CN106831015A - A kind of bacillus laterosporus microbial manure and its production and use - Google Patents
A kind of bacillus laterosporus microbial manure and its production and use Download PDFInfo
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- CN106831015A CN106831015A CN201710098364.1A CN201710098364A CN106831015A CN 106831015 A CN106831015 A CN 106831015A CN 201710098364 A CN201710098364 A CN 201710098364A CN 106831015 A CN106831015 A CN 106831015A
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- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F11/00—Other organic fertilisers
- C05F11/08—Organic fertilisers containing added bacterial cultures, mycelia or the like
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- 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
Abstract
The present invention relates to a kind of preparation method of bacillus laterosporus microbial manure, comprise the following steps：S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor；S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained；S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid；S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid；S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains fermenting agent；S6：Fermenting agent and organic matter mixture are carried out into compost fermentation treatment, the microbial manure is obtained.Methods described is by being mutually combined between unique multiple technical characteristic and cooperates with, so as to obtain the fermenting agent of the final bacillus laterosporus with excellent technique effect, the widespread demand of multiple fields is disclosure satisfy that, is had a good application prospect and industrial production potential.
Description
Technical field
The present invention relates to a kind of microbial manure and its production and use, more specifically, it is related to a kind of side spore bud
Spore bacillus microorganisms fertilizer and its production and use, belongs to microculture and microbial manure applied technical field.
Background technology
Bacillus laterosporus (Bacilluslaterosporus) be gram-positive bacteria, its gemma for ellipse, side life,
Middle life is near middle raw, and sporangiocyst expands, and the gemma that dissociates is while, presentation canoe shape thicker than another side.
Bacillus laterosporus has important application in multiple fields, for example：1st, it can secrete substantial amounts of chitinase, suppression
Fungus disease processed；2nd, there can be very strong inhibitory action to harmful bacterias such as vibrios, Escherichia coli and baculovirals；3rd, can subtract
Few fish, shrimp disease occur, and improve yield；4th, residual bait, excrement, organic matter etc. in decomposable asymmetric choice net pond, eliminate blue-green algae, purify water；5、
Promote plant root growth, strengthen Root Absorption ability, so as to improve crop yield；6th, pathogen inside and outside plant processed is faced upward numerous
Grow, mitigate pest and disease damage, reduce residues of pesticides；7th, chesson is improved, soil compaction phenomenon is solved, so that activating soil, improves
Utilization rate of fertilizer；8th, strengthen plant metabolism, promote photosynthesis and reinforcing blade protection film, resist pathogen；9th, strengthen
Photosynthesis, improves chemical fertilizer utilization ratio, reduces nitrate content；10th, solidify some heavy metals, reduce heavy metal in plant and contain
Amount.
Just because of above-mentioned all multipurposes of bacillus laterosporus, so as in animal husbandry, feed industry, agricultural chemicals, fertilizer, kill
All had a wide range of applications in the multiple fields such as bacterium, gardens industry, aquaculture, proportion of crop planting, cultivating seedlings, and its application field
With new purposes also always in research and expansion.
Also just because of bacillus laterosporus so important effect, people for various bacillus laterosporus products and
Cultural method and culture medium of bacillus laterosporus etc. have carried out substantial amounts of further investigation, achieve many achievements in research, example
Such as：
CN102399710A discloses a kind of new bacillus laterosporus, and preserving number is CGMCC No.5090 and by the bacterium
Microorganism fungus kind agent and microbial manure and their preparation method prepared by strain.The microorganism fungus kind produced using the strain
Agent has effective bacterium number and the shelf-life more long higher.
CN103289937A discloses a kind of method that high density solid fermentation produces bacillus laterosporus viable bacteria, and it includes
Following steps：Strain selection, inclined-plane seed culture, one-level solid seed culture, two grades of solid seed cultures, solid fermentation trainings
Support, tunning drying and tunning are flashed.The culture medium raw material for being used all is to use agricultural byproducts, low cost, formula
Rationally, culture propagation nutritional need is disclosure satisfy that, the pH value change of culture medium in calcium carbonate buffering incubation is particularly added,
The bacillus laterosporus viable bacteria content of solid fermentation can be substantially increased, experiment and production prove what is prepared using the inventive method
The living bacteria count of bacillus laterosporus up to 50,000,000,000/gram, product miscellaneous bacteria rate is less than 5%, and gemma rate is up to more than 90%,
The production cost of enterprise can be substantially reduced, the need for meeting microbial bacterial agent production.
CN103416321A discloses a kind of bacillus laterosporus fermentation bed and preparation method thereof.Under shown fermentation bed passes through
State method making：By bedding and padding raw material and chicken manure 3-5 by volume:1 is well mixed, and is inoculated with bacillus laterosporus preparation, water content
It is 40-60%, carries out piling up pre fermentation 5-7 days；Pre fermentation bedding and padding are laid in hen house, the bedding and padding of same thickness are laid thereon
Raw material, is mixed evenly to prepare fermentation bed, and the fermentation bed water content is 30-60%, and the gross thickness of the fermentation bed is 8-25cm；Institute
It is 2-5 that bedding and padding raw material is stated by volume ratio:Husk and the sawdust composition of 5-8.The bacillus laterosporus fermenting bed padding thickness is small,
The usage amount of bedding and padding is few, is significantly reduced the work workload of raiser, cost-effective；Effect is still fine at high temperature, can
To suppress and kill mosquito, improve birds function of intestinal canal, efficient degradation albumen, starch, fat, fecaluria, and ammonia taste, stink compared with
It is few；Use and expand column convenient, can Continuous disinfection.
CN103725626A discloses a kind of bacillus laterosporus preparation, and its preparation technology is as follows：Bacillus laterosporus bacterium
Plant (ampoul tube)-test tube slant Spawn incubation-shaking flask Spawn incubation-seeding tank deep fermentation culture-closed solid hair
Ferment case culture-bacillus laterosporus preparation.It uses continuously fermenting for advanced " deep fermentation+closed type solid fermentation "
Technique, using totally-enclosed autocontrol solid fermentation apparatus, makes the whole process of solid fermentation all in totally enclosed automatic monitoring
Under, automation inspection is carried out to environmental conditions such as the medium component in fermentation process, formula, temperature, humidity, ventilation, pH value
Survey, electronical display, automatically control, not only increase substantially yield, and ensure that the microbial foliage prepared by said preparation increases
Imitate the quality of agent.
