CN113862189A - Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof - Google Patents
Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof Download PDFInfo
- Publication number
- CN113862189A CN113862189A CN202111214350.4A CN202111214350A CN113862189A CN 113862189 A CN113862189 A CN 113862189A CN 202111214350 A CN202111214350 A CN 202111214350A CN 113862189 A CN113862189 A CN 113862189A
- Authority
- CN
- China
- Prior art keywords
- c1tb
- culture medium
- c1gb
- preparation
- bacterial
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- 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
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/22—Bacillus
-
- 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
- A01N63/27—Pseudomonas
-
- 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
- A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
- A01N63/36—Penicillium
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
- C05G3/60—Biocides or preservatives, e.g. disinfectants, pesticides or herbicides; Pest repellants or attractants
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
- C05G3/80—Soil conditioners
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Abstract
The invention discloses a corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation and a preparation method thereof, wherein the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation comprises bacillus aryabhattai C1TB-10, bacillus pumilus C1TB-17, enterobacter cloacae C1TB-26, pseudomonas putida C1GB-15, lactobacillus plantarum C1GB-21 and penicillium microflavus C1 TF-23; and bacillus aryabhattai C1TB-10, bacillus pumilus C1TB-17, enterobacter cloacae C1TB-26, pseudomonas putida C1GB-15, lactobacillus plantarum C1GB-21 and penicillium microflavum C1TF-23 in a volume ratio of 1: 1: 1: 1: 1: 1, and mixing the components in a ratio of 1. The invention is compounded by a plurality of microbial strains, and achieves the purposes of promoting plant growth, improving soil environment and fertilizer efficiency, inhibiting the growth of soil-borne pathogenic bacteria and improving plant disease resistance by adding an exogenous plant growth-promoting bacterium, probiotic bacteria and biocontrol bacterium integrated microbial preparation.
Description
Technical Field
The invention belongs to the technical field of microbial preparations, and particularly relates to a corn growth-promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and a preparation method thereof.
Background
The Gansu province climate has the characteristics of southeast moistening, middle drought and northwest high cold, so that the planting quantity of middle and late maturing corn varieties is large, the growth period is long, and the environmental conditions and the phenological period are prolonged to create a good living environment for pest and disease damage; on the other hand, the corn in Hexi province is continuously planted for years, so that the disease residues and the pathogenic bacteria source base number in soil are increased in an accumulated mode, and soil-borne diseases are widely spread due to excessive application of nitrogen fertilizers and flood irrigation.
The corn diseases become the most main problems of yield reduction and income reduction of the corn planting industry in China, and the economic benefits of farmers are damaged. Chemical control can prevent and control the disease rate of corn diseases in a wide range within a very short time, but the chemical control has the problems of drug resistance of germs and insect pests, excessive pesticide residue, serious environmental pollution, threat to human health and the like. Biological control avoids a series of problems in plant protection, environment, energy, health and the like caused by the use of chemical pesticides.
Microbial preparations have a plant growth promoting effect and plant growth is improved by plant rhizosphere or symbiotic microorganisms producing plant hormones, such as: indoleacetic acid, gibberellin, cytokinin, ethylene, and the like; microorganisms produce vitamins such as: nicotinic acid, pantothenic acid, biotin, VB12 and the like, which are coenzymes or prosthetic groups of various enzymes in organisms; the growth process of the microorganism secretes nucleic acid substances, promotes the absorption of phosphorus and potassium by the roots of the plants, and enhances the photosynthesis of the plants. The microorganism converts inorganic trace elements which are not easy to be absorbed by the plant into organic trace elements which are easy to be absorbed, and the absorption of nutrition by the plant is promoted; some inducers secreted by the microorganisms improve the activity of soil enzymes, are beneficial to the transformation of soil nutrients and improve the absorption of plants to the nutrients.
The microbial preparation has biocontrol effect on plants, including antagonism, and biocontrol bacteria can be obtained by secreting antibacterial substances, such as: antibiotics, bacteriocins and other antibacterial proteins, etc., change the living environment, thereby inhibiting the growth of pathogenic bacteria; the method has the advantages that the method has a heavy parasitic effect, active substances secreted by biocontrol strains are identified by pathogenic bacteria, the biocontrol bacteria tightly wind pathogenic bacteria hyphae, and the cell walls of the pathogenic bacteria are degraded by secreting extracellular hydrolases, so that the hyphae are broken or disintegrated, and the growth of the pathogenic bacteria is hindered; competition effect, biocontrol bacteria are planted in the plant body and rapidly propagated, and a large amount of nutrition is consumed in the process, so that the pathogenic bacteria are lack of nutrition to inhibit growth; the induction of resistance and the excitation of biocontrol bacteria to the potential disease resistance of plants can play a role in preventing diseases.
The composite microbial preparation has better functionality than a single microbial preparation, 3 functional microorganisms including plant growth promoting microorganisms, probiotic microorganisms and biocontrol microorganisms are screened out and compounded with the functional microorganisms to prepare the preparation, and metabolites and various bioactive factors of the preparation are compounded, so that the function of the composite microbial preparation is diversified, and the stability of the preparation is enhanced.
Disclosure of Invention
The invention provides a corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and a preparation method thereof.
