CN106754459A - One plant of bacterial strain Bacillus altitudinis SEM 1 and its application from silkworm excrement - Google Patents

One plant of bacterial strain Bacillus altitudinis SEM 1 and its application from silkworm excrement Download PDF

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CN106754459A
CN106754459A CN201610915079.XA CN201610915079A CN106754459A CN 106754459 A CN106754459 A CN 106754459A CN 201610915079 A CN201610915079 A CN 201610915079A CN 106754459 A CN106754459 A CN 106754459A
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sem
bacillus
culture
highland
bacterial strain
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CN106754459B (en
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杨琼
李庆荣
廖森泰
支胡钰
邢东旭
叶明强
肖阳
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Guangdong Geolong Biotechnology Co ltd
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Abstract

The invention belongs to microbial technology field, and in particular to one plant of bacterial strain Bacillus altitudinis SEM 1 and its application from silkworm excrement.The bacterial strain is preserved in Guangdong Province's Culture Collection on May 16th, 2016, and deposit number is GDMCC NO.60038.The bacterial strains of Bacillus altitudinis SEM 1 of the invention can efficiently suppress the growth of the harmful pathogenic microorganism of crop in the soil such as sickle-like bacteria (Fusarium), Ralstonia solanacearum (Ralstonia dolaanacearum), play a part of to prevent and treat crop disease and improve crop disease-resistant insect pest ability, so as to improve the yield and quality of crop.

Description

One plant from silkworm excrement bacterial strain Bacillus altitudinis SEM-1 and its Using
Technical field
The invention belongs to microbial technology field, and in particular to one plant of bacterial strain Bacillus from silkworm excrement Altitudinis SEM-1 and its application.
Background technology
Soil-borne disease is the general designation of the plant disease propagated as medium with soil, refers to the disease lived in the middle of soil The disease that substance causes when condition is suitable from root or stem infringement crop, common local products disease has：Capsicum, eggplant, Cucumber root rot, damping-off, samping off, epidemic disease, yellow blight, tomato, the bacterial wilt of capsicum, damping-off, nematodiasis of cotton etc., Sclerotiniose of red disease of withering, verticillium wilt, take-all, soft rot of Chinese cabbage, rape and lettuce etc..
The root of soil-borne disease main harm crop and stem.Once there is disease, seedling root rot in plant growth early stage Or stem rot is rotten, seedling will soon be dead；Plant growth deuterogenesis's disease, the underproduction for causing can according to order of severity difference Up to 20%-60%, or even total crop failure.After soil-borne disease occurs, relatively more difficult preventing and treating, germ is ensconced and is survived the winter in soil, it is difficult to be killed, The coming year continues to encroach on crop, so circulation, and disease is increasingly severe.
The prevention and control approach of soil-borne disease mainly has shift of crops, disease-resistant variety, medical treatment and biological control etc..Crop is taken turns Work can to a certain degree control various soil-borne diseases, but because most of pathogen can survive a very long time in soil, Once disease occurs, the validity of crop rotation can be caused to be restricted；Disease-resistant variety is with strong points, and overall preventive effect is not high；And it is long Phase can destroy the ecological balance using chemical agent, cause the problems such as rampant Minor diseases, medicament residual, environmental pollution.
Biological control is mainly and is colonized by antagonistic microbe soil, and nutrition and living space, secretion are competed with pathogen The comprehensive function shape such as antibacterial substance and protease, the resistance immunologic mechanism for exciting plant, promotion plant growth, reinforcing resistance The prevention of paired soil-borne disease and prevention effect.People are right for the attention of food security and the enhancing of environmental consciousness in recent years The sustainable development of agricultural proposes requirement higher, and biological control is because of its low cost, high efficiency, environment-friendly and residual without medicine The features such as staying has turned into the study hotspot of current control of plant disease both at home and abroad.
Silkworm excrement is the main organic waste in producing of breeding silkworms, not only containing abundant nutriment, also containing abundant Microorganism species, such as fixed nitrogen, phosphorus decomposing and potassium decomposing and its microorganism species of cause of disease antagonism.Therefore how separation screening is simultaneously effective Using microbial resources therein, as the focal point that this is studied.