CN105219676A discloses a kind of cultural method of bacillus laterosporus, and the Microbiological Culture Collection number is
CGMCC No.5090, the cultural method is as follows：1st, by 1 volume bacteria suspension access 20 times of volume No. 1 culture mediums in, 30-38
DEG C aerobic culture 8 hours；2nd, the nutrient solution that step 1 is obtained is added in about 20 times of No. 2 culture mediums of volume, 30-38 DEG C has
Oxygen culture 8 hours；3rd, the nutrient solution that step 2 is obtained is added in about 20 times of No. 3 culture mediums of volume, 35-38 DEG C of culture 8 is small
When, obtain final product can production microorganism fungus kind culture.
CN105272406A discloses a kind of microbial manure prepared by bacillus laterosporus and preparation method thereof, the fertilizer
Expect that gained culture and turf powder are mixed by preserving number for the microorganism fungus kind of CGMCC No.5090 obtains culture by culture
Conjunction obtains microorganism fungus kind agent, and gained microorganism fungus kind agent is mixed with common fertilizer forms again, wherein the microbial bacteria
The culture side for planting culture is as follows：1st, by 1 volume bacteria suspension access 20 times of volume No. 1 culture mediums in, 30-38 DEG C of aerobic culture
8 hours；2nd, the nutrient solution that step 1 is obtained is added in about 20 times of No. 2 culture mediums of volume, 30-38 DEG C of aerobic culture 8 is small
When；3rd, the nutrient solution that step 2 is obtained is added in about 20 times of No. 3 culture mediums of volume, 35-38 DEG C is cultivated 8 hours, and obtaining final product can
The culture of production.
As described above, disclosing the multiple-microorganism fertilizer comprising bacillus laterosporus and side spore gemma in the prior art
The cultural method of bacillus, but for new bacillus laterosporus microbial manure and preparation method thereof, still suffer from continuing to study
Necessity and demand, this is currently the study hotspot and emphasis in microbial manure field, even more the present invention be accomplished it is dynamic
Lean on where power and basis.
The content of the invention
In order to seek bacillus laterosporus microbial manure and its production and use, present inventor has performed substantial amounts of
Further investigation, after creative work has been paid, so as to complete the present invention.
Specifically, the present invention relates to following aspects.
One side, the present invention relates to a kind of preparation method of bacillus laterosporus microbial manure, methods described bag
Include following steps：
S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor；
S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained；
S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid；
S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid；
S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains bacillus laterosporus
Fermenting agent；
S6：Bacillus laterosporus fermenting agent and organic matter mixture are carried out into compost fermentation treatment, it is described so as to obtain
Bacillus laterosporus microbial manure.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S1 is specific as follows：
S1-1：Selection bacillus laterosporus bacterial strain, and be aseptically inoculated in activation training is carried out on slant medium
Support, obtain activated strains；
S1-2：Sterile water wash inclined-plane, obtains the seed liquor.
Wherein, the bacillus laterosporus bacterial strain in step S1 may be selected any of spore Bacillus strain, for example
CGMCC No.5090, CGMCC No.1755, spore Bacillus strain BL-11, spore Bacillus strain BL-22, the spore bud known
Spore bacillus strain BL-21, spore Bacillus strain 1.864, spore Bacillus strain BL-1, spore Bacillus strain 2-Q-9 etc.
Deng these bacterial strains can be obtained by various commercial channel, and this is no longer going to repeat them.
Wherein, in step S1, the composition of slant medium is described in per 100g：Agar 5-6g, peptone 3-4g, yeast leaching
Cream 0.02-0.03g, analysis for soybean powder 2-3g, corn flour 0.8-1.4g, beef extract 3-4g, sodium chloride 0.2-0.5g, glucose 0.06-
0.1g, balance of sterilized water, and adjust pH value for 7-7.2 (regulating measure is routine techniques, is repeated no more).
Wherein, analysis for soybean powder and corn flour cross 200 mesh sieves.
Wherein, in step S1, the temperature of the activation culture is 30-34 DEG C, for example, can be 30 DEG C, 32 DEG C or 34 DEG C.
Wherein, in step S1, time of the activation culture is 34-40 hour, for example can be 34 hours, 36 hours, 38
Hour or 40 hours.
Wherein, in step S2, using aseptic water washing inclined-plane, flushing liquor is collected, and it is 1.3 × 10 to adjust bacterial concentration8-
1.4×108Individual/ml, so as to obtain the seed liquor.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S2 is specific as follows：
S2-1：Prepare the first nutrient solution
Following material is weighed respectively：Metallic element aqueous solution 25ml, vitamin E 1.5g, citric acid 0.15g, L- dried meat ammonia
Sour 0.2mg, sucrose 2.5g, agar 8g, peptone 12g, yeast extract 1.4g, ammonium sulfate 1g, sodium phosphate 2.5g, beef extract 3.5g
With sodium nitrate 4g；
Above-mentioned substance is added in deionized water, is sufficiently stirred for completely, and 1000ml is settled to deionized water, obtained final product
To first nutrient solution.
S2-2：According to 1:The volume ratio of 40-50, the seed liquor is inoculated in first nutrient solution, at 34-36 DEG C
Lower ventilation shaken cultivation 20-24 hours, so as to obtain the first amplification cultivation liquid.
Wherein, in step S2-1, the metallic element aqueous solution be by 2.5g manganese sulfates, 3g zinc chloride, 4g magnesium sulfate,
1.5g cobalt chlorides, 2g copper nitrates, 3g calcium chloride, 4g frerrous chlorides, 3g iron chloride are completely dissolved in 1000ml distilled water and obtain
Arrive；Then 20-30ml therein is measured, the 20-30ml metallic element aqueous solution used in as above-mentioned steps S2-1.
Wherein, in step S2-2, it is 1 that the seed liquor is connect with the volume ratio of first nutrient solution:40-50, for example may be used
It is 1:40、1:45 or 1:50.
Wherein, in step S2-2, the ventilating ratio of the ventilation is 1:0.8-1, for example, can be 1:0.8、1:0.9 or 1:1.
Wherein, in step S2-2, in the be passed through gas of the ventilation, oxygen volume content is 14-20%, for example, can be
14%th, 16%, 18% or 20%, preferably 16-18%, most preferably 17%.
Inventor has found that the oxygen content in the ventilation has significant impact for final result, in suitable body
Best technique effect can be obtained under product content.