The technical scheme adopted by the invention is as follows:
the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation comprises bacillus aryabhattai C1TB-10, bacillus pumilus C1TB-17, enterobacter cloacae C1TB-26, pseudomonas putida C1GB-15, lactobacillus plantarum C1GB-21 and penicillium microflavus C1 TF-23; and bacillus aryabhattai C1TB-10, bacillus pumilus C1TB-17, enterobacter cloacae C1TB-26, pseudomonas putida C1GB-15, lactobacillus plantarum C1GB-21 and penicillium microflavum C1TF-23 in a volume ratio of 1: 1: 1: 1: 1: 1 in proportion;
the effective viable count is: bacillus aryabhattai 5.0X 108cfu/g, Bacillus pumilus 4.0 × 108cfu/g, Enterobacter cloacae 4.0X 108cfu/g, Pseudomonas putida 6.0X 108cfu/g, Lactobacillus plantarum 5.0 × 108cfu/g, Penicillium chrysogenum 6.0X 108cfu/g; wherein the content of the first and second substances,
bacillus aryabhattai (Bacillus aryabhattai) C1TB-10 is milk white in bacterial culture medium, convex in surface, smooth, opaque and neat in edge; the strain is a plant growth-promoting strain, produces plant hormone, is preserved in Gansu provincial industry microorganism strain preservation center in 2021, 9 months and 9 days, and has the preservation number: GSICC 32850;
the bacillus pumilus (Bacillus pumilus) C1TB-17 is milk white in colony on a bacterial culture medium, and has wrinkled surface and irregular edge; the thallus is rod-shaped, and two ends of the thallus are blunt and round; the strain is a plant probiotic, has the functions of passivating soil heavy metals and improving soil quality, is preserved in Gansu province industry microorganism strain preservation center in 2021, 9 months and 9 days, and has the preservation number: GSICC 32852;
the Enterobacter cloacae (Enterobacterercoloacae) C1TB-26 has small colony in a bacterial culture medium, is convex on the surface, is yellowish and opaque, is short and rod-shaped, and has no spore; the strain is a plant growth-promoting strain, has the functions of dissolving phosphorus and fixing nitrogen and improving the structure of a soil microbial flora, is stored in the Gansu province labor-saving industry microbial strain collection center in 2021, 9 months and 9 days, and has the preservation number: GSICC 30522;
the pseudomonas putida (Pseudomonas putida) C1GB-15 is white in bacterial colony in a bacterial culture medium, the surface is smooth, the edge is neat, and the thallus is in a short rod shape; the strain is a plant biocontrol strain, has the functions of inhibiting the growth of pathogenic bacteria and producing plant hormone, is stored in the Gansu province labor industry microorganism strain collection center at 9 months and 9 days in 2021, and has the following preservation numbers: GSICC 31637;
the bacterial colony of the lactobacillus plantarum C1GB-21 in a bacterial culture medium is milky white, round, smooth in surface and opaque; the strain is a plant probiotic, has the functions of generating various amino acids and vitamins, promoting plant growth and preventing and treating pathogenic bacteria, is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, 9 months and 9 days, and has the preservation number: GSICC 31252;
the bacterial colony of the penicillium microflavum (Penicillium miniatum) C1TF-23 in a fungus culture medium is changed from initial white to dark green, the hypha is in a compact velvet shape, the hypha is provided with septate hypha, the hypha is divided into a plurality of cells, the top does not form an expanded apical sac, the conidiophore chain is in a broom shape, and the conidia are in an oval shape; the strain is a plant biocontrol strain, has the function of inhibiting the growth of pathogenic bacteria, is preserved in the Gansu province industry microorganism strain preservation center at 9 months and 9 days in 2021, and has the preservation number: GSICC 61609.
The invention also provides a preparation method of the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation, which comprises the following steps:
the method comprises the following steps: preparing microorganism slant seeds: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 to a bacterial solid slant culture medium, and culturing at constant temperature of 30 ℃ for 48h to obtain a seed slant; inoculating lactobacillus plantarum C1GB-21 to an MRS solid slant culture medium, and culturing at a constant temperature of 37 ℃ for 48 hours to obtain a seed slant; inoculating Penicillium chrysogenum C1TF-23 to a fungus solid slant culture medium, and culturing at constant temperature of 30 ℃ for 48h to obtain a seed slant;
step two: culturing liquid seeds of the microorganisms: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a bacterial liquid culture medium, and performing shake culture at 30 ℃ and 200r/min for 30h to obtain liquid seeds; inoculating lactobacillus plantarum C1GB-21 in MRS liquid culture medium, and shake-culturing at 37 deg.C and 200r/min for 30h to obtain liquid seed; inoculating Penicillium chrysogenum C1TF-23 to a fungus liquid slant culture medium, and shake-culturing at 30 deg.C and 200r/min for 30h to obtain liquid seeds;
step three: liquid amplification culture of the strain: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a 50L fermentation tank, wherein the culture medium is a bacterial liquid culture medium, the inoculation amount is 10%, and the culture is carried out for 30h at the temperature of 30 ℃ and the rotating speed of 200r/min until the effective viable count in the Bacillus aryabhattai C1TB-10 bacterial liquid is 5 multiplied by 108The effective viable count of the bacterial liquid of CFU/mL and the Bacillus pumilus C1TB-17 is 4 multiplied by 108CFU/mL, the effective viable count of the bacterial liquid of the enterobacter cloacae C1TB-26 is 5 multiplied by 108CFU/mL, the effective viable count of the pseudomonas putida C1GB-15 bacterial liquid is 6 multiplied by 108CFU/mL；
Inoculating Lactobacillus plantarum C1GB-21 in 50L fermenter, wherein the culture medium is MRS liquid culture medium with an inoculum size of 10%, and culturing at 37 deg.C and 200r/min for 30h until the effective viable count of Lactobacillus plantarum C1GB-21 bacterial liquid is 4 × 108CFU/mL；
Inoculating Penicillium microflavum C1TF-23 into 50L fermentation tank, wherein the culture medium is fungus liquid culture medium with an inoculum size of 10%, and culturing at 30 deg.C and 200r/min for 36h until the number of effective active spores in the bacterial liquid is 6 × 108CFU/mL；
Step four: preparation of microbial preparation: respectively carrying out amplification culture on the microbial agents in the third step according to the volume ratio of 1: 1: 1: 1: 1: 1, uniformly mixing to obtain the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation;
step five: preparing a microbial fertilizer: and D, uniformly mixing the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation obtained in the step four with an organic fertilizer according to the volume ratio of 1:20 to obtain the corn growth promoting, probiotic and biocontrol microbial fertilizer.
Wherein, the bacteria solid slant culture medium in the first step, the second step and the third step is as follows: 10g of tryptone, 5g of yeast extract, 10g of sodium chloride, 20g of agar and 1000mL of distilled water, wherein the pH value is 7.0; sterilizing with high pressure steam at 121 deg.C for 20 min;
the MRS solid slant culture medium comprises: 10g of casein peptone, 10g of beef extract, 5g of yeast powder, 5g of glucose, 5g of sodium acetate, 2g of diammonium citrate, 801g of Tween, 2g of dipotassium phosphate, 0.2g of magnesium sulfate heptahydrate, 0.05g of manganese sulfate, 20g of calcium carbonate, 20g of agar, 1000mL of distilled water and pH of 6.8; sterilizing with high pressure steam at 121 deg.C for 20 min;
the fungus solid slant culture medium comprises: 200g of potato, 20g of glucose, 20g of agar and 1000ml of distilled water, and the pH value is natural; cleaning and peeling potato, weighing 200g, cutting into small pieces, adding water, boiling for 20 minutes, filtering with eight layers of gauze, heating, adding agar, continuously heating, stirring uniformly, adding glucose after the agar is dissolved, stirring uniformly, slightly cooling, supplementing water to 1000ml, and sterilizing at 121 ℃ for 20 minutes at high pressure for later use.
