CN105272406A - Microbial fertilizer prepared by bacillus laterosporus and preparation method of microbial fertilizer - Google Patents
Microbial fertilizer prepared by bacillus laterosporus and preparation method of microbial fertilizer Download PDFInfo
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- CN105272406A CN105272406A CN201510699985.6A CN201510699985A CN105272406A CN 105272406 A CN105272406 A CN 105272406A CN 201510699985 A CN201510699985 A CN 201510699985A CN 105272406 A CN105272406 A CN 105272406A
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Abstract
The invention relates to microbial fertilizer prepared by bacillus laterosporus and a preparation method of the microbial fertilizer. The fertilizer is prepared by culturing a microbial strain with a collection number of CGMCC (China General Microbiological Culture Collection Center) No. 5090 to form a culture, mixing the culture and grass carbon powder to form a microbial strain agent and mixing the microbial strain agent and conventional fertilizer. A culture method of the microbial strain culture comprises the steps of: 1) inoculating a volume of strain suspension to 20 volumes of No. 1 culture medium for aerobic culture for 8h at 30-38 DEG C, 2) adding a culture solution obtained in Step 1) into about 20 volumes of No. 2 culture medium for aerobic culture for 8h at 30-38 DEG C, and 3) adding a culture solution obtained in Step 2) into about 20 volumes of No. 3 culture medium for culture for 8h at 35-38 DEG C to produce the culture.
Description
Technical field
The present invention relates to a kind of microbial fertilizer prepared by new bacillus laterosporus with and preparation method thereof, belong to microbial fertilizer field.
Background technology
China has a large population, it is few to plough, and along with successively decreasing year by year in the development arable land in population growth, industry and city.For this reason, China increases the unique feasible approach of agricultural output is increase the yield per unit area.Could meet industrial development and the demand of population increase to agricultural-food, what chemical fertilizer also just followed a well mapped-out plan is widely applied.Unreasonable use chemical fertilizer brings for a long time
Soil compaction, soil fertility decline, ecological damage, environmental pollution and agricultural byproducts quality decline problem, be subject to the great attention of national departments concerned and agricultural science and technology circle.In recent decades, China makes great efforts to explore the rational application of fertilizer always, overcomes the effective way of the drawback of chemical fertilizer and raising chemical fertilizer utilization ratio.
Agricultural fertilizer comprises chemical fertilizer and bio-feritlizer, is the irreplaceable important production means of China's agriculture production.
Wherein, no matter the important branch of bio-feritlizer as agro-biological engineering subject and the important component part of soil fertilizer science, be obtained for and develop rapidly for nearly ten years in fundamental research and applied research.China's microbial fertilizer development is in recent years very fast, and have many enterprises to be engaged in production and operation, kind constantly increases, and also has some external products to squeeze into domestic market simultaneously.Due to reasons such as bacterial classifications, the quality product of existing market is uneven, production application has occurred some problems.According to Ministry of Agriculture's random checking recent years, follow the tracks of selective examination, the situation of country's system inspection is seen, selective examination qualification rate is less than 60%, and mainly effectively bacterium number is not up to standard for product quality problem, and the quality guaranteed period is too short.In microbial fertilizer, living bacteria count is the important indicator of quality product, living bacteria count is not up to standard, quality product just can not be guaranteed, what reflect is that effect is unstable, wherein reason mainly bacterial classification problem, bacterial classification is after being prepared as microbial strains agent, and the survival ability in preserving process declines, and the survival time shortens.This problem is actually the bacterial classification because these microbial strains agent use or can not produces gemma, the ability producing gemma is more weak, or the gemma produced sprouts ability, stress conditions in microbial strains agent production process and stock, preserving process can not be tided over very well, in production and application process, show as bacterial activity decline, production performance weakens, and reproduction speed is slack-off, and resistance is poor.Therefore, bacterial classification problem perplexs the prodution and development of China's microbial fertilizer always.
Chinese patent discloses the microbial strains that a kind of preserving number is CGMCCNo.5090, but be through experiment to find, cultural method defectiveness wherein, yield poorly, price is high, for improving output, the present inventor improves cultural method, the preserving number obtaining a kind of high yield is unexpectedly the microbial strains culture of CGMCCNo.5090, greatly reduces the price of fertilizer, is conducive to scale operation.