Wherein, in step S2-2, the rotating speed of the shaken cultivation is 150-170 revs/min.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S3 is specific as follows：
S3-1：Yeast extract consumption in first nutrient solution is halved (be reduced to 0.7g), and the metallic element aqueous solution
Consumption increase be twice (increase to 50ml), other are constant, so as to obtain second nutrient solution (namely according to first
The same procedure of nutrient solution is prepared, be nothing but yeast extract consumption halve with metallic element amount of aqueous solution used increase be twice and
)；
S3-2：According to 1:The volume ratio of 30-35, the first amplification cultivation liquid is connect and is added to second nutrient solution
In, air surge culture is passed through at 34-36 DEG C 14-16 hours, so as to obtain the second amplification cultivation liquid.
Wherein, in step S3-1, the consumption of the metallic element aqueous solution is increased by reducing the consumption of yeast extract, is sent out
Superior technique effect can now be obtained, it should be now to have been obtained for sufficient amount and activity in the first amplification cultivation liquid
Activation flora, so that the consumption but the demand to metallic element that can reduce yeast extract rise (if yeast extract consumption
It is constant, excessively fast passage, but viable count can be caused to reduce on the contrary, instead result in last effect reduction).
Wherein, normal air is passed through in step S3-2, during shaken cultivation, without as wanted in step S2-3
The content for seeking oxygen is 14-20%.
Wherein, in step S3-2, the ventilating ratio for being passed through air is 1:1.
Wherein, in step S3-2, the rotating speed of the shaken cultivation is 150-170 revs/min.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S4 is specific as follows：
The second amplification cultivation liquid is added in the liquid fermentation medium for 20-30 times of its volume carries out two-part
Fermentation at elevated temperatures culture, fermented and cultured total time is 10-12 hours, and period is passed through air always, and the ventilating ratio for being passed through air is 1:
0.6-1, for example, can be 1:0.6、1:0.8 or 1:1, so as to obtain the zymotic fluid.
Wherein, the liquid fermentation medium is with every liter (L) (the 1L compounds for finally giving, namely 1L liquid fermentations
Culture medium) meter, comprising yeast extract 0.5g, peptone 2g, maltose 4g, glucose 1g, manganese sulfate 0.6g, sodium chloride 3.8g,
Analysis for soybean powder 7g, boric acid 1.5g, potassium nitrate 2.5g, urea 6g, lysine 0.7g and ammonium sulfate 1.5g, balance of sterilized water.
Wherein, analysis for soybean powder crosses 200 mesh sieves.
Wherein, the two-part fermentation at elevated temperatures culture refers to (to start meter in the first half time of fermented and cultured total time
5-6 hours for rising) in, fermentation temperature is 32-34 DEG C, and temperature then is increased into 38-40 DEG C in 10 minutes, and in the temperature
The lower fermentation for completing follow-up remaining time.
Inventor's discovery, by two-part fermentation at elevated temperatures culture so, can obtain best technique effect, it should be
By after Preliminary fermentation, continuing to heat up, can further improve fermentation efficiency, improve viable count.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S5 is specific as follows：
According to mass ratio 1:The ratio of 10-14, the zymotic fluid is drenched in solid medium, adds appropriate deionization
Water, makes overall biodiversity content for 32-36%, then fully turn it is uniform, load length be 3 meters, width be 1.5 meters and
During depth is 1.5 meters of fermentation tank, natural packing ferments 60-70 hours, and period remains that overall biodiversity content is
32-36%, and be passed through sterile wind to keep the central temperature inside fermentation tank is 56-60 DEG C；After fermentation ends, lead to heated-air drying,
Until biodiversity content is less than 5%, fermenting agent is obtained.
Wherein, during fermentation, if interval of the biodiversity content of entirety less than 32-36%, by drenching into going
The method of ionized water is held it in the range of this, and this is the routine that those skilled in the art should possess after the present invention is read
Ability, is no longer described in detail herein.
Wherein, the solid medium is obtained as follows：By 3 weight portion cotton dregs, 4 weight portion peanut shells
Powder, 2 weight portion wheat bran, 0.2 weight portion molasses, 2 weight portion peanut cakes, 2.5 weight portion dry silkworm excrements, the dry corn of 10 weight portions
Powder of straw, 0.8 weight portion plant ash, 2 weight portion fishbone dusts, the dry bagasse of 4 weight portions, 0.008 part by weight of cellulose enzyme,
1.8 weight portion Dry chick faeces, 0.1 weight portion ammonium chloride, crushed 50 mesh sieves, then 130-140 DEG C of high temperature after being well mixed
Vapor fully sterilizes, that is, obtain the solid medium.
In the preparation method of bacillus laterosporus microbial manure of the invention, the step S6 is specially：According to 1:
The mass ratio of 40-50, the bacillus laterosporus fermenting agent and organic matter mixture is sufficiently mixed, and carry out compost
Fermentation process, obtains final product the bacillus laterosporus microbial manure after post processing.
Wherein, the organic matter mixture is the arbitrary proportion mixing of all any organic matters that can be used to prepare organic fertilizer
Thing, for example can be dry maize straw, drying wheat stalk, wheat bran, drying bagasse, vinasse, dregs of beans, cotton dregs, peanut shell powder,
Dry rice straw, biogas residue, dry pig manure, Dry chick faeces, various bone meal etc. in any various mixtures with arbitrary proportion, but
For comprehensive nutrition meter, it is preferred to use at least 5 kinds therein, and preferably add appropriate corn flour, beancake powder, peanut cake
The assisted fermentations such as powder, molasses, cellulase, amylase and decomposition nutrition matter.
Wherein, the compost fermentation treatment can be using the conventional fermentation techniques in compost fermentation field, until it has fermented
It is all only, is no longer described one by one herein.
Wherein, after the completion of the post processing refers to compost fermentation, it is dried to biodiversity content less than 5%, obtains final product
The bacillus laterosporus microbial manure.
Second aspect, the present invention relates to the bacillus laterosporus microorganism fertilizer according to obtained in above-mentioned preparation method
Material.
Preparation method of the invention is by unique multiple technical characteristics synergy each other and mutually promotes, from
And good technique effect is achieved, can provide with high activity, highdensity bacillus laterosporus fermenting agent, and then
The bacillus laterosporus microbial manure with excellent properties is arrived, so as to industrially have a good application prospect and produce
Potentiality.
