing a microbial preparation for controlling tobacco mosaic virus is provided, which comprises the following steps,
s1, preparing seed solutions of various strains of the microbial mixed inoculant, fermenting and propagating the seed solutions respectively, and uniformly mixing propagation products according to the proportion;
s2 adding acetic acid into tobacco mosaic virus solution, mixing uniformly, standing, centrifuging to remove RNA precipitate, separating supernatant through desalting column, removing acetic acid, dialyzing the obtained solution in deionized water for 12-24 h, centrifuging, re-dissolving the obtained precipitate with phosphoric acid buffer solution to obtain tobacco mosaic virus capsid protein solution;
s3, spraying the tobacco mosaic virus capsid protein solution obtained in the step S2 on leaves of tobacco plants, picking the leaves of the plants after the tobacco plants generate immunoreaction, crushing the leaves, extracting mixed liquor by a water extraction method, and filtering solid impurities to obtain a leaf extracting solution;
s4, uniformly mixing the products obtained in the steps S1-S3 according to the proportion to obtain a mixed solvent;
s5 adding the surfactant into the mixed solvent in the step S4 according to the mixture ratio, and stirring for 1-2 h.
On the basis of the above technical solution, preferably, in step S3, after spraying the tobacco mosaic virus capsid protein solution on the leaves of the tobacco plant for 1 to 2 hours, extracting the leaf extract, then separating a single component from the leaf extract by using ion separation chromatography and high performance liquid chromatography, and obtaining the composition of the single component by using a mass spectrometry identification method to identify whether the tobacco plant produces an immunoreaction product.
Compared with the prior art, the microbial preparation for preventing and treating tobacco mosaic virus and the preparation method thereof have the following beneficial effects:
(1) the plant immunity inducer plays a role of an immunity exciting factor, after contacting tobacco plant leaves, capsid protein in a liquid agent can trigger a plant immune system to generate stress immune response to induce the plant immune system to generate immune resistance aiming at tobacco mosaic virus, meanwhile, a large amount of biochemical substances capable of limiting virus proliferation are contained in a leaf extracting solution, and when the immune response of the plant does not fully exert efficacy, the biochemical substances in the leaf extracting solution play a certain virus prevention and control effect, so that the plant specific immunity is effectively improved, and the plant immunity inducer is safe and reliable, and avoids the risks of drug residue and drug resistance.
(2) The microbial mixed bacterial agent can effectively improve the basic immunity of plants as a beneficial flora, and the microbial flora can also generate various substances, promote the absorption and utilization of nutrient substances by the plants, greatly improve the soil structure and increase the soil fertility after entering soil along with rainfall, and further improve the planting quality of tobacco leaves.
(3) The surfactant can enhance the dispersive adhesion capacity of microbial flora on tobacco leaves, reduce the adverse effect of precipitation on the adhesion of the flora, prolong the time for the flora to exert the effect on the leaves and further improve the microbial control effect.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without any inventive step based on the embodiments of the present invention, are within the scope of the present invention.
The first embodiment is as follows:
the invention provides a microbial preparation for preventing and treating tobacco mosaic virus, which comprises a microbial mixed microbial inoculum, a tobacco mosaic virus capsid protein solution and a leaf extract, wherein the leaf extract is the leaf extract of a tobacco plant which produces immune reaction after being sprayed with a plant immune inducer, and the mass percentages of the microbial mixed microbial inoculum, the plant immune inducer and the leaf extract are 10 percent of the microbial mixed microbial inoculum, 5 percent of the plant immune inducer, 15 percent of the leaf extract and the balance of sterile water; the microbial mixed microbial inoculum comprises bacillus amyloliquefaciens and lactobacillus plantarum, wherein the mass ratio of the bacillus amyloliquefaciens to the lactobacillus plantarum is 1: 1.
the preparation method comprises the following steps of,
s1, preparing seed solutions of various strains of the microbial mixed inoculant, fermenting and propagating the seed solutions respectively, and uniformly mixing propagation products according to the proportion;
s2 adding acetic acid into tobacco mosaic virus solution, mixing uniformly, standing, centrifuging to remove RNA precipitate, separating supernatant through desalting column, removing acetic acid, dialyzing the obtained solution in deionized water for 12-24 h, centrifuging, re-dissolving the obtained precipitate with phosphoric acid buffer solution to obtain tobacco mosaic virus capsid protein solution;
s3, spraying the tobacco mosaic virus capsid protein solution obtained in the step S2 on leaves of tobacco plants, picking the leaves of the plants after the tobacco plants generate immunoreaction, crushing the leaves, extracting mixed liquor by a water extraction method, and filtering solid impurities to obtain a leaf extracting solution;
s4, uniformly mixing the products obtained in the steps S1-S3 according to the proportion to obtain a mixed solvent;
s5 adding the surfactant into the mixed solvent in the step S4 according to the mixture ratio, and stirring for 1-2 h.