Wherein, the liquid culture medium in the second step and the third step is the same as the bacterial culture medium, the MRS culture medium and the fungus culture medium in the first step, and the difference is that agar is not added.
The invention adopts a compound microbial preparation, wherein bacillus aryabhattai can synthesize a siderophore, generate phytohormones such as indoleacetic acid (IAA) and the like, and has broad spectrum on the growth promotion effect of plants; can also protect plants, prevent the toxic action of heavy metal and nitrite on the plants and promote the absorption of phosphorus and potassium fertilizer. The bacillus pumilus has the function of passivating soil heavy metals, and can repair soil pollution and promote plant growth; meanwhile, various antibacterial substances are secreted to inhibit the growth of various soil-borne pathogenic fungi. The enterobacter cloacae has the functions of dissolving phosphorus and fixing nitrogen, improves the diversity of soil bacteria flora, and improves the activity and the fertilizer efficiency of soil enzyme; meanwhile, the composition has a biological control effect on plant diseases. The pseudomonas putida fully utilizes root exudates and nutrients in rhizosphere soil through stronger planting capacity, reduces nutrient substances necessary for the growth of pathogenic bacteria, and thus inhibits the growth of the pathogenic bacteria; can also produce antibiotics, siderophores and other growth promoting substances to promote plant growth. The lactobacillus plantarum generates various amino acids and vitamins and promotes the nutrient absorption capacity of plant roots; produce a large amount of lactic acid and bacteriocin, effectively inhibit the growth of harmful bacteria, and have good probiotic function. The penicillium microflavum can inhibit the growth of pathogenic bacteria by secreting metabolites, and by aspects of nutrition competition, parasitism, antibiotic action and the like, and can also produce phytohormones such as indoleacetic acid and the like. The strains have no mutual antagonism and mutual inhibition, different strains have different functions, the strains are compounded to form a compound microbial preparation, the compound microbial preparation is applied to soil in an externally added manner, and a great amount of dominant floras are formed through propagation, so that the effects of promoting the growth of plants, benefiting life and preventing life are achieved.
The invention has the following advantages:
(1) the microbial preparation is compounded by 6 microorganisms with 3 functions of growth promotion, benefit and biological control, no inhibition effect exists among strains, 6 functional microbial strains have synergistic effect, and the functions of growth promotion, benefit and biological control are exerted together;
(2) the growth-promoting strain in the microbial preparation not only secretes a large amount of plant hormone and promotes the growth of plants, but also has the functions of dissolving phosphorus and fixing nitrogen, improving the soil fertilizer efficiency and improving the structure of soil microbial flora;
(3) the probiotic bacterial strain in the microbial preparation can secrete plant hormone, generate various amino acids, vitamins and other substances beneficial to plant growth, and has the function of inhibiting the growth of pathogenic bacteria;
(4) the biocontrol strain in the microbial preparation has good effects of inhibiting the growth of pathogenic bacteria and preventing and treating plant diseases, and can secrete phytohormone to promote healthy growth of plants;
(5) the microbial preparation and the organic fertilizer are uniformly mixed and applied, so that the recycling of agricultural and pastoral wastes is realized, the pollution of livestock and poultry manure is reduced, the soil fertilizer efficiency is improved, and the growth of plants is facilitated;
(6) after the microbial preparation is applied, the effects of plant growth promotion, probiotics and biocontrol are simultaneously exerted, the production process is simple, and the industrial production is convenient to realize.
Detailed Description
The present invention is described in terms of particular embodiments, other advantages and features of the invention will become apparent to those skilled in the art from the following disclosure, and it is to be understood that the described embodiments are merely exemplary of the invention and that it is not intended to limit the invention to the particular embodiments disclosed. All modifications directly derivable or suggested based on the present disclosure are within the scope of the invention.
Example 1
Screening of functional strains
(1) Separation and purification of bacterial strains
The microbial strain is screened and separated from the rhizosphere soil of the root of healthy corn in the Gansu Hexi corn seed production base, and specifically comprises the following steps: adding 10g of healthy angelica rhizosphere soil sample into a triangular flask filled with 100mL of sterile water, shaking for 30min on a shaking table at 200rpm, adding 1mL of uniformly mixed liquid soil sample into a new test tube filled with 9mL of sterile water, shaking for 5min in a shaker, adding 1mL of liquid soil sample into a new test tube filled with 9mL of sterile water, and diluting the soil sample to 10-8. Sequentially selecting soil samples with concentrations of 10-4, 10-5, 10-6 and 10-7, respectively sucking 100 μ L of the soil samples, coating the soil samples on a bacteria solid culture medium, an MRS solid culture medium and a fungus solid culture medium, culturing at a constant temperature of 30 ℃, separating and purifying strains by a plate marking method after the bacteria grow out, preparing the strains into inclined planes, and storing at 4 ℃ for later use.
The bacterial solid slant culture medium comprises: 10g of tryptone, 5g of yeast extract, 10g of sodium chloride, 20g of agar and 1000mL of distilled water, wherein the pH value is 7.0. Sterilizing with high pressure steam at 121 deg.C for 20 min.
The MRS solid slant culture medium comprises: 10g of casein peptone, 10g of beef extract, 5g of yeast powder, 5g of glucose, 5g of sodium acetate, 2g of diammonium citrate, 801g of Tween, 2g of dipotassium phosphate, 0.2g of magnesium sulfate heptahydrate, 0.05g of manganese sulfate, 20g of calcium carbonate, 20g of agar, 1000mL of distilled water and 6.8 of pH. Sterilizing with high pressure steam at 121 deg.C for 20 min.