3rd aspect, the present invention relates to purposes of the bacillus laterosporus microbial manure in agriculture field.
Find that the bacillus laterosporus microbial manure has superperformance by research, thus can in agriculture field,
For example in the plantation and/or cultivation of crops, veterinary antibiotics, flowers or nursery stock etc. have a good application prospect and potentiality.
As described above, the invention provides a kind of bacillus laterosporus microbial manure and its production and use, institute
Preparation method is stated by being mutually combined between unique multiple technical characteristics and is cooperateed with, so as to obtain imitating with excellent technology
The final bacillus laterosporus microbial manure of fruit, disclosure satisfy that the widespread demand of agricultural technology field, with good application
Prospect and industrial production potential.
Specific embodiment
Below by specific embodiment, the present invention is described in detail, but the purposes of these exemplary embodiments and
Purpose is only used for enumerating the present invention, not constitutes any type of any restriction to real protection scope of the invention, more non-to incite somebody to action
Protection scope of the present invention is confined to this.
Preparation example 1：Slant medium
Often the composition of 100g slant mediums is：Agar 5.5g, peptone 3.5g, yeast extract 0.025g, analysis for soybean powder
2.5g, corn flour 1.1g, beef extract 3.5g, sodium chloride 0.35g, glucose 0.08g, balance of sterilized water, its pH value is 7-
7.2。
Wherein, analysis for soybean powder and corn flour cross 200 mesh sieves.
Preparation example 2：The preparation of the metallic element aqueous solution
By 2.5g manganese sulfates, 3g zinc chloride, 4g magnesium sulfate, 1.5g cobalt chlorides, 2g copper nitrates, 3g calcium chloride, 4g protochlorides
Iron, 3g iron chloride are completely dissolved in 1000ml distilled water, so as to obtain the metallic element aqueous solution.
Preparation example 3：Liquid fermentation medium
Liquid fermentation medium in terms of every liter (L), comprising yeast extract 0.5g, peptone 2g, maltose 4g, glucose
1g, manganese sulfate 0.6g, sodium chloride 3.8g, analysis for soybean powder 7g, boric acid 1.5g, potassium nitrate 2.5g, urea 6g, lysine 0.7g and sulfuric acid
Ammonium 1.5g, balance of sterilized water.
Wherein, analysis for soybean powder crosses 200 mesh sieves.
Preparation example 4：The preparation (i.e. solid medium in step S5) of solid medium
By 3 weight portion cotton dregs, 4 weight portion peanut shell powders, 2 weight portion wheat bran, 0.2 weight portion molasses, 2 weight portion peanuts
Cake, 2.5 weight portion dry silkworm excrements, the dry corn stalk powder of 10 weight portions, 0.8 weight portion plant ash, 2 weight portion fishbone dusts, 4 weights
The dry bagasse of amount part, 0.008 part by weight of cellulose enzyme, 1.8 weight portion Dry chick faeces, 0.1 weight portion ammonium chloride, mixing are equal
50 mesh sieves were crushed after even, then 130-140 DEG C of high-temperature vapor fully sterilizes, that is, obtained the solid medium.
Unless otherwise stated, following all of slant medium, the metallic element aqueous solution, liquid fermentation medium and
Solid medium is the respective substance in corresponding above-mentioned preparation example 1-4.
Embodiment 1
S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor
S1-1：Selection bacillus laterosporus bacterial strain CGMCC No.1755, and aseptically it is inoculated in slant medium
On carry out activation culture, the temperature of the activation culture is 32 DEG C, and the time of activation culture is 37 hours, so as to obtain activating bacterium
Strain；
S1-2：Sterile water wash inclined-plane, obtains the seed liquor
Using aseptic water washing inclined-plane, flushing liquor is collected, and it is 1.35 × 10 to adjust bacterial concentration8Individual/ml, so as to obtain
The seed liquor；
S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained
S2-1：Prepare the first nutrient solution
Following material is weighed respectively：Metallic element aqueous solution 25ml, vitamin E 1.5g, citric acid 0.15g, L- dried meat ammonia
Sour 0.2mg, sucrose 2.5g, agar 8g, peptone 12g, yeast extract 1.4g, ammonium sulfate 1g, sodium phosphate 2.5g, beef extract 3.5g
With sodium nitrate 4g；
Above-mentioned substance is added in deionized water, is sufficiently stirred for completely, and 1000ml is settled to deionized water, obtained final product
To the first nutrient solution；
S2-2：According to 1:45 volume ratio, the seed liquor is inoculated in first nutrient solution, at 34-36 DEG C
(ventilating ratio is 1 to ventilation shaken cultivation within 22 hours:0.9, oxygen volume content is 17%, and the rotating speed of shaken cultivation is 160 revs/min
Clock), so as to obtain the first amplification cultivation liquid；
S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid
S3-1：Yeast extract consumption in first nutrient solution is halved (be reduced to 0.7g), and the metallic element aqueous solution
Consumption increase be twice (increase to 50ml), other are constant, so as to obtain the second nutrient solution；
S3-2：According to 1:32.5 volume ratio, the first amplification cultivation liquid is connect and is added in second nutrient solution,
Air surge culture is passed through at 34-36 DEG C, and (ventilating ratio is 1 within 15 hours:1, the rotating speed of shaken cultivation is 160 revs/min), from
And obtain the second amplification cultivation liquid；
S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid
The second amplification cultivation liquid is added in the liquid fermentation medium for 25 times of its volume carries out two-part liter
Warm fermented and cultured, fermented and cultured total time is 11 hours, and period is passed through air always, and the ventilating ratio for being passed through air is 1:0.8, from
And obtain zymotic fluid；
Wherein, the two-part fermentation at elevated temperatures culture refer to fermented and cultured total time the first half time it is (i.e. preceding 5.5
Hour) in, fermentation temperature is 33 DEG C, and temperature then is increased into 39 DEG C in 10 minutes, and completes follow-up surplus at such a temperature
The fermentation of remaining time (5.5 hours after i.e.)；
S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains bacillus laterosporus
Fermenting agent, specially：
According to mass ratio 1:12 ratio, the zymotic fluid is drenched in solid medium, adds appropriate amount of deionized water,
Make overall biodiversity content for 32-36%, then fully turn uniformly, load length for 3 meters, width are for 1.5 meters and deep
Spend in the fermentation tank for 1.5 meters, natural packing ferments 65 hours, period remains that overall biodiversity content is 32-
36%, and be passed through sterile wind to keep the central temperature inside fermentation tank is 56-60 DEG C；After fermentation ends, lead to heated-air drying, until
Biodiversity content is less than 5%, obtains fermenting agent, is named as F1；
S6：Bacillus laterosporus fermenting agent and organic matter mixture are carried out into compost fermentation treatment, so as to obtain side spore
Micro-organisms bacillus fertilizer, specially：
According to 1:45 mass ratio, the bacillus laterosporus fermenting agent and organic matter mixture are fully mixed
Close, and compost fermentation treatment is carried out according to routine techniques, after the completion of compost fermentation, be dried to biodiversity content and be less than
5%, bacillus laterosporus microbial manure is obtained final product, it is named as C1；
Wherein, the organic matter mixture is mass ratio 1:1:0.1:2:2:1:0.3 1 weight portion dries maize straw, 1
Weight portion wheat bran, 0.1 weight portion dregs of beans, 2 weight portion drying bagasses, 2 weight portion biogas residues, 1 weight portion Dry chick faeces, 0.3 weight
Amount part fishbone dust, 0.05 parts by weight Corn powder, 0.01 weight portion molasses, the mixture of 0.005 weight portion.