In step S3, after spraying tobacco mosaic virus capsid protein solution on the leaves of the tobacco plant for 1 to 2 hours, extracting the leaf extract, separating a single component from the leaf extract by ion separation chromatography and high performance liquid chromatography, and obtaining the composition of the single component by mass spectrometry to identify whether the tobacco plant produces an immunoreaction product.
Example two:
the mass percentages of the mixed microbial inoculum, the plant immunity inducer and the leaf extract in the first embodiment are 5% of the mixed microbial inoculum, 3% of the plant immunity inducer and 10% of the leaf extract, and other conditions are kept unchanged.
Example three:
besides the conditions of the embodiment are kept unchanged, the detergent also comprises a surfactant.
Wherein the surfactant is Tween 80, and the mass of the Tween 80 accounts for 3% of the mass of the solution.
Bacillus amyloliquefaciens and Lactobacillus plantarum are probiotic strains widely existing in the nature, belong to gram-positive bacteria, and are widely applied to the fields of food fermentation, industrial production, planting and breeding, medical care and the like or are widely used as biocontrol bacteria. The two can release a large amount of biochemical substances and nutrient substances beneficial to the growth of plants into the environment after being sprayed to the plants or the soil as microbial preparations, and can greatly improve the soil structure, further promote the growth of the plants and improve the disease-resistant immunity of the plants.
Compared with the reaction mechanism of the immune system of higher animals and human beings, the reaction mechanism of the plant immune system is more special. Plants and animals are distinguished in that plants do not have a complete circulatory system, so that immune cells cannot be directly generated in blood and transported to a pathogenic site through the circulatory system to perform an immune reaction, as in animals. The mechanisms of the plant immune system are therefore independently responsive to infection, and each requires the presence of the correct amount of a particular protein, called immunoreceptor, in the correct location to respond accordingly, or in the correct combination, to allow the plant to respond effectively and efficiently to the immune response, to promote the death of plant cells at the site of infection, to spread the pathogen across the tissue, while the tissues at other healthy sites are regulated by hormonal signals to develop resistance in advance to the attack of the pathogen. Meanwhile, the plant immune system also has a systemic anaphylactic reaction similar to that of the animal immune system, namely a certain memory function.
The immune response of the plant immune system is divided into two types, namely PTI and ETI.
When the pathogen infects in the early stage, the pathogen can secrete some toxic factors to attack plant cells; in order to avoid pathogen attack, plants gradually evolve cell membrane surface pattern recognition receptors for recognizing pathogen-associated molecular patterns conserved in pathogen inheritance and stimulating the basic immune response of plants, namely PTI. This is a ubiquitous but weak resistance to infection by a variety of pathogens.
When the infection of the pathogen is aggravated, the pathogen can evolve effector molecules to enter plant cells, so that PTI (protein-regulated indicator) generated by the plant is inhibited, and the disease resistance of the plant is overcome; the plant can evolve corresponding disease-resistant protein aiming at the pathogenic effect molecules and generate new immunoreaction, namely ETI; pathogens escape recognition through variation of effector molecules, or secrete new effector molecules to interfere with ETI. The immune system and pathogens of plants are co-evolved in this constant battle until one wins the elimination of pathogens or the plant is pathogenic.
Plant ETI has stronger disease resistance compared to PTI, but only targets a single pathogen, and nutrients for immune response in the plant immune system are limited, so that PTI exerts efficacy to delay or reduce the occurrence and development of diseases at the initial stage of pathogen infection, and ETi requires a certain reaction time to exert full efficacy.
Therefore, the tobacco mosaic virus capsid protein is adopted as a plant immune inducer to induce a plant immune system to generate a systemic anaphylactic reaction to the capsid protein so as to generate immune memory, which is beneficial to the targeted excitation of ETI when the plant is infected by virus; meanwhile, the leaf extract is extracted from tobacco plants which have undergone immune reaction, so that the solution contains a large amount of biochemical substances capable of limiting virus proliferation, and a certain virus prevention and treatment effect can be achieved when the immune reaction of the plants does not fully exert efficacy. And genetic materials of the tobacco mosaic virus are not contained in the coat protein of the tobacco mosaic virus, so that the coat protein can not infect plants to cause pathogenicity, is safe and reliable, and avoids the problems of drug resistance enhancement or drug residue.