The fungus solid slant culture medium comprises: 200g of potato, 20g of glucose, 20g of agar and 1000ml of distilled water, and the pH is natural. Cleaning and peeling potato, weighing 200g, cutting into small pieces, adding water, boiling for 20 minutes, filtering with eight layers of gauze, heating, adding agar, continuously heating, stirring uniformly, adding glucose after the agar is dissolved, stirring uniformly, slightly cooling, supplementing water to 1000ml, and sterilizing at 121 ℃ for 20 minutes at high pressure for later use.
(2) Determination of phosphate solubilizing ability of Strain
Respectively inoculating the strains to a PKO inorganic phosphorus culture medium, culturing for 48 hours at 28 ℃ and 180r/min, determining the phosphorus dissolving capacity by adopting a molybdenum-antimony anti-chromogenic method, centrifuging a strain culture solution for 15 minutes at 10000r/min, absorbing 2-10 mL of supernatant into a 50mL volumetric flask, diluting to 20mL by using water, adding 2 drops of 2, 6-dinitrophenol indicator, adjusting the pH value of the solution to be just yellowish by using 10% sodium hydroxide or dilute sulfuric acid solution, adding 5mL of molybdenum-antimony anti-chromogenic agent, shaking up, and fixing the volume to scale. Standing at room temperature of 20-25 deg.C for 30min, and determining OD690 value.
The PKO inorganic phosphorus culture medium comprises: (NH)4)2SO40.1g, 10.0g of sucrose, 0.5g of NaCl, 0.2g of KCl and MgSO40.1g，FeSO40.03g，MnSO40.03g，Ca3(PO4)23.0g, 0.5g of yeast extract powder and 1000mL of distilled water;
(3) determination of nitrogen fixation ability of strain
By adopting a streak inoculation method, strains are selected to be streaked on an Ashby solid culture medium, the strains are cultured for 10 days at the temperature of 30 ℃, the strains are continuously transferred for 3 times, and the 3 rd time bacteria which can still grow on the Ashby solid culture medium can be regarded as bacteria with nitrogen fixing capacity.
The Ashby culture medium is as follows: KH (Perkin Elmer)2PO40.2g，MgSO4·7H2O 0.2g，NaCl 0.2g,CaCO35.0g, mannitol 10.0g, CaSO4·2H20.1g of O, 1000mL of distilled water, and pH 6.8-7.0.
(4) Quantitative determination of indole acetic acid secretion of strain
Drawing of IAA standard curve
100. mu.g/mL of IAA mother liquor was prepared, and 5. mu.g/mL, 10. mu.g/mL, 15. mu.g/mL, 20. mu.g/mL and 25. mu.g/mL of IAA mother liquor were prepared by gradient dilution with distilled water, and the experiment was repeated for 3 groups of IAA mother liquor. Measuring by a Salkowski method, sucking 1mL of the solution to be measured, dripping 1mL of Salkowski colorimetric solution, developing for 30min in a dark condition, rapidly carrying out colorimetry (530nm) by using a spectrophotometer, and drawing a standard curve according to statistical data.
Quantitative determination of IAA secretion of strain
Respectively inoculating the strains into a bacterial culture solution containing L-tryptophan, culturing at 28 ℃ and 180r/min for 48h, centrifuging at 10000r/min for 10min, sucking 1mL of centrifugal supernatant, adding 4mL of Salkowski colorimetric solution, developing under dark conditions for 30min, and quickly carrying out color comparison (530nm) by using a spectrophotometer. Three sets of data were measured and the results averaged to find the corresponding amount of IAA produced on the standard curve.
Salkowski colorimetric solution: 50mL 35% HCLO4+1mL0.5mol/LFeCL3。
(5) Quantitative determination of gibberellin secretion of strains
Drawing of standard curve
Respectively preparing standard gibberellin solutions with the concentrations of 0, 0.1, 0.2, 0.4, 0.8, 1.2, 1.6, 2.0, 2.4 and 3.0 mu g/mL, dissolving 5mL of the standard solution in a proper amount of 70% (volume fraction) ethanol, adding 0.5mL of the standard solution into 4.5mL of 98% (volume fraction) concentrated sulfuric acid, uniformly mixing, fixing the volume to 25mL, and determining the light absorption value of a reaction product at 412 nm. And drawing a corresponding standard curve.
Quantitative determination of gibberellin secretion of strains
Respectively inoculating the strains into a bacterial culture solution, culturing at 28 ℃ and 180r/min for 48h, centrifuging at 10000r/min for 10min, dissolving 5mL of fermentation liquor in a proper amount of 70% ethanol, adding 0.5mL of 98% concentrated sulfuric acid into 0.5mL of fermentation liquor, uniformly mixing, metering to 25mL, and measuring the absorbance at 412 nm. Three sets of data were measured and the results averaged to find the corresponding amount of gibberellin production on the standard curve.
(6) Pathogenic bacterium antagonistic strain screening
And (3) screening pathogenic bacteria: corn stalk rot and corn blight disease strains are collected from a corn breeding base in Hexi, Gansu province, pathogenic microorganisms are respectively separated and screened from the corn disease strains through fungus plate separation and screening, and the pathogenic microorganisms are respectively fusarium graminearum (Fusarium graminearum) and rhizoctonia solani (Rhizoctonia solani) through the identification of fungus ITS specific regions.
Bacteriostatic experiments: preparing a nutrient agar solid culture medium, taking 4mm fungus cakes of pathogenic microorganisms including fusarium graminearum and rhizoctonia solani, placing the fungus cakes at the central part of a culture dish, culturing at the constant temperature of 30 ℃ for 24 hours, then respectively taking each strain fungus cake, placing the fungus cakes in 4 directions of about 25mm from pathogenic bacteria, repeating the treatment for 3 times, and simultaneously setting a blank control group; and (3) performing constant-temperature culture at 30 ℃, observing the bacteriostatic effect when the blank control is about to grow over the whole culture dish, and determining the bacteriostatic rate. The bacteriostatic rate (%) (control growth radius-treatment growth radius)/control growth radius × 100%.
The nutrient agar solid culture medium comprises: 10g of peptone, 3g of beef extract, 5g of sodium chloride, 20g of agar and 1000mL of distilled water, and the pH value is 7.0. Autoclaving at 121 deg.C for 20min, and pouring into a culture dish.