Embodiment 2
S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor
S1-1：Selection bacillus laterosporus bacterial strain CGMCC No.1755, and aseptically it is inoculated in slant medium
On carry out activation culture, the temperature of the activation culture is 30 DEG C, and the time of activation culture is 40 hours, so as to obtain activating bacterium
Strain；
S1-2：Sterile water wash inclined-plane, obtains the seed liquor
Using aseptic water washing inclined-plane, flushing liquor is collected, and it is 1.3 × 10 to adjust bacterial concentration8Individual/ml, so as to obtain
The seed liquor；
S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained
S2-1：Prepare the first nutrient solution
Following material is weighed respectively：Metallic element aqueous solution 25ml, vitamin E 1.5g, citric acid 0.15g, L- dried meat ammonia
Sour 0.2mg, sucrose 2.5g, agar 8g, peptone 12g, yeast extract 1.4g, ammonium sulfate 1g, sodium phosphate 2.5g, beef extract 3.5g
With sodium nitrate 4g；
Above-mentioned substance is added in deionized water, is sufficiently stirred for completely, and 1000ml is settled to deionized water, obtained final product
To the first nutrient solution；
S2-2：According to 1:40 volume ratio, the seed liquor is inoculated in first nutrient solution, at 34-36 DEG C
(ventilating ratio is 1 to ventilation shaken cultivation within 20 hours:0.8, oxygen volume content is 17%, and the rotating speed of shaken cultivation is 150 revs/min
Clock), so as to obtain the first amplification cultivation liquid；
S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid
S3-1：Yeast extract consumption in first nutrient solution is halved (be reduced to 0.7g), and the metallic element aqueous solution
Consumption increase be twice (increase to 50ml), other are constant, so as to obtain the second nutrient solution；
S3-2：According to 1:30 volume ratio, the first amplification cultivation liquid is connect and is added in second nutrient solution,
Air surge culture is passed through at 34-36 DEG C, and (ventilating ratio is 1 within 14 hours:1, the rotating speed of shaken cultivation is 150 revs/min) so that
Obtain the second amplification cultivation liquid；
S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid
The second amplification cultivation liquid is added in the liquid fermentation medium for 20 times of its volume carries out two-part liter
Warm fermented and cultured, fermented and cultured total time is 10 hours, and period is passed through air always, and the ventilating ratio for being passed through air is 1:0.6, from
And obtain zymotic fluid；
Wherein, the two-part fermentation at elevated temperatures culture refers to that (i.e. preceding 5 small in the first half time of fermented and cultured total time
When) in, fermentation temperature is 32 DEG C, and temperature then is increased into 38 DEG C in 10 minutes, and completes follow-up remaining at such a temperature
The fermentation of time (5 hours after i.e.)；
S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains bacillus laterosporus
Fermenting agent, specially：
According to mass ratio 1:10 ratio, the zymotic fluid is drenched in solid medium, adds appropriate amount of deionized water,
Make overall biodiversity content for 32-36%, then fully turn uniformly, load length for 3 meters, width are for 1.5 meters and deep
Spend in the fermentation tank for 1.5 meters, natural packing ferments 60 hours, period remains that overall biodiversity content is 32-
36%, and be passed through sterile wind to keep the central temperature inside fermentation tank is 56-60 DEG C；After fermentation ends, lead to heated-air drying, until
Biodiversity content is less than 5%, obtains fermenting agent, is named as F2；
S6：Bacillus laterosporus fermenting agent and organic matter mixture are carried out into compost fermentation treatment, so as to obtain side spore
Micro-organisms bacillus fertilizer, specially：
According to 1:40 mass ratio, the bacillus laterosporus fermenting agent and organic matter mixture are fully mixed
Close, and compost fermentation treatment is carried out according to routine techniques, after the completion of compost fermentation, be dried to biodiversity content and be less than
5%, bacillus laterosporus microbial manure is obtained final product, it is named as C2；
Wherein, the organic matter mixture is with the organic matter mixture in embodiment 1.