The content of the invention
Primary and foremost purpose of the invention is to provide one plant of bacterial strain Bacillus altitudinis SEM- from silkworm excrement 1。
Soil is being prepared another object of the present invention is to provide above-mentioned bacterial strains Bacillus altitudinis SEM-1 Application in microbial bacterial agent or biological organic fertilizer.
The object of the invention is achieved through the following technical solutions：
One plant of bacterial strain Bacillus altitudinis (highland bacillus) SEM-1 from silkworm excrement, in 2016 On May 16, in is preserved in Guangdong Province's Culture Collection, and deposit number is GDMCC NO.60038.
Above-mentioned highland bacillus are obtained after being isolated and purified from silkworm excrement, and its proterties is as follows：
Colonial morphology：After highland bacillus suspension of the invention is through coated plate and NA culture mediums, in 30 DEG C of biochemical cultivation cases Culture 2 days, observes its colony morphology characteristic, as a result as shown in Figure 1.As seen from Figure 1, Bacillus colonies of the invention are milky white Color, circular, surface is smooth, translucent, neat in edge.
Highland bacillus of the invention are inoculated in NB culture mediums in 1% ratio, after cultivating 18h in 30 DEG C of shaking tables, are taken Suspension, point sample to slide, roasting piece, through Gram's staining, oil mirror observes the morphological feature of bacterial strain, as a result as shown in Figure 2.By scheming 2 is visible, and highland bacillus of the invention are gram-positive bacteria, raw in gemma, is oval, and thalline is shaft-like.
The 16srDNA identifications of bacillus of the present invention：Strain suspensions are template, 16srDNA universal primer 27F/1492R, PCR reaction systems are 50 μ L, and reaction condition is：94 DEG C of predegeneration 5min；94 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 2min, circulation 30 It is secondary；72 DEG C of extension 10min.PCR primer sample presentation sequencing after purification.
The part variable region sequences sequencing result of the 16srDNA of highland bacillus SEM-1 of the present invention passes through NCBI/ Blast has found and highland bacillus strain HN-8 homology highests after comparing, and is highland bacillus up to more than 99%.
Above-mentioned bacterial strains Bacillus altitudinis SEM-1 are in edaphon microbial inoculum or biological organic fertilizer is prepared Application.
Above-mentioned bacterial strains Bacillus altitudinis SEM-1 are used to prepare highland bacillus original powder, including following system Standby step：
(1) activated spawn
Activation highland bacillus (Bacillus altitudinis) SEM-1 bacterial strains, from inclined-plane picking colony and The line culture of NA solid mediums, 32 DEG C of condition of culture, after SEM-1 is in NA cultured on solid medium 18-24h, picking list Bacterium colony, for the preparation of primary seed solution；
(2) preparation of primary seed solution
The single bacterium colony on NA solid mediums is chosen, is inoculated in first order seed shaking flask, Flask volume is 250mL, one-level The liquid amount 100ml of seed NB liquid culture mediums (at 121 DEG C, sterilize 20min), by first order seed shaking flask in 32 DEG C, 200r/ 12-14h is cultivated under min, primary seed solution is obtained；
(3) preparation of secondary seed solution
Culture medium in seeding tank is still NB fluid nutrient mediums, and at 121 DEG C, sterilize 20min, primary seed solution is accessed and is planted In sub- tank, inoculum concentration is 0.5~1% (V/V)；Condition of culture is：Ventilation ratio 1:1, speed of agitator is 250~400r/min, training 32 DEG C of temperature is supported, tank pressure 0.05MPa, incubation time is 10 hours, and secondary seed solution is obtained at the end of culture, and its viable bacteria content is 1~3 × 107, 0.7~0.8 or so, now spawn activity is strong for cfu/mL, OD value；