(7) Identification of functional strains
1) Bacterial strain identification
Extracting genome DNA of each strain by using a kit method (OMEGA USA), selecting bacterial 16SrDNA universal primers 27F:5'-AGAGTTTGATCCTGGCTCAG-3' and 1492R:5'-GGTTACCTTGTTACGACTT-3' for amplification, sending amplified products to Shanghai worker for sequencing, and comparing and analyzing obtained DNA sequences with sequences in a GenBank database.
2) Identification of fungal strains
Extracting genome DNA of each strain by using a kit method (OMEGA USA), and selecting a fungus ITS universal primer ITS 1: 5'-TCCGTAGGTGAACCTGCGG-3', ITS 4: 5'-TCCTCCGCTTATTGATATGC-3' amplifying, sending the amplified product to Shanghai work sequencing, and comparing the obtained DNA sequence with the sequence in GenBank database for analysis.
(8) Results
The identified C1TB-10 is Bacillus aryabhattai, has the best capability of producing indoleacetic acid and gibberellin, and simultaneously has the capability of fixing nitrogen and dissolving phosphorus. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 32850;
c1TB-17 has the best phosphate solubilizing capability for Bacillus pumilus, can produce indoleacetic acid and gibberellin, and has certain inhibition effect on pathogenic bacteria. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 32852;
c1TB-26 is enterobacter cloacae which has the best gibberellin and indoleacetic acid producing capability and simultaneously has nitrogen fixing and phosphorus dissolving capabilities. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 30522;
c1GB-15 has good function of inhibiting the growth of pathogenic bacteria for pseudomonas putida, and can produce indoleacetic acid. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 31637;
the C1GB-21 is lactobacillus plantarum, has a good function of producing indoleacetic acid, and has a certain effect of inhibiting the growth of pathogenic bacteria. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 31252;
c1TF-23 has the best bacteriostatic effect on Penicillium chrysogenum. The strain is preserved in the Gansu province labor industry microorganism strain preservation center in 2021, and the preservation number is as follows: GSICC 61609;
TABLE 1 measurement results of functionality of microorganism strains
Example 2
The preparation method of the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation comprises the following steps:
the method comprises the following steps: preparing microorganism slant seeds: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 to a bacterial solid slant culture medium, and culturing at constant temperature of 30 ℃ for 48h to obtain a seed slant; inoculating lactobacillus plantarum C1GB-21 to an MRS solid slant culture medium, and culturing at a constant temperature of 37 ℃ for 48 hours to obtain a seed slant; the micro-yellow penicillium C1TF-23 is planted in a fungus solid slant culture medium and cultured for 48 hours at the constant temperature of 30 ℃, thus obtaining the seed slant.
The bacterial solid slant culture medium comprises: 10g of tryptone, 5g of yeast extract, 10g of sodium chloride, 20g of agar and 1000mL of distilled water, wherein the pH value is 7.0. Sterilizing with high pressure steam at 121 deg.C for 20 min.
The MRS solid slant culture medium comprises: 10g of casein peptone, 10g of beef extract, 5g of yeast powder, 5g of glucose, 5g of sodium acetate, 2g of diammonium citrate, 801g of Tween, 2g of dipotassium phosphate, 0.2g of magnesium sulfate heptahydrate, 0.05g of manganese sulfate, 20g of calcium carbonate, 20g of agar, 1000mL of distilled water and 6.8 of pH. Sterilizing with high pressure steam at 121 deg.C for 20 min.
The fungus solid slant culture medium comprises: 200g of potato, 20g of glucose, 20g of agar and 1000ml of distilled water, and the pH is natural. Cleaning and peeling potato, weighing 200g, cutting into small pieces, adding water, boiling for 20 minutes, filtering with eight layers of gauze, heating, adding agar according to actual needs, continuously heating, stirring uniformly, adding glucose after the agar is dissolved, stirring uniformly, slightly cooling, supplementing water to 1000ml, and sterilizing at 121 ℃ under high pressure for 20 minutes for later use.
Step two: culturing liquid seeds of the microorganisms: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a bacterial liquid culture medium, and performing shake culture at 30 ℃ and 200r/min for 30h to obtain liquid seeds; inoculating lactobacillus plantarum C1GB-21 in MRS liquid culture medium, and shake-culturing at 37 deg.C and 200r/min for 30h to obtain liquid seed; the method comprises the steps of planting the Penicillium chrysogenum C1TF-23 in a fungus liquid slant culture medium, and carrying out shake culture at the temperature of 30 ℃ and the rotation speed of 200r/min for 30 hours to obtain liquid seeds.
The liquid culture medium is the same as the bacteria culture medium, the MRS culture medium and the fungus culture medium in the step one, and the difference is that agar is not added.
Step three: liquid amplification culture of the strain: respectively inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a 50L fermentation tank, wherein the culture medium is a bacterial liquid culture medium, the inoculation amount is 10%, and the culture is carried out for 30h at the temperature of 30 ℃ and the rotating speed of 200r/min until the effective viable count in the Bacillus aryabhattai C1TB-10 bacterial liquid is 5 multiplied by 108The effective viable count of the bacterial liquid of CFU/mL and the Bacillus pumilus C1TB-17 is 4 multiplied by 108CFU/mL, the effective viable count of the bacterial liquid of the enterobacter cloacae C1TB-26 is 5 multiplied by 108CFU/mL, the effective viable count of the pseudomonas putida C1GB-15 bacterial liquid is 6 multiplied by 108CFU/mL. Inoculating Lactobacillus plantarum C1GB-21 in 50L fermenter, wherein the culture medium is MRS liquid culture medium with an inoculum size of 10%, and culturing at 37 deg.C and 200r/min for 30h until the effective viable count of Lactobacillus plantarum C1GB-21 bacterial liquid is 4 × 108CFU/mL。
Inoculating Penicillium microflavum C1TF-23 into 50L fermentation tank, wherein the culture medium is fungus liquid culture medium with an inoculum size of 10%, and culturing at 30 deg.C and 200r/min for 36h until the number of effective active spores in the bacterial liquid is 6 × 108CFU/mL。
And the bacterial liquid culture medium, the MRS liquid culture medium and the fungus liquid culture medium are the same as the step two.
Step four: preparation of microbial preparation: respectively carrying out amplification culture on the microbial agents in the third step according to the volume ratio of 1: 1: 1: 1: 1: 1, mixing uniformly to obtain the integrated preparation of the corn growth promoting, probiotic and biocontrol bacterial manure.