Summary of the invention
The object of this invention is to provide a kind of microbial fertilizer be prepared from by the microbial strains culture of CGMCCNo.5090 and preparation method thereof.
In order to achieve the above object, the invention provides following technical scheme:
Cultivate a method for the bacillus laterosporus (Bacilluslaterosporus) of high yield, described bacillus laterosporus preserving number is the microbial strains of CGMCCNo.5090.
This bacterial classification is named as " new side genus bacillus No. 1 ", and this strain separating is from high-yielding grain fields, and through domestication, proterties is stablized for a long time.
New side genus bacillus No. 1 bacterial strain submits the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms on July 22nd, 2010, deposit number is CGMCCNo.5090, biology name Bacilluslaterosporus, depositary institution address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica.
The biological characteristics of this bacterial classification is that thalline is shaft-like, belong to bacillus, its cell size is 0.15 ~ 0.18 × 215 ~ 510um, Gram-positive, can be changed into feminine gender, gemma is oval, and side is raw, middle life or near middle raw, sporangiocyst expands, free gemma thicker than the other side (canoe shape).Multiple nitrogenous source, carbon source can be utilized, growth temperature 20-55 DEG C, optimum growth temp 37 DEG C, optimal pH 7.2.Under bad condition, thalline transforms and produces gemma, and gemma, to poor environment strong stress resistance, can tolerate the mal-condition such as 100 DEG C of high temperature 20 minutes and drying.
The preservation inclined-plane of bacterial classification of the present invention is preserved, during cultivation first after inclined-plane is got kind, and line access slant medium, 30-38 DEG C of incubators are cultivated 18 hours under leaving standstill dark condition; Use aseptic water washing inclined-plane, form bacteria suspension, as original bacteria liquid.
Deposit number of the present invention is the microbial strains culture of CGMCCNo.5090, and its cultural method is: with above-mentioned bacteria suspension for original bacteria liquid, carry out amplification cultivation to bacterial classification, and to reach the object of productive culture thing, concrete cultural method step is as follows:
1) access in No. 1 substratum of 20 times of volumes by 1 volume bacteria suspension, 30-38 DEG C of aerobics cultivate 8 hours;
2) joined by the nutrient solution that step 1) obtains in No. 2 substratum of about 20 times of volumes, 30-38 DEG C of aerobics cultivate 8 hours;
3) by step 2) nutrient solution that obtains joins in No. 3 substratum of about 20 times of volumes, and cultivate 8 hours, obtain the culture that can produce for 35-38 DEG C.
The present invention further provides microbial strains agent, the preparation method of described inoculum agent is as follows:
Cool after peat composed of rotten mosses powder is carried out dry sterilization, join in the culture that the present invention obtains and fully stir, with granulator granulation, oven drying at low temperature obtains microbial strains agent.The granulation of grain machine, oven drying at low temperature obtains microbial strains agent, and wherein the part by weight of peat composed of rotten mosses powder and culture is 3-5:1.
Preferably, the part by weight of peat composed of rotten mosses powder and culture is 4:1.
Preparation method of the present invention, wherein the part by weight of microbial strains agent and common fertilizer is 1:9-11, and preferably, the part by weight of microbial strains agent and common fertilizer is 1:10.
The invention reside in the spawn culture method providing new, of the present inventionly focus on the formula improving substratum, the present invention obtains following substratum by adding sodium alginate:
Slant culture based formulas used in culturing process is: sodium alginate 5g/L, peptone 5g/L, sodium-chlor 5g/L, extractum carnis 5g/L, and agar 1.5%-2.0%, PH is adjusted to 6.5-7.8.
No. 1 culture medium prescription is: sodium alginate 5g/L, peptone 5g/L, sodium-chlor 5g/L, and extractum carnis 5g/L, PH is adjusted to 6.5-7.8.
No. 2 substratum: sodium alginate 5g/L, starch 5g/L, sucrose 1g/L, Yeast diffusion juice 0.2g/L, peptone 5g/L, extractum carnis 5g/L, ammonium sulfate 1g/L, potassium primary phosphate 0.5g/L, magnesium sulfate 0.2g/L, sodium-chlor 5g/L, PH is adjusted to 6.5-7.8.