(4) the bacterium solution fermentation of highland bacillus SEM-1
Fermentative medium formula (percentage by weight)：Cornstarch 2-3%g；Bean cake powder 1.5-2.5%；Yeast extract 0.5~ 1%, CaCO30.3-0.6%, K2HPO40.02-0.03%, MgSO4·7H2O 0.015-0.025%, MnSO4·H2O 0.015-0.025%, (NH4)2SO40.05-0.15%；PH6.5-7.5,121 DEG C of sterilizing 30min；
It is that 0.5~1% (V/V) accesses fermentation tank according to inoculum concentration by the secondary seed solution of highland bacillus SEM-1 In, condition of culture is：Inoculum concentration is 0.5-1% (V/V), and condition of culture is：Ventilation ratio 1:(1-1.5), speed of agitator is 200- 400r/min, 32 DEG C of cultivation temperature, tank pressure 0.05MPa keeps more than 5% dissolved oxygen condition, and incubation time is 48h or so；Under The tank time is defined by sampling the release of microscopy gemma more than 90%, highland bacillus SEM-1 living spores contents in maturing fermentation liquid It is 50~80 × 108Cfu/mL, obtains fermenting-ripening microbial inoculum；
(5) preparation of highland bacillus original powder
(living spores content is 50~80 × 10 to fermenting-ripening microbial inoculum obtained by step (4)8Cfu/mL) through disk plate centrifuge Or after microfiltration equipment concentration, precipitated calcium carbonate being added, the ripe fermenting agent of concentration and the weight ratio of precipitated calcium carbonate are 100:3, Liquid-solid mixture is obtained, after the liquid-solid mixture is stirred into 30min, liquid-solid mixture atomizer is sent into using dehvery pump Dried in tower, obtain highland bacillus SEM-1 original powder.
Highland bacillus (Bacillus altitudinis) SEM-1 of the invention has the following advantages that and beneficial effect Really：
(1) highland bacillus SEM-1 bacterium sources of the invention, in silkworm excrement, are the spontaneous bacterium in silkworm excrement composting process, Conformability with silkworm excrement organic fertilizer is good；
(2) highland bacillus SEM-1 bacterial strains heat-resisting quantity of the invention is good, is cultivated under 60 degree of high temperature and transported To survive, there is suitable advantage in terms of edaphon microbial inoculum preparation；
(3) highland bacillus SEM-1 bacterial strains Saline alkali tolerance of the invention is good, can be in 10% NaCl concentration and PH5- Well-grown in the range of 10, has suitable advantage in terms of alkaline land improving；
(4) highland bacillus SEM-1 bacterial strains of the invention can efficiently suppress sickle-like bacteria (Fusarium), Ralstonia solanacearum The growth of the harmful pathogenic microorganism of crop in soil such as (Ralstonia dolaanacearum), play prevent and treat crop disease and The effect of crop disease-resistant insect pest ability is improved, so as to improve the yield and quality of crop；
(5) highland bacillus SEM-1 bacterial strains of the invention can be prepared into soil with efficient-decomposition soil mineral potassium Microbial bacterial agent or after being added to silkworm excrement organic fertilizer, with significant potassium decomposing effect, increases soil fertility, and the bacterial strain can be used to make Standby single microorganism microbial inoculum, complex micro organism fungicide, biological organic fertilizer etc..
Brief description of the drawings
Fig. 1 is the colony morphology characteristic figure of highland bacillus of the present invention (Bacillus altitudinis) SEM-1；
Fig. 2 is the strain morphology characteristic pattern of highland bacillus of the present invention (Bacillus altitudinis) SEM-1；
Fig. 3 is embodiment of the present invention gained highland bacillus (Bacillus altitudinis) SEM-1 to four kinds of sickles The antagonistic experiment design sketch of knife bacterium.
Specific embodiment
With reference to embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention are not limited In this.