Embodiment 3
S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor
S1-1：Selection bacillus laterosporus bacterial strain CGMCC No.1755, and aseptically it is inoculated in slant medium
On carry out activation culture, the temperature of the activation culture is 34 DEG C, and the time of activation culture is 34 hours, so as to obtain activating bacterium
Strain；
S1-2：Sterile water wash inclined-plane, obtains the seed liquor
Using aseptic water washing inclined-plane, flushing liquor is collected, and it is 1.4 × 10 to adjust bacterial concentration8Individual/ml, so as to obtain
The seed liquor；
S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained
S2-1：Prepare the first nutrient solution
Following material is weighed respectively：Metallic element aqueous solution 25ml, vitamin E 1.5g, citric acid 0.15g, L- dried meat ammonia
Sour 0.2mg, sucrose 2.5g, agar 8g, peptone 12g, yeast extract 1.4g, ammonium sulfate 1g, sodium phosphate 2.5g, beef extract 3.5g
With sodium nitrate 4g；
Above-mentioned substance is added in deionized water, is sufficiently stirred for completely, and 1000ml is settled to deionized water, obtained final product
To the first nutrient solution；
S2-2：According to 1:50 volume ratio, the seed liquor is inoculated in first nutrient solution, at 34-36 DEG C
(ventilating ratio is 1 to ventilation shaken cultivation within 24 hours:1, oxygen volume content is 17%, and the rotating speed of shaken cultivation is 170 revs/min
Clock), so as to obtain the first amplification cultivation liquid；
S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid
S3-1：Yeast extract consumption in first nutrient solution is halved (be reduced to 0.7g), and the metallic element aqueous solution
Consumption increase be twice (increase to 50ml), other are constant, so as to obtain the second nutrient solution；
S3-2：According to 1:35 volume ratio, the first amplification cultivation liquid is connect and is added in second nutrient solution,
Air surge culture is passed through at 34-36 DEG C, and (ventilating ratio is 1 within 16 hours:1, the rotating speed of shaken cultivation is 170 revs/min) so that
Obtain the second amplification cultivation liquid；
S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid
The second amplification cultivation liquid is added in the liquid fermentation medium for 30 times of its volume carries out two-part liter
Warm fermented and cultured, fermented and cultured total time is 12 hours, and period is passed through air always, and the ventilating ratio for being passed through air is 1:1, so that
Obtain zymotic fluid；
Wherein, the two-part fermentation at elevated temperatures culture refers to that (i.e. preceding 6 small in the first half time of fermented and cultured total time
When) in, fermentation temperature is 34 DEG C, and temperature then is increased into 40 DEG C in 10 minutes, and completes follow-up remaining at such a temperature
The fermentation of time (6 hours after i.e.)；
S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains bacillus laterosporus
Fermenting agent, specially：
According to mass ratio 1:14 ratio, the zymotic fluid is drenched in solid medium, adds appropriate amount of deionized water,
Make overall biodiversity content for 32-36%, then fully turn uniformly, load length for 3 meters, width are for 1.5 meters and deep
Spend in the fermentation tank for 1.5 meters, natural packing ferments 70 hours, period remains that overall biodiversity content is 32-
36%, and be passed through sterile wind to keep the central temperature inside fermentation tank is 56-60 DEG C；After fermentation ends, lead to heated-air drying, until
Biodiversity content is less than 5%, obtains fermenting agent, is named as F3；
S6：Bacillus laterosporus fermenting agent and organic matter mixture are carried out into compost fermentation treatment, so as to obtain side spore
Micro-organisms bacillus fertilizer, specially：
According to 1:50 mass ratio, the bacillus laterosporus fermenting agent and organic matter mixture are fully mixed
Close, and compost fermentation treatment is carried out according to routine techniques, after the completion of compost fermentation, be dried to biodiversity content and be less than
5%, bacillus laterosporus microbial manure is obtained final product, it is named as C2；
Wherein, the organic matter mixture is with the organic matter mixture in embodiment 1.
Comparative example 1-6：The investigation of nutrient solution
Comparative example 1-3：In addition to second nutrient solution of S3 replaces with the first nutrient solution the step of respectively by embodiment 1-3 (i.e.
First liquid culture medium is used in step S2-S3), other operations are constant, so as to repeat embodiment 1-3, point
Comparative example 1-3 is not obtained, and gained bacillus laterosporus fermenting agent is sequentially named as DF1, DF2 and DF3, gained is finally fertile
Material is sequentially named as DC1, DC2 and DC3.
Comparative example 4-6：In addition to first nutrient solution of S2 replaces with the second nutrient solution the step of respectively by embodiment 1-3 (i.e.
Second liquid culture medium is used in step S2-S3), other operations are constant, so as to repeat embodiment 1-3, point
Comparative example 4-6 is not obtained, and gained bacillus laterosporus fermenting agent is sequentially named as DF4, DF5 and DF6, gained is finally fertile
Material is sequentially named as DC4, DC5 and DC6.
Comparative example 7-10：The investigation of oxygen content in being divulged information in step S2-2
The oxygen volume content (other operations are constant) in the step S2-2 shown in following table is respectively adopted, institute is accumulated using oxysome
Content, correspondence embodiment, the name of gained bacillus laterosporus fermenting agent and final fertilizer see the table below 1.
Table 1
Comparative example 11-16：The investigation that two-part heats up in step S4
Comparative example 11-13：Except respectively by embodiment 1-3 the step of S4 two-part heat up be revised as respective first
Whole at a temperature of section to have implemented all of fermentation (the correspondingly warming temperature without 10 minutes), other operations are constant, so that
Repeated embodiment 1-3, respectively obtain comparative example 11-13 (i.e. comparative example 11,12,13 the step of S3 in whole fermentation
Temperature is respectively 33 DEG C, 32 DEG C, 34 DEG C), by gained bacillus laterosporus fermenting agent be sequentially named as DF11, DF12 and
DF13, DC11, DC12 and DC13 are sequentially named as by the final fertilizer of gained.
Comparative example 14-16：Except respectively by embodiment 1-3 the step of S4 two-part heat up be revised as respective second
Whole at a temperature of section to have implemented all of fermentation (the correspondingly warming temperature without 10 minutes), other operations are constant, so that
Repeated embodiment 1-3, respectively obtain comparative example 14-16 (i.e. comparative example 11,12,13 the step of S3 in whole fermentation
Temperature is respectively 39 DEG C, 38 DEG C, 40 DEG C), by gained bacillus laterosporus fermenting agent be sequentially named as DF14, DF15 and
DF16, DC14, DC15 and DC16 are sequentially named as by the final fertilizer of gained.
Performance
A, the investigation of colony counting
According to the traditional test methods in this area, the different bacillus laterosporuses obtained to each embodiment and comparative example
Fermenting agent carries out colony counting investigation at once after preparing, investigated respectively viable count in every gram of fermenting agent and
Gemma rate, as a result see the table below 2.
Wherein, multiple fermenting agents of identical group take its average.
Table 2
As can be seen here：1st, the F1-F3 that the inventive method is obtained has viable count very high and gemma rate；2 and ought make always
During with the first nutrient solution or the second nutrient solution, viable count and gemma rate are significantly reduced, and especially the second nutrient solution is reduced more
It is obvious (see DF4-DF6)；3rd, in step S2-2 divulge information oxygen content it is extremely important, when for 17% when can obtain best skill
Art effect, it is bigger to deviate the value, then viable count and gemma rate reduction more obvious (see DF7-DF10), and this proves the determination of oxygen content
With unobviousness, and effect is unexpected；4th, the two-part fermentation at elevated temperatures in step S4 equally has for final result
Significant impact the most, when warming temperature is not carried out, always in the first stage at a temperature of when fermenting, viable count and gemma rate are most
Difference is (see DF11-DF13)；It is straight in the first stage at a temperature of when fermenting, viable count and gemma rate are also reduced significantly (see DF14-
DF16), but it is better than DF11-DF13, this demonstrate that only being heated up using two-part of the invention, best technology could be obtained
Effect.