No. 3 culture medium prescriptions are: sodium alginate 5g/L, starch 10g/L, sucrose 5g/L, Yeast diffusion juice 0.1g/L, peptone 0.5g/L, soybean cake powder 10g/L, ammonium sulfate 1g/L, potassium primary phosphate 0.5g/L, and sodium-chlor 5g/L, PH is adjusted to 6.5-7.8.
All substratum are all before use through high pressure steam sterilization (121 DEG C, 1.1kg/cm3,30 minutes) above, cooling.
Sodium alginate is a kind of natural polysaccharide, 1881, and first Englishize scholar E.C.Stanford carries out scientific research to the alginates extract in brown seaweed.He finds that this alignic extract has several very interesting characteristic, and it has concentrated solution, forms the ability of gel and film forming, can be used for foodstuffs industry and makes emulsion stabilizer and thickening material.
The present invention is by adding sodium alginate in the medium, and unexpected discovery incubation time shortens greatly, and culture efficiency, survival rate and the fertilizer efficiency of bacterial classification improve greatly.
Below for the parameter of cultural method of the present invention and prior art compares:
| Incubation time | Operation steps | 3 months bacterium survival rates deposited by the microbial inoculum made | Fertilizer efficiency is relative to the volume increase per-cent of blank | |
| Cultural method of the present invention | 24 hours | 3 steps | 100% | 30 |
| Prior art cultural method 1 | 61 hours | 4 steps | 90% | 20 |
| Prior art cultural method 2 | 62 hours | 3 steps | 85% | 15 |
Prior art cultural method 1 is the cultural method of embodiment 2 in Chinese patent 201110260431.8.
Prior art cultural method 2 is the cultural method of embodiment 1 in Chinese patent 201110260431.8.
Fungal counting method
Dilution plate number scale conveniently detects the viable bacteria content in microbial strains agent.Take microbial strains agent 1 gram (stock's two weeks), dissolve with sterilized water, being diluted to thalline number with sterilized water is 20-100/ml, then 1ml diluent is pipetted in sterile petri dish, add new 1 substratum (slant medium) in right amount, fully mix after coagulation, at 37 DEG C of insulation cans
Middle inversion cultivation 48 hours, the bacterium colony on statistics flat board, in triplicate, is multiplied by this gram of microbial strains dilution agent for milliliter number when cell concentration is 20-100/ml with mean value and is the strain number of every gram of microbial strains agent.
Fertilizer efficiency detection method:
The community fertilizer efficiency of being carried out by the microbial fertilizer of the present invention made comparing with prior art fertilizer and matrix fertilizer in Different Crop is tested, each plot area 0.16 mu.Each 25 kilograms of microbial fertilizer of the present invention and prior art fertilizer 1 and fertilizer 2 is executed respectively for every mu during experiment; Matrix fertilizer (mix with 1:2:3 weight percent via chicken manure, ground corn core, cinder, adjustment water content is after 30%, piles up make for 7-10 days in temperature under being greater than the condition of 30 DEG C) every mu executes 25 kilograms; All as base manure, once execute, do not added.Blank does not apply fertilizer.
Experimental result following (data are the volume increase per-cent of each fertilizer relative to blank)
Romaine lettuce sponge gourd corn, rice peanut cucumber
Microbial fertilizer 202530353536 of the present invention
Prior art microbial fertilizer 1142515141730
Prior art microbial fertilizer 2142515141730
Matrix fertilizer 1191091025
Result shows, microbial strains agent production-increasing function of the present invention is best.
Embodiment
Embodiment 1:
Cultural method of the present invention
Step 1, first slant culture activation is carried out to bacterial classification, then uses 5ml aseptic water washing inclined-plane, form bacteria suspension,
Step 2, bacteria suspension access 500ml triangular flask, this triangular flask is containing 100ml1 substratum, and use shaking table to cultivate 8 hours, culture condition is 37 DEG C, 180-200rpm,
Step 3, transfer in 5000ml triangular flask by the 105ml bacterium liquid in 500ml triangular flask, containing 2100ml2 substratum in bottle, use shaking table to cultivate 8 hours, culture condition is 37 DEG C, 180-200rpm;
Step 4, by 5000ml triangular flask 2205ml bacterium liquid access 60m
3in fermentor tank, containing 44.1L3 substratum in this fermentor tank, fermentation condition is: temperature 36-38 DEG C, tank pressure 0.5kg/cm, air flow 20m3/h, cultivates 8 hours; Obtain culture.