Embodiment 1
The separation and identification of highland bacillus (Bacillus altitudinis) SEM-1 bacterial strains
(1) Sample Dilution：Sampled from the silkworm excrement compost of fermentation maturity, weigh 5g and be dissolved into equipped with 45ml sterilized waters three In the bottle of angle, 150rp oscillators vibrate into uniform 10-1Dilution, then draws dilution 1ml to being equipped with 1ml liquid-transfering guns In the test tube of 9ml sterilized waters, fully shake up, that is, be made 10-2Sample diluting liquid, and be diluted to 10 successively-6；
(2) culture medium prepares：
Potassium decomposing screening and culturing medium：Glucose 10.0g, NH4NO33.0g, NaH2PO41.0g, FeSO4·7H2O1.0g, MgSO4·7H2O 1.0g, potassium feldspar 1.5g, distilled water 1000ml, agar 20.0g, pH 7.0~7.4, sterilize 20min；
Potato potato culture medium：200 grams of potato, 20 grams of glucose, 18 grams of agar, 1000 milliliters of distilled water, pH 6.8~7.2；
NA culture mediums：Peptone 10g, beef extract 5g, NaCl5g, agar powder 20g, distilled water 1000ml, PH7.0；121℃ Sterilizing 20min, aseptic culture dish is poured onto in 45 DEG C -55 DEG C；
(3) strain separating, purifying and screening：The dilution 0.1ml of different extension rates is drawn respectively, is spread evenly across cold Good potassium decomposing culture medium, place 30 DEG C of incubators in be inverted culture 5 days, observe and select grow fine, potassium decomposing circle it is significant Bacterium colony is in NA medium cultures；By picked out bacterium in after the multiple squamous subculture of NA culture mediums, stood facing each other method by flat board, detection with Sickle-like bacteria, the antagonistic effect of Ralstonia solanacearum, take the significant bacterial strain of antagonistic effect, and name bacterial strain SEM-1；
(5) identification of bacterial strain
A, Morphological Identification：The colonial morphology of highland bacillus (Bacillus altitudinis) SEM-1 bacterial strains is breast White, circular, surface is smooth, semi-opaque, neat in edge.Gram's staining result shows that highland bacillus SEM-1 is leather Lan Shi positive bacterias, bud is satisfied oval, middle life, and thalline is shaft-like.
B, molecular biology identification：The 16S rDNA serial analysises of SEM-1, particular sequence is shown in sequence table.
(6) bacterial strain SEM-1 expert's conclusions
Morphology and 16rDNA gene serial analysises according to bacterial strain SEM-1, identify that SEM-1 is highland bacillus (Bacillus altitudinis), is preserved in Guangdong Province's Culture Collection on May 16th, 2016, and preservation is compiled Number be GDMCC NO.60038.Preservation address：5 building, the building of compound the 59th of GuangZhou, China city martyr Road 100, postcode is 510075。
The present embodiment gained highland bacillus (Bacillus altitudinis) SEM-1 potassium decomposing effect measurings：
Using atomic absorption spectrophotometry 766nm determine fermented liquid supernatant in effectively potassium content.With compare bacterium (potassium decomposing Bacterium, purchased from Guangzhou infinitesimal Bioisystech Co., Ltd) compare, highland bacillus (Bacillus of the invention Altitudinis) SEM-1 potassium decomposings effect is significant compares bacterium higher than potassium decomposing.
The present embodiment gained highland bacillus (Bacillus altitudinis) SEM-1 antagonism sickle-like bacteria effect is surveyed It is fixed：
Using flat board face-off method, 6 kinds of sickle-like bacteria (Fusarium graminearums, Fusarium are inoculated with respectively in flat board center sambucinum；Double fusarium oxysporums, Fusarium catenulatum；Fusarium oxysporum, Fusarium oxysporum；Layer goes out Sickle-like bacteria, Fusarium proliferatum；Fusarium solani, Fusarium equiseti；Fusarium solani, Fusarium Equiseti), filter paper is placed at its about 1-1.5cm, (removal surfactant, suds immersion, rinses well, dries It is dry, sterilizing), the zymotic fluid of point 5-6ul highland bacillus SEM-1 after fully absorbing, is inverted culture 2-3 days, observation, knot Fruit shows：Highland bacillus SEM-1 has the antagonistic effect of highly significant to surveying six kinds of sickle-like bacteria (Fusarium).Fig. 3 is arranged SEM-1 is gone out to the wherein four kinds antagonistic experiment design sketch of sickle-like bacteria.