The investigation of B, heat-resisting quantity
Each embodiment and comparative example 40 minutes after different bacillus laterosporus fermenting agents are obtained, respectively in difference
Temperature (70 DEG C, 80 DEG C, 90 DEG C and 100 DEG C) under constant temperature keep 20 minutes, its viable count, and and initial live are then measured again
Bacterium number is calculated, and so as to obtain the survival rate after high temperature, as a result see the table below 3.
Wherein, multiple fermenting agents of identical group take its average.
Table 3
As can be seen here：1st, the F1-F3 that the inventive method is obtained has excellent high temperature resistant property, and this prepares organic in fermentation
There is more preferable high-temperature adaptability when fertile；2 and the resistance to elevated temperatures of the fermenting agent of comparative example, especially when higher than 80 DEG C
With obvious reduction, especially DF11-DF13, bacterium colony resistance to elevated temperatures now is significantly reduced.
The investigation of C, storage stability
By each embodiment and comparative example after different bacillus laterosporus fermenting agents are obtained, kept in dark place under normal temperature
Regular hour, and viable count when investigating different time respectively, and calculated with initial viable count, so as to obtain different storages
The survival rate after the time is deposited, 4 are as a result see the table below.
Wherein, multiple fermenting agents of identical group take its average.
Table 4
As can be seen here：1st, the F1-F3 that the inventive method is obtained has excellent storage stability, and this is in feed and microorganism
Fertilizer field has good application value, can store the longer time；2 and the storage stability of the fermenting agent of comparative example
Then there is obvious reduction, especially significantly, wherein DF11-DF13 is worst for DF7-DF8, DF11-DF16 reduction, and this proves step
Oxygen content in rapid S3 and the two-part fermentation at elevated temperatures in step S4 can unpredictably influence final performance.
The plantation culture Performance of fertilizer
I, corn planting volume increase are investigated
According to every 100 square metres as one piece of sample plot (corn variety be good beautiful 99), be divided into control group, chemical group,
Microbial manure group, it is specific as follows：
Control group：Any fertilizer is not used.
Chemical group：Using normal chemical fertilizer, i.e. N, P, K fertilizer, 5kg ammonium chlorides, 5kg potassium sulfates and 3kg phosphorus can be used
Sour ammonium is applied fertilizer.
Microbial manure group：Using the microbial manure of the 5kg embodiment of the present invention or comparative example, (each sample plot is only respectively
Use a kind of microbial manure), and N, P, K of the same ratio of the amount of additional chemical group 1/4 are fertile.
After normal field management and harvesting, per mu yield yield is converted to, as a result see the table below 5.
Table 5
Wherein, wherein, the data of C1-C3 carry out correspondence, example, " 805,801,799 " meaning in that same order respectively
C1 for 805, C2 is that 801 and C3 is 799.Numbered corresponding relation (be also so tool in following all forms
Have same implication, no longer list one by one when the time comes), this is no longer going to repeat them.
As can be seen here, microbial manure C1-C3 of the invention has effect of increasing production the most excellent, increases relative to chemical group
Produce>10%, although other contrast fertilizer will be significantly below C1-C3, but still chemical group is significantly better than, also achieve good
Effect of increasing production.
II, growing tomatoes are investigated
According to every 100 square metres as one piece of sample plot (tomato variety be in miscellaneous 109), be divided into control group, chemistry
Group, microbial manure group, it is specific as follows：
Control group：Any fertilizer is not used.
Chemical group：Using normal chemical fertilizer, i.e. N, P, K fertilizer, 8kg ammonium chlorides, 4kg potassium sulfates and 6kg phosphoric acid are used
Ammonium is applied fertilizer.
Microbial manure group：Using the microbial manure of the 6kg embodiment of the present invention or comparative example, (each sample plot is only respectively
Use a kind of microbial manure), and N, P, K of the same ratio of the amount of additional chemical group 1/4 are fertile.
After normal field management and harvesting, per mu yield yield is converted to, (is as benchmark using the yield of control group
100) fractional yield of chemical group and microbial manure group, is calculated, 6 are as a result see the table below.
Wherein, due to each group in multiple fertilizer corresponding to tomato production deviation less, therefore use each group
Average value investigated.
Table 6
As can be seen here, microbial manure C1-C3 of the invention has berry effect of increasing production the most excellent, relative to control
Group increases 27.8%, also has relative to chemical group and significantly increases production.And contrast microbial manure then has relative to C1-C3
Significant reduction, especially DC11-DC13, this performance trend also with fermenting agent are consistent.But even significantly reduce,
Still it is significantly better than control group and chemical group.
In sum, the invention provides a kind of bacillus laterosporus microbial manure and its production and use, institute
Preparation method is stated by being mutually combined between unique multiple technical characteristics and is cooperateed with, so as to obtain imitating with excellent technology
The final bacillus laterosporus microbial manure of fruit, disclosure satisfy that the widespread demand of agricultural technology field, with good application
Prospect and industrial production potential.
It should be appreciated that the purposes of these embodiments is merely to illustrate the present invention and is not intended to limit protection model of the invention
Enclose.Additionally, it will also be appreciated that after technology contents of the invention have been read, those skilled in the art can make each to the present invention
Plant and change, change and/or modification, all these equivalent form of value equally falls within the guarantor that the application appended claims are limited
Within the scope of shield.
In sum, the invention provides a kind of Novel culture method of bacillus laterosporus, methods described is by uniqueness
Multiple technical characteristics between be mutually combined and cooperate with, so as to obtain the final bacillus laterosporus with excellent technique effect
Fermenting agent, disclosure satisfy that the widespread demand of multiple fields, have a good application prospect and industrial production potential.
It should be appreciated that the purposes of these embodiments is merely to illustrate the present invention and is not intended to limit protection model of the invention
Enclose.Additionally, it will also be appreciated that after technology contents of the invention have been read, those skilled in the art can make each to the present invention
Plant and change, change and/or modification, all these equivalent form of value equally falls within the guarantor that the application appended claims are limited
Within the scope of shield.