Detect cell concentration, concentration is 10
10the order of magnitude.
Then by 1:4 part by weight with carry out dry sterilization after the high-quality peat composed of rotten mosses powder (commercially available) that cools mix, then carry out granulation, oven drying at low temperature becomes microbial strains agent.
Namely the agent of gained microbial strains and common fertilizer mixing can be used for the production of farm crop, and blending ratio is.The part by weight of microbial strains agent and common fertilizer is 1:10.
Comparative example:
Prior art culture method, the cultural method of embodiment 2 in Chinese patent 201110260431.8.
First bacterial classification is activated, then use 5ml aseptic water washing inclined-plane, form bacteria suspension, bacteria suspension access 500ml triangular flask, this triangular flask is containing new 1 liquid nutrient medium of 100ml, and use shaking table to cultivate 18 hours, culture condition is 37 DEG C, 180-200rpm, detects cell concentration, reaches 10 in concentration
8the order of magnitude, carries out next step; Transfer in 2000ml triangular flask by the 100ml bacterium liquid in 500ml triangular flask, this triangular flask is containing new 2 substratum of 700ml, and use shaking table to cultivate 20 hours, culture condition is 37 DEG C, 180-200rpm; Detect cell concentration, reach 10 in concentration
8the order of magnitude, carries out next step; By in the bacterium liquid access 1m3 fermentor tank in 2000ml triangular flask, access amount is 6 liters, and containing new 3 substratum of 500L in this fermentor tank, fermentation condition is: temperature 36-38 DEG C, tank pressure 0.5kg/cm, air flow 20m3/h, cultivates 8 hours; Detect cell concentration, reach 10 in concentration
8the order of magnitude, carries out next step; By 1m
3bacterium liquid access 5m in fermentor tank
3fermentor tank, access amount is 500 liters, and this fermentor tank contains new 4 substratum of 3m3, and fermentation condition is: temperature 36-38 DEG C, tank pressure 0.5kg/cm, air flow 120m3/h, cultivates 15 hours.Detect cell concentration, reach 10 in concentration
9the order of magnitude, stops fermentation.Then in 1:4 ratio with carry out dry sterilization after the high-quality peat composed of rotten mosses powder (commercially available) that cools mix, then carry out granulation, oven drying at low temperature becomes microbial strains agent.
Claims (6)
1. the preparation method of a microbial fertilizer, the microbial strains that this fertilizer is CGMCCNo.5090 by preserving number obtains culture through cultivation, gained culture and peat composed of rotten mosses powder are mixed to get microbial strains agent, the agent of gained microbial strains is mixed with common fertilizer and forms, and the cultural method of wherein said microbial strains culture is as follows:
First the microbial strains of CGMCCNo.5090 is carried out slant culture activation, then use aseptic water washing inclined-plane, form bacteria suspension, take bacteria suspension as original bacteria liquid, carry out amplification cultivation, cultural method step is as follows:
1) access in No. 1 substratum of 20 times of volumes by 1 volume bacteria suspension, 30-38 DEG C of aerobics cultivate 8 hours;
2) joined by the nutrient solution that step 1) obtains in No. 2 substratum of about 20 times of volumes, 30-38 DEG C of aerobics cultivate 8 hours;
3) by step 2) nutrient solution that obtains joins in No. 3 substratum of about 20 times of volumes, and cultivate 8 hours, obtain the culture that can produce for 35-38 DEG C;
Wherein, described slant culture based formulas is: sodium alginate 5g/L, peptone 5g/L, sodium-chlor 5g/L, extractum carnis 5g/L, agar 1.5%-2.0%, and PH is adjusted to 6.5-7.8;
No. 1 culture medium prescription is: sodium alginate 5g/L, peptone 5g/L, sodium-chlor 5g/L, extractum carnis 5g/L, and PH is adjusted to 6.5-7.8;
No. 2 substratum: sodium alginate 5g/L, starch 5g/L, sucrose 1g/L, Yeast diffusion juice 0.2g/L, peptone 5g/L, extractum carnis 5g/L, ammonium sulfate 1g/L, potassium primary phosphate 0.5g/L, magnesium sulfate 0.2g/L, sodium-chlor 5g/L, PH is adjusted to 6.5-7.8;
No. 3 culture medium prescriptions are: sodium alginate 5g/L, starch 10g/L, sucrose 5g/L, Yeast diffusion juice 0.1g/L, peptone 0.5g/L, soybean cake powder 10g/L, ammonium sulfate 1g/L, potassium primary phosphate 0.5g/L, sodium-chlor 5g/L, and PH is adjusted to 6.5-7.8;
All substratum are all before use through high pressure steam sterilization (121 DEG C, 1.1kg/cm3,30 minutes) above, cooling.