Embodiment 2
The preparation of highland bacillus (Bacillus altitudinis) SEM-1 microbial inoculums original powder
(1) activated spawn
Activation highland bacillus (Bacillus altitudinis) SEM-1 bacterial strains, from inclined-plane picking colony and The line culture of NA solid mediums, 32 DEG C of condition of culture, after SEM-1 is in NA cultured on solid medium 18-24h, picking list Bacterium colony, for the preparation of primary seed solution；
(2) preparation of primary seed solution
The single bacterium colony on NA solid mediums is chosen, is inoculated in first order seed shaking flask, Flask volume is 250mL, one-level The liquid amount 100ml of seed NB liquid culture mediums (at 121 DEG C, sterilize 20min), by first order seed shaking flask in 32 DEG C, 200r/ 12h is cultivated under min, primary seed solution is obtained；
(3) preparation of secondary seed solution
Culture medium in seeding tank is still NB fluid nutrient mediums, and at 121 DEG C, sterilize 20min, primary seed solution is accessed and is planted In sub- tank, inoculum concentration is 0.5~1% (V/V)；Condition of culture is：Ventilation ratio 1:1, speed of agitator is 250~400r/min, training 32 DEG C of temperature is supported, tank pressure 0.05MPa, incubation time is 10 hours, and secondary seed solution is obtained at the end of culture, and its viable bacteria content is 1~3 × 107, 0.7~0.8 or so, now spawn activity is strong for cfu/mL, OD value；
(4) the bacterium solution fermentation of highland bacillus SEM-1
Fermentative medium formula (percentage by weight)：Cornstarch 2-3%g；Bean cake powder 1.5-2.5%；Yeast extract 0.5~ 1%, CaCO30.3-0.6%, K2HPO40.02-0.03%, MgSO4·7H2O 0.015-0.025%, MnSO4·H2O 0.015-0.025%, (NH4)2SO40.05-0.15%；PH6.5-7.5,121 DEG C of sterilizing 30min；
It is that 0.5~1% (V/V) accesses fermentation tank according to inoculum concentration by the secondary seed solution of highland bacillus SEM-1 In, condition of culture is：Inoculum concentration is 0.5-1% (V/V), and condition of culture is：Ventilation ratio 1:(1-1.5), speed of agitator is 200- 400r/min, 32 DEG C of cultivation temperature, tank pressure 0.05MPa keeps more than 5% dissolved oxygen condition, and incubation time is 48h or so；Under The tank time is defined by sampling the release of microscopy gemma more than 90%, highland bacillus SEM-1 living spores contents in maturing fermentation liquid It is 50~80 × 108Cfu/mL, obtains fermenting-ripening microbial inoculum；
(5) preparation of highland bacillus original powder
(living spores content is 50~80 × 10 to fermenting-ripening microbial inoculum obtained by step (4)8Cfu/mL) through disk plate centrifuge Or after microfiltration equipment concentration, precipitated calcium carbonate being added, the ripe fermenting agent of concentration and the weight ratio of precipitated calcium carbonate are 100:3, Liquid-solid mixture is obtained, after the liquid-solid mixture is stirred into 30min, liquid-solid mixture atomizer is sent into using dehvery pump Dried in tower, obtain highland bacillus SEM-1 original powder.
Viable bacteria concentration is 5~800,000,000/gram dry powder in gained highland bacillus SEM-1 original powder.
Embodiment 3
The preparation of one kind addition highland bacillus (Bacillus altitudinis) SEM-1 silkworm excrement microbial-bacterial fertilizers.
The biological organic fertilizer is by the silkworm excrement after silkworm excrement or compost maturityization treatment, highland bacillus SEM-1 microbial inoculums and shellfish Shell powder is mixed with and forms.
The dry powder of highland bacillus SEM-1 is obtained with reference to the method for embodiment 2, wherein viable bacteria concentration is in 5-8 hundred million/gram dry Powder, is prepared into microbial bacterial agent.By microbial inoculum in proportion with compost maturity after silkworm excrement and oyster shell whiting mix, obtain silkworm excrement microorganism Bacterial manure.The weight/mass percentage composition of silkworm excrement, microbial bacterial agent and oyster shell whiting after the silkworm excrement or compost maturityization treatment is respectively 70%~90%, 1%~10% and 10%~30%.