Claims (10)
1. a kind of preparation method of bacillus laterosporus microbial manure, methods described comprises the following steps：
S1：Bacillus laterosporus bacterial strain is activated, and aseptic water washing obtains seed liquor；
S2：Amplification cultivation is carried out to seed liquor using the first nutrient solution, the first amplification cultivation liquid is obtained；
S3：The first amplification cultivation liquid is cultivated using the second nutrient solution, obtains the second amplification cultivation liquid；
S4：Second amplification cultivation liquid carries out liquid fermentation and culture, obtains zymotic fluid；
S5：Zymotic fluid is drenched carries out solid fermentation in solid medium, and is post-processed, and obtains bacillus laterosporus fermentation
Microbial inoculum；
S6：Bacillus laterosporus fermenting agent and organic matter mixture are carried out into compost fermentation treatment, so as to obtain the side spore
Micro-organisms bacillus fertilizer.
2. preparation method as claimed in claim 1, it is characterised in that：The step S1 is specific as follows：
S1-1：Selection bacillus laterosporus bacterial strain, and be aseptically inoculated in activation culture is carried out on slant medium, obtain
To activated strains；
S1-2：Sterile water wash inclined-plane, obtains the seed liquor；
In step S1-1, the composition of slant medium is described in per 100g：Agar 5-6g, peptone 3-4g, yeast extract 0.02-
0.03g, analysis for soybean powder 2-3g, corn flour 0.8-1.4g, beef extract 3-4g, sodium chloride 0.2-0.5g, glucose 0.06-0.1g, it is remaining
It is sterilized water to measure, and adjusts pH value for 7-7.2.
3. the preparation method as described in claim any one of 1-2, it is characterised in that：The step S2 is specific as follows：
S2-1：Prepare the first nutrient solution
Following material is weighed respectively：Metallic element aqueous solution 25ml, vitamin E 1.5g, citric acid 0.15g, L-PROLINE
0.2mg, sucrose 2.5g, agar 8g, peptone 12g, yeast extract 1.4g, ammonium sulfate 1g, sodium phosphate 2.5g, beef extract 3.5g and
Sodium nitrate 4g；
Above-mentioned substance is added in deionized water, is sufficiently stirred for completely, and 1000ml is settled to deionized water, that is, obtain institute
State the first nutrient solution.
S2-2：According to 1:The volume ratio of 40-50, the seed liquor is inoculated in first nutrient solution, is led at 34-36 DEG C
Wind shaken cultivation 20-24 hours, so as to obtain the first amplification cultivation liquid.
4. preparation method as claimed in claim 3, it is characterised in that：In step S2-1, the metallic element aqueous solution be by
2.5g manganese sulfates, 3g zinc chloride, 4g magnesium sulfate, 1.5g cobalt chlorides, 2g copper nitrates, 3g calcium chloride, 4g frerrous chlorides, 3g iron chloride
It is completely dissolved in obtained from 1000ml distilled water.
5. the preparation method as described in claim any one of 3-4, it is characterised in that：It is described to be passed through in step S2-2
Gas in, oxygen volume content be 14-20%, preferably 16-18%, most preferably 17%.
6. the preparation method as described in claim any one of 1-5, it is characterised in that：The step S3 is specific as follows：
S3-1：Yeast extract consumption in first nutrient solution is halved, and the consumption increasing of the metallic element aqueous solution is twice, other
It is constant, so as to obtain second nutrient solution；
S3-2：According to 1:The volume ratio of 30-35, the first amplification cultivation liquid is connect and is added in second nutrient solution,
Air surge culture is passed through at 34-36 DEG C 14-16 hours, so as to obtain the second amplification cultivation liquid.
7. the preparation method as described in claim any one of 1-6, it is characterised in that：The step S4 is specific as follows：
The second amplification cultivation liquid is added in the liquid fermentation medium for 20-30 times of its volume carries out two-part intensification
Fermented and cultured, fermented and cultured total time is 10-12 hours, and period is passed through air always, and the ventilating ratio for being passed through air is 1:0.6-
1, so as to obtain the zymotic fluid；
Wherein, the liquid fermentation medium is in terms of every liter (L), comprising yeast extract 0.5g, peptone 2g, maltose 4g, Portugal
Grape sugar 1g, manganese sulfate 0.6g, sodium chloride 3.8g, analysis for soybean powder 7g, boric acid 1.5g, potassium nitrate 2.5g, urea 6g, lysine 0.7g and
Ammonium sulfate 1.5g, balance of sterilized water；
Wherein, the two-part fermentation at elevated temperatures culture refers to that fermentation temperature is in the first half time of fermented and cultured total time
32-34 DEG C, temperature is then increased to 38-40 DEG C in 10 minutes, and completes the fermentation of follow-up remaining time at such a temperature.
8. the preparation method as described in claim any one of 1-7, it is characterised in that：The step S5 is specific as follows：
According to mass ratio 1:The ratio of 10-14, the zymotic fluid is drenched in solid medium, adds appropriate amount of deionized water,
Make overall biodiversity content for 32-36%, then fully turn uniformly, load length for 3 meters, width are for 1.5 meters and deep
Spend in the fermentation tank for 1.5 meters, natural packing ferments 60-70 hours, period remains that overall biodiversity content is 32-
36%, and be passed through sterile wind to keep the central temperature inside fermentation tank is 56-60 DEG C；After fermentation ends, lead to heated-air drying, until
Biodiversity content is less than 5%, obtains fermenting agent.
9. the bacillus laterosporus microbial manure obtained in preparation method according to claim any one of 1-8.
10. purposes of the bacillus laterosporus microbial manure in agriculture field described in claim 9.
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| CN113046253A (en) * | 2021-04-14 | 2021-06-29 | 武汉新华扬生物股份有限公司 | Culture method for improving heat resistance of kluyveromyces marxianus |
| CN113046253B (en) * | 2021-04-14 | 2022-09-06 | 武汉新华扬生物股份有限公司 | Culture method for improving heat resistance of kluyveromyces marxianus |
| CN114717142A (en) * | 2022-03-09 | 2022-07-08 | 山东劲牛集团股份有限公司 | Preparation and application of streptomycete complex microbial inoculum |
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Application publication date: 20170613 |