Step five: preparing a microbial fertilizer: and D, uniformly mixing the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation obtained in the step four with an organic fertilizer according to the volume ratio of 1:20 to obtain the corn growth promoting, probiotic and biocontrol microbial fertilizer.
Example 3
Corn growth promoting, probiotic and biocontrol bacterial fertilizerThe difference from example 2 in the preparation method of the transformant preparation is that the strain liquid amplification culture: respectively inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a 50L fermentation tank, wherein the culture medium is a bacterial liquid culture medium, the inoculation amount is 10%, and the culture is carried out for 30h at the temperature of 30 ℃ and the rotating speed of 200r/min until the effective viable count in the Bacillus aryabhattai C1TB-10 bacterial liquid is 4 multiplied by 108CFU/mL, the effective viable count of the bacillus pumilus C1TB-17 bacterial liquid is 6 multiplied by 108CFU/mL, the effective viable count of the enterobacter cloacae C1TB-26 bacterial liquid is 4 multiplied by 108The effective viable count of the bacterial liquid of the CFU/mL and the pseudomonas putida C1GB-15 is 5 multiplied by 108CFU/mL. Inoculating Lactobacillus plantarum C1GB-21 in 50L fermenter, wherein the culture medium is MRS liquid culture medium with an inoculum size of 10%, and culturing at 37 deg.C and 200r/min for 30h until the effective viable count of Lactobacillus plantarum C1GB-21 bacterial liquid is 6 × 108CFU/mL. Inoculating Penicillium microflavum C1TF-23 into 50L fermentation tank, wherein the culture medium is fungus liquid culture medium with an inoculum size of 10%, and culturing at 30 deg.C and 200r/min for 36h until the number of effective active spores in the bacterial liquid is 5 × 108CFU/mL. Respectively carrying out amplification culture on the microbial agents according to the volume ratio of 1: 1: 1: 1: 1: 1, uniformly mixing to obtain a corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation, and uniformly mixing the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and an organic fertilizer according to the volume ratio of 1:20 to obtain the corn growth promoting, probiotic and biocontrol microbial fertilizer.
Example 4
The preparation method of the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation is different from the preparation method of the example 2 in that the strain liquid is subjected to amplification culture: respectively inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a 50L fermentation tank, wherein the culture medium is a bacterial liquid culture medium, the inoculation amount is 10%, and the culture is carried out for 30h at the temperature of 30 ℃ and the rotating speed of 200r/min until the effective viable count in the Bacillus aryabhattai C1TB-10 bacterial liquid is 5 multiplied by 108CFU/mL, the effective viable count of the bacillus pumilus C1TB-17 bacterial liquid is 6 multiplied by 108CFU/mL, Enterobacter cloacae CThe effective viable count of the 1TB-26 bacterial liquid is 5 multiplied by 108CFU/mL, the effective viable count of the pseudomonas putida C1GB-15 bacterial liquid is 4 multiplied by 108CFU/mL. Inoculating Lactobacillus plantarum C1GB-21 in 50L fermenter, wherein the culture medium is MRS liquid culture medium with an inoculum size of 10%, and culturing at 37 deg.C and 200r/min for 30h until the effective viable count of Lactobacillus plantarum C1GB-21 bacterial liquid is 6 × 108CFU/mL. Inoculating Penicillium microflavum C1TF-23 into 50L fermentation tank, wherein the culture medium is fungus liquid culture medium with an inoculum size of 10%, and culturing at 30 deg.C and 200r/min for 36h until the number of effective active spores in the bacterial liquid is 4 × 108CFU/mL. Respectively carrying out amplification culture on the microbial agents according to the volume ratio of 1: 1: 1: 1: 1: 1, uniformly mixing to obtain a corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation, and uniformly mixing the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and an organic fertilizer according to the volume ratio of 1:20 to obtain the corn growth promoting, probiotic and biocontrol microbial fertilizer.
Example 5
The field application effect is as follows:
the microbial fertilizer prepared in the example 2, the example 3 and the example 4 is applied to a corn seed production base in Hexi area of Gansu, and meanwhile, a blank control group without adding microbial inoculum is arranged, each treatment is repeated for 3 times, and the test adopts a completely random block design. And (5) counting the corn yield, the stalk rot and the blight morbidity.
And (3) test results: compared with a control group, after the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation is applied in the examples 2, 3 and 4, the incidence rates of common diseases, namely stem rot and blight of corn are reduced to different degrees, and the yield of the corn for seed production is obviously improved. The corn growth promoting, benefiting and biocontrol bacterial manure integrated preparation can relieve the morbidity of corn stalk rot and withered grain disease, improve the ecological environment of corn roots, promote the corn growth and improve the yield of corn seed production.
TABLE 2 Effect of microbial preparations on corn yield and disease control
It is noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention.
Claims (4)
1. The corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation is characterized in that: comprises Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26, Pseudomonas putida C1GB-15, Lactobacillus plantarum C1GB-21 and Penicillium chrysogenum C1 TF-23; and bacillus aryabhattai C1TB-10, bacillus pumilus C1TB-17, enterobacter cloacae C1TB-26, pseudomonas putida C1GB-15, lactobacillus plantarum C1GB-21 and penicillium microflavum C1TF-23 in a volume ratio of 1: 1: 1: 1: 1: 1 in proportion;
the effective viable count is: bacillus aryabhattai 5.0X 108cfu/g, Bacillus pumilus 4.0 × 108cfu/g, Enterobacter cloacae 4.0X 108cfu/g, Pseudomonas putida 6.0X 108cfu/g, Lactobacillus plantarum 5.0 × 108cfu/g, Penicillium chrysogenum 6.0X 108cfu/g; wherein the content of the first and second substances,
bacillus aryabhattai (Bacillusaryabyabhattai) C1TB-10, which is preserved in the Gansu province labor industry microorganism strain collection center at 9 months and 9 days in 2021 with the preservation number: GSICC 32850;
the bacillus pumilus (Bacillus pumilus) C1TB-17 is preserved in the Gansu province labor industry microorganism strain collection center at 9 months and 9 days in 2021, and the preservation number is as follows: GSICC 32852;
the enterobacter cloacae (Enterobactererccloacae) C1TB-26 is preserved in the culture collection of the strain of the labor saving industry in Gansu at 9 months and 9 days in 2021, and the preservation number is as follows: GSICC 30522;
the pseudomonas putida (pseudomonas putida) C1GB-15 is preserved in the Gansu province industry microorganism strain collection center at 9 months and 9 days in 2021 with the preservation number: GSICC 31637;
the lactobacillus plantarum C1GB-21 is preserved in the Gansu province labor-saving microorganism strain collection center in 2021, 9 months and 9 days, and the preservation number is as follows: GSICC 31252;
the penicillium microflavum (Penicillium miniatum) C1TF-23 is preserved in the Gansu province labor-saving microorganism strain collection center at 9 months and 9 days in 2021 with the preservation number: GSICC 61609.