Tween 80, organic silicon, ethoxy modified polytrisiloxane or succinic acid diester sodium sulfonate are common surfactants, and when the surfactants are added into the liquid, the dispersion coefficient of the liquid and the adhesion capability of the liquid can be increased, the liquid is prevented from converging on leaves to form liquid drops, and the leaf surface adhesion effect of the liquid containing the microbial agent is further improved.
Test methods and analysis of results:
the formulations of examples 1 to 3 were diluted to 1000 times with water and stirred uniformly for use, and tobacco mosaic virus suspensions were prepared for use. The test is carried out in a green insect-proof greenhouse to reduce errors caused by other objective factors.
The identification method of the tobacco leaves infected with tobacco mosaic virus disease is performed according to GB/T23222-2008 (tobacco pest and disease damage grading and investigation method).
1. Passivation experiment
Taking tobacco seedling as a dead-spot host, respectively and uniformly mixing 180uL of the preparation prepared in the embodiment 1 to 3 with 20uL of tobacco mosaic virus suspension, and standing at room temperature for 1h with intermittent shaking. In addition, sterile water treatment was used as a blank control, and a mixed microbial inoculum treatment of the same concentration was used as a comparative example.
After 1h, accurately taking 20uL of the liquid agent of each test group by using a liquid transfer device, inoculating the liquid agent on the whole leaf of the tobacco seedling plant with consistent position and size, and uniformly smearing the liquid agent by using a medicinal cotton swab.
Then, the above seedlings were cultured at room temperature and humidity of 70 to 80% for 3 to 4 days, and the number of formation of scorched spots was recorded. 6 replicates per sample. The inhibition rate was calculated according to the following formula:
the inhibition ratio was [ (C-T)/C ]. times.100%
Wherein, C represents the number of dead spots formed by the blank control; t represents the number of scorched spots formed after the treatment.
2. Prevention test
The tobacco seedling is used as a dead-spot host to prepare the preparation of the embodiment 1 to 3, and meanwhile, sterile water is used as a blank control, and a mixed microbial inoculum with the same concentration is used as a comparative example.
The liquid of each test group is dipped in absorbent cotton and then is uniformly smeared on the whole leaf of the tobacco seedling plant with consistent position and size, preferably, the liquid does not drip.
After 3h treatment, 20uL of the virus suspension was inoculated onto the treated leaves of each test group and smeared with a medicinal cotton swab.
Then, the above seedlings were cultured at room temperature and humidity of 70 to 80% for 3 to 4 days, and the number of formation of scorched spots was recorded. 6 replicates per sample. The inhibition rate was calculated according to the following formula:
the inhibition ratio was [ (C-T)/C ]. times.100%
Wherein, C represents the number of dead spots formed by the blank control; t represents the number of scorched spots formed after the treatment.
3. Experiment of treatment
The tobacco seedling is used as a dead-spot host to prepare the preparation of the embodiment 1 to 3, and meanwhile, sterile water is used as a blank control, and a mixed microbial inoculum with the same concentration is used as a comparative example.
20uL of the tobacco mosaic virus suspension is taken by a pipette, inoculated on the whole leaf of the tobacco seedling plant with consistent position and size, and evenly smeared by a medicinal cotton swab.
After inoculation for 3h, the liquid of each test group is dipped by medicinal absorbent cotton and is uniformly smeared on the leaves which are inoculated with viruses in advance, and the liquid medicine is preferably not dropped.
Then, the above seedlings were cultured at room temperature and humidity of 70 to 80% for 3 to 4 days, and the number of formation of scorched spots was recorded. 6 replicates per sample. The inhibition rate was calculated according to the following formula:
the inhibition ratio was [ (C-T)/C ]. times.100%
Wherein, C represents the number of dead spots formed by the blank control; t represents the number of scorched spots formed after the treatment.
The above test results are shown in Table 1.