Above-described embodiment is the present invention preferably implementation method, but embodiments of the present invention are not by above-described embodiment Limitation, it is other it is any without departing from Spirit Essence of the invention and the change, modification, replacement made under principle, combine, simplification, Equivalent substitute mode is should be, is included within protection scope of the present invention.

Claims (3)

1. one plant of bacterial strain Bacillus altitudinis SEM-1 from silkworm excrement, is preserved on May 16th, 2016 Guangdong Province's Culture Collection, deposit number is GDMCC NO.60038.
2. the bacterial strain Bacillus altitudinis SEM-1 described in claim 1 are preparing edaphon microbial inoculum or biology Application in organic fertilizer.
3. application according to claim 2, it is characterised in that：The bacterial strain Bacillus altitudinis SEM-1 are used In preparation highland bacillus original powder, including following preparation process：
(1) activated spawn
Activated b acillus altitudinis SEM-1 bacterial strains, from inclined-plane picking colony and NA solid mediums rule train Support, 32 DEG C of condition of culture, after SEM-1 is in NA cultured on solid medium 18-24h, picking single bacterium colony, for primary seed solution Preparation；
(2) preparation of primary seed solution
The single bacterium colony on NA solid mediums is chosen, is inoculated in first order seed shaking flask, first order seed shaking flask is trained in NB liquid Support base, 32 DEG C, cultivate 12-14h under 200r/min, obtain primary seed solution；
(3) preparation of secondary seed solution
Culture medium in seeding tank is still NB fluid nutrient mediums, and at 121 DEG C, sterilized 20min, and primary seed solution is accessed into seeding tank In, inoculum concentration volume ratio is 0.5~1%；Condition of culture is：Ventilation ratio 1:1, speed of agitator is 250~400r/min, culture temperature 32 DEG C of degree, tank pressure 0.05MPa, incubation time is 10 hours, and secondary seed solution is obtained at the end of culture, and its viable bacteria content is 1~3 ×107Cfu/mL, OD value are 0.7~0.8；
(4) the bacterium solution fermentation of highland bacillus SEM-1
Fermentative medium formula is by weight percentage：Cornstarch 2-3%, bean cake powder 1.5-2.5%, yeast extract 0.5~1%, CaCO30.3-0.6%, K2HPO40.02-0.03%, MgSO4·7H2O 0.015-0.025%, MnSO4·H2O 0.015- 0.025%, (NH4)2SO40.05-0.15%；PH 6.5-7.5,121 DEG C of sterilizing 30min；
It is 0.5~1% to access fermentation tank according to inoculum concentration volume ratio by the secondary seed solution of highland bacillus SEM-1, training Foster condition is：Ventilation ratio 1:(1-1.5), speed of agitator is 200-400r/min, and 32 DEG C of cultivation temperature, tank pressure 0.05MPa keeps More than 5% dissolved oxygen condition, incubation time is 48h；The lower tank time will be defined by sampling the release of microscopy gemma more than 90%, maturation hair Highland bacillus SEM-1 living spores contents are 50~80 × 10 in zymotic fluid8Cfu/mL, obtains fermenting-ripening microbial inoculum；
(5) preparation of highland bacillus original powder
After fermenting-ripening microbial inoculum obtained by step (4) is concentrated through disk plate centrifuge or microfiltration equipment, precipitated calcium carbonate, fermentation are added Ripe microbial inoculum is 100 with the weight ratio of precipitated calcium carbonate:3, liquid-solid mixture is obtained, after the liquid-solid mixture is stirred into 30min, Dried during liquid-solid mixture is sent into atomizer tower using dehvery pump, obtain highland bacillus SEM-1 original powder.