2. The preparation method of the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation according to claim 1, characterized in that: the method comprises the following steps:
the method comprises the following steps: preparing microorganism slant seeds: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 to a bacterial solid slant culture medium, and culturing at constant temperature of 30 ℃ for 48h to obtain a seed slant; inoculating lactobacillus plantarum C1GB-21 to an MRS solid slant culture medium, and culturing at a constant temperature of 37 ℃ for 48 hours to obtain a seed slant; inoculating Penicillium chrysogenum C1TF-23 to a fungus solid slant culture medium, and culturing at constant temperature of 30 ℃ for 48h to obtain a seed slant;
step two: culturing liquid seeds of the microorganisms: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a bacterial liquid culture medium, and performing shake culture at 30 ℃ and 200r/min for 30h to obtain liquid seeds; inoculating lactobacillus plantarum C1GB-21 in MRS liquid culture medium, and shake-culturing at 37 deg.C and 200r/min for 30h to obtain liquid seed; inoculating Penicillium chrysogenum C1TF-23 to a fungus liquid slant culture medium, and shake-culturing at 30 deg.C and 200r/min for 30h to obtain liquid seeds;
step three: liquid amplification culture of the strain: inoculating Bacillus aryabhattai C1TB-10, Bacillus pumilus C1TB-17, Enterobacter cloacae C1TB-26 and Pseudomonas putida C1GB-15 into a 50L fermentation tank, wherein the culture medium is a bacterial liquid culture medium, the inoculation amount is 10%, and the culture is carried out for 30h at the temperature of 30 ℃ and the rotating speed of 200r/min until the effective viable count in the Bacillus aryabhattai C1TB-10 bacterial liquid is 5 multiplied by 108The effective viable count of the bacterial liquid of CFU/mL and the Bacillus pumilus C1TB-17 is 4 multiplied by 108CFU/mL, the effective viable count of the bacterial liquid of the enterobacter cloacae C1TB-26 is 5 multiplied by 108CFU/mL, Pseudomonas putidaThe effective viable count of the C1GB-15 bacterial liquid is 6 multiplied by 108CFU/mL；
Inoculating Lactobacillus plantarum C1GB-21 in 50L fermenter, wherein the culture medium is MRS liquid culture medium with an inoculum size of 10%, and culturing at 37 deg.C and 200r/min for 30h until the effective viable count of Lactobacillus plantarum C1GB-21 bacterial liquid is 4 × 108CFU/mL；
Inoculating Penicillium microflavum C1TF-23 into 50L fermentation tank, wherein the culture medium is fungus liquid culture medium with an inoculum size of 10%, and culturing at 30 deg.C and 200r/min for 36h until the number of effective active spores in the bacterial liquid is 6 × 108CFU/mL；
Step four: preparation of microbial preparation: respectively carrying out amplification culture on the microbial agents in the third step according to the volume ratio of 1: 1: 1: 1: 1: 1, uniformly mixing to obtain the corn growth promoting, probiotic and biocontrol bacterial manure integrated preparation;
step five: preparing a microbial fertilizer: and D, uniformly mixing the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation obtained in the step four with an organic fertilizer according to the volume ratio of 1:20 to obtain the corn growth promoting, probiotic and biocontrol microbial fertilizer.
3. The preparation method of the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation as claimed in claim 2, wherein the preparation method comprises the following steps: the bacterial solid slant culture medium in the first step is as follows: 10g of tryptone, 5g of yeast extract, 10g of sodium chloride, 20g of agar and 1000mL of distilled water, wherein the pH value is 7.0; sterilizing with high pressure steam at 121 deg.C for 20 min;
the MRS solid slant culture medium comprises: 10g of casein peptone, 10g of beef extract, 5g of yeast powder, 5g of glucose, 5g of sodium acetate, 2g of diammonium citrate, 801g of Tween, 2g of dipotassium phosphate, 0.2g of magnesium sulfate heptahydrate, 0.05g of manganese sulfate, 20g of calcium carbonate, 20g of agar, 1000mL of distilled water and pH of 6.8; sterilizing with high pressure steam at 121 deg.C for 20 min;
the fungus solid slant culture medium comprises: 200g of potato, 20g of glucose, 20g of agar and 1000ml of distilled water, and the pH value is natural; cleaning and peeling potato, weighing 200g, cutting into small pieces, adding water, boiling for 20 minutes, filtering with eight layers of gauze, heating, adding agar, continuously heating, stirring uniformly, adding glucose after the agar is dissolved, stirring uniformly, slightly cooling, supplementing water to 1000ml, and sterilizing at 121 ℃ for 20 minutes at high pressure for later use.