TABLE 1
Experiments show that compared with a single microbial agent, the mixed preparation provided by the invention has better effects on passivation, prevention and treatment of tobacco mosaic disease.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (5)
1. A microbial preparation for preventing and controlling tobacco mosaic virus is characterized in that: the tobacco leaf extract is a tobacco plant leaf extract which generates immune reaction after being sprayed with the plant immune inducer, and the mass percentages of the microorganism mixed microbial inoculum, the plant immune inducer and the leaf extract are 5-10%, 3-5% and 10-15% respectively, and the balance is sterile water; the mixed microbial inoculum comprises bacillus amyloliquefaciens and lactobacillus plantarum, wherein the mass ratio of the bacillus amyloliquefaciens to the lactobacillus plantarum is (2-3): (2-3); the plant immunity inducer is a tobacco mosaic virus capsid protein solution.
2. The microbial preparation for controlling tobacco mosaic virus according to claim 1, wherein: the cleaning agent also comprises a surfactant, wherein the surfactant is tween 80, organic silicon or sodium sulfosuccinate diester succinate.
3. The microbial preparation for controlling tobacco mosaic virus according to claim 2, wherein: the mass percent of the Tween 80 is 3%, or the mass percent of the organic silicon is 2%, or the mass percent of the sodium sulfosuccinate is 1.5%.
4. The method for preparing a microbial preparation for controlling tobacco mosaic virus according to claim 2 or 3, wherein the microbial preparation comprises the following steps: comprises the following steps of (a) carrying out,
s1, preparing seed solutions of various strains of the microbial mixed inoculant, fermenting and propagating the seed solutions respectively, and uniformly mixing propagation products according to a ratio;
s2 adding acetic acid into tobacco mosaic virus solution, mixing uniformly, standing, centrifuging to remove RNA precipitate, separating supernatant through desalting column, removing acetic acid, dialyzing the obtained solution in deionized water for 12-24 h, centrifuging, re-dissolving the obtained precipitate with phosphoric acid buffer solution to obtain tobacco mosaic virus capsid protein solution;
s3, spraying the tobacco mosaic virus capsid protein solution obtained in the step S2 on leaves of tobacco plants, picking the leaves of the plants after the tobacco plants generate immunoreaction, crushing the leaves, obtaining extraction mixed liquor by adopting a water extraction method, and then filtering solid impurities to obtain a leaf extracting solution;
s4, uniformly mixing the products obtained in the steps S1-S3 according to the proportion to obtain a mixed solvent;
s5, adding a surfactant into the mixed solvent in the step S4 according to the mixture ratio, and stirring for 1-2 hours.
5. The method for preparing a microbial preparation for controlling tobacco mosaic virus according to claim 4, wherein the microbial preparation comprises the following components in percentage by weight: in the step S3, after spraying the tobacco mosaic virus capsid protein solution on the tobacco plant lamina for 1 to 2 hours, extracting a lamina extract, then separating a single component from the lamina extract by using ion separation chromatography and high performance liquid chromatography, and obtaining the composition of the single component by using a mass spectrometry identification method to identify whether the tobacco plant produces an immunoreaction product.
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Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5832808A (en) * | 1981-08-24 | 1983-02-25 | Kureha Chem Ind Co Ltd | Controllong agent against plant viral disease and its preparation |
| CN1131439A (en) * | 1993-08-17 | 1996-09-18 | 莫根国际公司 | Chitinase DNA coding thereof and plants containing same |
| US7598346B1 (en) * | 2006-05-16 | 2009-10-06 | Pioneer Hi-Bred International, Inc. | Antifungal polypeptides |
| JP4472945B2 (en) * | 2003-04-14 | 2010-06-02 | 片倉チッカリン株式会社 | Microorganisms for controlling plant viruses, plant virus control agents comprising the microorganisms, and methods for controlling plant viruses using the microorganisms |
| CN102154164A (en) * | 2010-12-31 | 2011-08-17 | 山东农业大学 | Separation and application of antiviral bacillus amyloliquefaciens |
| WO2012079370A1 (en) * | 2010-12-17 | 2012-06-21 | 中国农业科学院植物保护研究所 | Verticillium dahliae protein elicitor gene, protein sequence and use thereof |
| CN103449892A (en) * | 2013-01-24 | 2013-12-18 | 杜新明 | Novel plant vaccine |
| RU2621860C1 (en) * | 2016-02-15 | 2017-06-07 | Федеральное государственное бюджетное учреждение науки Институт элементоорганических соединений им. А.Н. Несмеянова Российской академии наук (ИНЭОС РАН) | Inhibitor of tobacco mosaic virus and method of its production |
| CN107568249A (en) * | 2017-07-19 | 2018-01-12 | 北京百丰天下生物科技有限公司 | Bioprotective applied to prevention and cure of viruses and preparation method thereof |
| CN108076931A (en) * | 2018-01-05 | 2018-05-29 | 扬州大学 | It is a kind of to improve method of the plant to tobacco mosaic virus (TMV) systemic resistance |
| CN110402085A (en) * | 2016-12-06 | 2019-11-01 | 卵石实验室美国股份有限公司 | System and method for BIOLOGICAL CONTROL phytopathogen |
| CN110558340A (en) * | 2019-08-02 | 2019-12-13 | 武汉生物工程学院 | biological composite plant vaccine and preparation method thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110627881B (en) * | 2019-10-17 | 2022-12-27 | 中国烟草总公司陕西省公司 | Preparation and application of hypersensitive protein for biological prevention and treatment of tobacco mosaic virus disease |
-
2020
- 2020-06-30 CN CN202010612440.8A patent/CN111771911B/en active Active
Patent Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5832808A (en) * | 1981-08-24 | 1983-02-25 | Kureha Chem Ind Co Ltd | Controllong agent against plant viral disease and its preparation |
| CN1131439A (en) * | 1993-08-17 | 1996-09-18 | 莫根国际公司 | Chitinase DNA coding thereof and plants containing same |
| JP4472945B2 (en) * | 2003-04-14 | 2010-06-02 | 片倉チッカリン株式会社 | Microorganisms for controlling plant viruses, plant virus control agents comprising the microorganisms, and methods for controlling plant viruses using the microorganisms |
| US7598346B1 (en) * | 2006-05-16 | 2009-10-06 | Pioneer Hi-Bred International, Inc. | Antifungal polypeptides |
| WO2012079370A1 (en) * | 2010-12-17 | 2012-06-21 | 中国农业科学院植物保护研究所 | Verticillium dahliae protein elicitor gene, protein sequence and use thereof |
| CN102154164A (en) * | 2010-12-31 | 2011-08-17 | 山东农业大学 | Separation and application of antiviral bacillus amyloliquefaciens |
| CN103449892A (en) * | 2013-01-24 | 2013-12-18 | 杜新明 | Novel plant vaccine |
| RU2621860C1 (en) * | 2016-02-15 | 2017-06-07 | Федеральное государственное бюджетное учреждение науки Институт элементоорганических соединений им. А.Н. Несмеянова Российской академии наук (ИНЭОС РАН) | Inhibitor of tobacco mosaic virus and method of its production |
| CN110402085A (en) * | 2016-12-06 | 2019-11-01 | 卵石实验室美国股份有限公司 | System and method for BIOLOGICAL CONTROL phytopathogen |
| CN107568249A (en) * | 2017-07-19 | 2018-01-12 | 北京百丰天下生物科技有限公司 | Bioprotective applied to prevention and cure of viruses and preparation method thereof |
| CN108076931A (en) * | 2018-01-05 | 2018-05-29 | 扬州大学 | It is a kind of to improve method of the plant to tobacco mosaic virus (TMV) systemic resistance |
| CN110558340A (en) * | 2019-08-02 | 2019-12-13 | 武汉生物工程学院 | biological composite plant vaccine and preparation method thereof |
Non-Patent Citations (4)
| Title |
|---|
| A geminivirus betasatellite encoded beta C1 protein interacts with PsbP and subverts PsbP-mediated antiviral defence in plants;Gnanasekaran, Prabu et,al;《MOLECULAR PLANT PATHOLOGY》;20190731;第20卷(第7期);第943-960页 * |
| A polysaccharide found in Dendrobium nobile Lindl stimulates calcium signaling pathway and enhances tobacco defense against TMV;Li, Zhurui et,al;《INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES》;20190915;第137卷;第1286-1297页 * |
| 以烟草花叶病毒为半抗原载体的活体免疫反应;赵霞等;《应用化学》;20151031;第32卷(第10期);第1107-1113页 * |
| 植物抗病毒分子机制;钱礼超等;《中国科学:生命科学》;20141231;第44卷(第10期);第999-1099页 * |
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Denomination of invention: A microbial preparation for the prevention and control of tobacco mosaic virus and its preparation method Granted publication date: 20210604 Pledgee: Guanggu Branch of Wuhan Rural Commercial Bank Co.,Ltd. Pledgor: WUHAN HEYUAN GREEN ORGANISM Co.,Ltd. Registration number: Y2024980008736 |
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