2. preparation method according to claim 1, is characterized in that, step is as follows:
Step 1, first slant culture activation is carried out to bacterial classification, then uses 5ml aseptic water washing inclined-plane, form bacteria suspension,
Step 2, bacteria suspension access 500ml triangular flask, this triangular flask is containing 100ml1 substratum, and use shaking table to cultivate 8 hours, culture condition is 37 DEG C, 180-200rpm,
Step 3, transfer in 5000ml triangular flask by the 105ml bacterium liquid in 500ml triangular flask, containing 2100ml2 substratum in bottle, use shaking table to cultivate 8 hours, culture condition is 37 DEG C, 180-200rpm;
Step 4, by 5000ml triangular flask 2205ml bacterium liquid access 60m
3in fermentor tank, containing 44.1L3 substratum in this fermentor tank, fermentation condition is: temperature 36-38 DEG C, tank pressure 0.5kg/cm, air flow 20m3/h, cultivates 8 hours; Obtain culture.
3. preparation method according to claim 1, is characterized in that, the preparation method of wherein said microbial strains agent is as follows:
Cool after peat composed of rotten mosses powder is carried out dry sterilization, join in culture and fully stir, with granulator granulation, oven drying at low temperature obtains microbial strains agent, and wherein the part by weight of peat composed of rotten mosses powder and culture is 3-5:1.
4. preparation method according to claim 3, is characterized in that, wherein the part by weight of peat composed of rotten mosses powder and culture is 4:1.
5. preparation method according to claim 1, is characterized in that, wherein the part by weight of microbial strains agent and common fertilizer is 1:9-11.
6. preparation method according to claim 5, is characterized in that, wherein the part by weight of microbial strains agent and common fertilizer is 1:10.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108719334A (en) * | 2017-04-19 | 2018-11-02 | 柏玉兰 | The antimicrobial plant vaccine and production method of yellow twig can be prevented |
| CN109735465A (en) * | 2019-01-25 | 2019-05-10 | 江苏大学 | A kind of improvement beach ground microorganism formulation of plant growth and preparation method thereof |
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| CN101200385A (en) * | 2006-12-13 | 2008-06-18 | 四川艾蒙爱生物科技有限公司 | Preparation of composite microbial fertilizer by using bacilluslaterosporus and bacillus subtilis |
| CN102101800A (en) * | 2009-12-17 | 2011-06-22 | 天津市施易得肥料有限公司 | Method for preparing microbial organic fertilizer |
| CN102424626A (en) * | 2011-09-05 | 2012-04-25 | 麻林涛 | Microbial fertilizer prepared by bacillus laterosporus and preparation method thereof |
| CN103614448A (en) * | 2013-11-26 | 2014-03-05 | 燕山大学 | Method for preparing bioethanol by taking sodium alginate or algae as active ingredients |
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2015
- 2015-10-26 CN CN201510699985.6A patent/CN105272406A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101200385A (en) * | 2006-12-13 | 2008-06-18 | 四川艾蒙爱生物科技有限公司 | Preparation of composite microbial fertilizer by using bacilluslaterosporus and bacillus subtilis |
| CN102101800A (en) * | 2009-12-17 | 2011-06-22 | 天津市施易得肥料有限公司 | Method for preparing microbial organic fertilizer |
| CN102424626A (en) * | 2011-09-05 | 2012-04-25 | 麻林涛 | Microbial fertilizer prepared by bacillus laterosporus and preparation method thereof |
| CN103614448A (en) * | 2013-11-26 | 2014-03-05 | 燕山大学 | Method for preparing bioethanol by taking sodium alginate or algae as active ingredients |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108719334A (en) * | 2017-04-19 | 2018-11-02 | 柏玉兰 | The antimicrobial plant vaccine and production method of yellow twig can be prevented |
| CN109735465A (en) * | 2019-01-25 | 2019-05-10 | 江苏大学 | A kind of improvement beach ground microorganism formulation of plant growth and preparation method thereof |
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