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Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107151639A (en) * 2017-06-06 2017-09-12 广东植物龙生物技术股份有限公司 One plant of bacterial strain Bacillus subtilis SEM 9 and its application from silkworm excrement
CN109504622A (en) * 2018-11-16 2019-03-22 广东植物龙生物技术股份有限公司 A kind of lysine bacillus and the composite bacteria agent of bacillus subtilis and preparation method thereof
CN110066754A (en) * 2019-05-14 2019-07-30 西南林业大学 A kind of highland bacillus bacterial strain and its application
CN111066790A (en) * 2019-11-06 2020-04-28 广东植物龙生物技术股份有限公司 Plant growth regulator combining diethyl aminoethyl hexanoate and bacillus altitudinis fungus-containing metabolites
CN111100818A (en) * 2020-01-10 2020-05-05 新疆根力多生物科技有限公司 Geobacillus altitudinis SWY137 and application thereof
CN112322518A (en) * 2020-10-15 2021-02-05 华南农业大学 Bacillus alpinus with potassium-dissolving effect and application thereof
CN112592845A (en) * 2020-11-19 2021-04-02 仲恺农业工程学院 Broad-spectrum antifungal highland bacillus ZKY02 and application thereof

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102703350A (en) * 2012-05-29 2012-10-03 北京大学 Application of salt-tolerant nitrogen and phosphorus removing bacillus alitudinis to wastewater treatment
CN104342395A (en) * 2014-10-31 2015-02-11 中国农业科学院农产品加工研究所 Bacillus altitudinis and application thereof to prevention and control of fusarium graminearum
CN105820981A (en) * 2016-04-27 2016-08-03 山东农业大学 Preparation and application of bacillus altitudinis bacterial agent

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102703350A (en) * 2012-05-29 2012-10-03 北京大学 Application of salt-tolerant nitrogen and phosphorus removing bacillus alitudinis to wastewater treatment
CN104342395A (en) * 2014-10-31 2015-02-11 中国农业科学院农产品加工研究所 Bacillus altitudinis and application thereof to prevention and control of fusarium graminearum
CN105820981A (en) * 2016-04-27 2016-08-03 山东农业大学 Preparation and application of bacillus altitudinis bacterial agent

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
KHALED ELBANNA 等: "Characterization of Bacillus altitudinis as a New Causative Agent of Bacterial Soft Rot", 《J PHYTOPATHOL》 *

Cited By (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107151639A (en) * 2017-06-06 2017-09-12 广东植物龙生物技术股份有限公司 One plant of bacterial strain Bacillus subtilis SEM 9 and its application from silkworm excrement
CN107151639B (en) * 2017-06-06 2020-11-13 广东植物龙生物技术股份有限公司 Strain Bacillus subtilis SEM-9 derived from silkworm excrement and application thereof
CN109504622A (en) * 2018-11-16 2019-03-22 广东植物龙生物技术股份有限公司 A kind of lysine bacillus and the composite bacteria agent of bacillus subtilis and preparation method thereof
CN109504622B (en) * 2018-11-16 2022-01-07 广东植物龙生物技术股份有限公司 Composite microbial inoculum of lysine bacillus and bacillus subtilis and preparation method thereof
CN110066754A (en) * 2019-05-14 2019-07-30 西南林业大学 A kind of highland bacillus bacterial strain and its application
CN111066790A (en) * 2019-11-06 2020-04-28 广东植物龙生物技术股份有限公司 Plant growth regulator combining diethyl aminoethyl hexanoate and bacillus altitudinis fungus-containing metabolites
CN111100818A (en) * 2020-01-10 2020-05-05 新疆根力多生物科技有限公司 Geobacillus altitudinis SWY137 and application thereof
CN111100818B (en) * 2020-01-10 2023-05-12 新疆根力多生物科技有限公司 Geobacillus altitudinalis SWY137 and application thereof
CN112322518A (en) * 2020-10-15 2021-02-05 华南农业大学 Bacillus alpinus with potassium-dissolving effect and application thereof
CN112592845A (en) * 2020-11-19 2021-04-02 仲恺农业工程学院 Broad-spectrum antifungal highland bacillus ZKY02 and application thereof
CN112592845B (en) * 2020-11-19 2022-09-23 仲恺农业工程学院 Broad-spectrum antifungal highland bacillus ZKY02 and application thereof

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