4. The preparation method of the corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation according to claim 1, characterized in that: the liquid culture medium in the second step is the same as that in the third step, and is the same as the bacteria culture medium, MRS culture medium and fungus culture medium in the first step, except that agar is not added.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111214350.4A CN113862189A (en) | 2021-10-19 | 2021-10-19 | Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111214350.4A CN113862189A (en) | 2021-10-19 | 2021-10-19 | Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113862189A true CN113862189A (en) | 2021-12-31 |
Family
ID=79000452
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111214350.4A Pending CN113862189A (en) | 2021-10-19 | 2021-10-19 | Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113862189A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114196601A (en) * | 2022-01-14 | 2022-03-18 | 甘肃省科学院生物研究所 | Microbial agent capable of promoting growth, biocontrol and degrading organophosphorus pesticide and preparation method thereof |
| CN114591867A (en) * | 2022-03-29 | 2022-06-07 | 甘肃省科学院生物研究所 | Composite microbial preparation for enhancing corn growth effect and preparation method and application thereof |
| CN114946883A (en) * | 2022-03-21 | 2022-08-30 | 云南师范大学 | Cucurbitacin capable of enhancing disease resistance of plants and application thereof |
| CN114982589A (en) * | 2022-04-21 | 2022-09-02 | 甘肃省科学院生物研究所 | Corn seeding time probiotic composite matrix and preparation method, matrix cup and application thereof |
| CN115305220A (en) * | 2022-07-26 | 2022-11-08 | 中国农业大学 | Pseudomonas capable of fixing nitrogen and promoting plant growth and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106396864A (en) * | 2016-08-29 | 2017-02-15 | 佛山市艳晖生物科技有限公司 | Microbial fertilizer for control of potato scab and preparation method thereof |
| CN107778035A (en) * | 2017-11-24 | 2018-03-09 | 山西省农业科学院生物技术研究中心 | A kind of growth-promoting volume increase microbial organic fertilizer and preparation method thereof |
| CN108865955A (en) * | 2018-07-30 | 2018-11-23 | 中山市承铭农业技术开发有限公司 | A kind of complex micro organism fungicide and preparation method thereof |
-
2021
- 2021-10-19 CN CN202111214350.4A patent/CN113862189A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106396864A (en) * | 2016-08-29 | 2017-02-15 | 佛山市艳晖生物科技有限公司 | Microbial fertilizer for control of potato scab and preparation method thereof |
| CN107778035A (en) * | 2017-11-24 | 2018-03-09 | 山西省农业科学院生物技术研究中心 | A kind of growth-promoting volume increase microbial organic fertilizer and preparation method thereof |
| CN108865955A (en) * | 2018-07-30 | 2018-11-23 | 中山市承铭农业技术开发有限公司 | A kind of complex micro organism fungicide and preparation method thereof |
Non-Patent Citations (3)
| Title |
|---|
| LOVET T. KIGIGHA ET AL.: "Microbial Characterization of Maize Fermentation Water during Ogi Production", 《JOURNAL OF ENVIRONNEMENTAL TREATMENT TECHNIQUES》, pages 41 - 45 * |
| 李兆周 等: "黑曲霉高产纤维素酶菌株的高通量筛选", 《甘肃农业大学学报》, pages 151 - 156 * |
| 祝英 等: "有机肥替代部分化肥对土壤肥力和微生物特征的影响", 《土壤通报》, pages 1161 - 1167 * |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114196601A (en) * | 2022-01-14 | 2022-03-18 | 甘肃省科学院生物研究所 | Microbial agent capable of promoting growth, biocontrol and degrading organophosphorus pesticide and preparation method thereof |
| CN114946883A (en) * | 2022-03-21 | 2022-08-30 | 云南师范大学 | Cucurbitacin capable of enhancing disease resistance of plants and application thereof |
| CN114591867A (en) * | 2022-03-29 | 2022-06-07 | 甘肃省科学院生物研究所 | Composite microbial preparation for enhancing corn growth effect and preparation method and application thereof |
| CN114982589A (en) * | 2022-04-21 | 2022-09-02 | 甘肃省科学院生物研究所 | Corn seeding time probiotic composite matrix and preparation method, matrix cup and application thereof |
| CN115305220A (en) * | 2022-07-26 | 2022-11-08 | 中国农业大学 | Pseudomonas capable of fixing nitrogen and promoting plant growth and application thereof |
| CN115305220B (en) * | 2022-07-26 | 2024-02-13 | 中国农业大学 | Pseudomonas with nitrogen fixation and plant growth promotion capabilities and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101659934B (en) | Antagonistic bacteria preventing and removing continuous cropping banana Panama wilt disease and microbial organic fertilizer thereof | |
| CN106591185B (en) | Bacillus amyloliquefaciens plant subspecies and preparation and application of microbial inoculum thereof | |
| CN113862189A (en) | Corn growth promoting, probiotic and biocontrol bacterial fertilizer integrated preparation and preparation method thereof | |
| CN110564646B (en) | Bacillus belgii CY30 and application thereof in prevention and treatment of tea round spot | |
| CN101974428A (en) | Complex microbial preparation capable of resisting replant obstacle resistance and preparation method thereof | |
| CN109306328A (en) | Saline-alkali tolerant bacterial manure microorganism be separately cultured and serial bacterial fertilizer products production technology | |
| CN109896900A (en) | A kind of preparation and application of complex micro organism fungicide and biological organic complex fertilizer | |
| CN109734524A (en) | A kind of ecological system fertilizer of biologic ferment and the preparation method and application thereof | |
| CN105439725A (en) | Paenibacillus polymyxa pesticide-fertilizer for farm onsite fermentation and applications thereof | |
| CN106146194B (en) | A kind of anti-continuous cropping microbial inoculum | |
| CN112006035B (en) | Microbial source bactericide for preventing and treating plant diseases and preparation method thereof | |
| CN104671853A (en) | Preparation method of compound photosynthetic bacterial fertilizer | |
| CN112342165A (en) | Bacillus mucilaginosus and application thereof in agricultural production | |
| CN101948780A (en) | Antagonist bacterium for preventing and treating continuous cropping hot pepper epidemic disease and microbial organic fertilizer thereof | |
| CN101463339B (en) | Inocula for preventing apple fungous diseases and increasing yield, and special strain therefore and use thereof | |
| CN108795797B (en) | Corn root system endophytic enterobacter cloacae and application thereof | |
| CN107628894A (en) | Composite bacteria agent increase soil fertility and its preparation method and application | |
| CN114437982A (en) | Bacillus amyloliquefaciens for improving soil fertilizer efficiency and application thereof | |
| CN110122485A (en) | A kind of trichoderma, Trichoderma and preparation method thereof | |
| CN112111431B (en) | Plant growth promoting strain XN-K13 and application thereof | |
| CN110791459B (en) | Bacillus subtilis for preventing and controlling continuous cropping lily soil-borne blight and application thereof | |
| CN109182194A (en) | One plant of Yang Ling rhizobium for promoting coronule flower growth and its cultural method and application | |
| CN108949635A (en) | Bacillusmusilaginosiengineering, microbial-bacterial fertilizer, preparation method and its application | |
| CN115651846A (en) | Trichoderma fungicide, biological organic fertilizer, preparation method and application thereof | |
| CN110819563B (en) | Brevibacillus brevis 3C and application thereof in promoting growth of pleurotus eryngii |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |