CN106967615A - One plant of gibberella and its application - Google Patents

One plant of gibberella and its application Download PDF

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Publication number
CN106967615A
CN106967615A CN201710147806.7A CN201710147806A CN106967615A CN 106967615 A CN106967615 A CN 106967615A CN 201710147806 A CN201710147806 A CN 201710147806A CN 106967615 A CN106967615 A CN 106967615A
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CN
China
Prior art keywords
bacterial strain
qjp2
inoculated
gibberella
mass ratio
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Pending
Application number
CN201710147806.7A
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Chinese (zh)
Inventor
杜丰玉
肖�琳
周远明
牛赡光
王学军
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Qingdao Nuubel Biological Engineering Co ltd
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Qingdao Nuubel Biological Engineering Co ltd
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Priority to CN201710147806.7A priority Critical patent/CN106967615A/en
Publication of CN106967615A publication Critical patent/CN106967615A/en
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/645—Fungi ; Processes using fungi
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
    • C12N1/145—Fungal isolates
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K17/00—Soil-conditioning materials or soil-stabilising materials
    • C09K17/14—Soil-conditioning materials or soil-stabilising materials containing organic compounds only
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K2101/00—Agricultural use

Abstract

The invention discloses one plantGibberella intermediaAnd its application, belong to phosphate solubilizing microorganism technical field of fertilizers.The bacterial strain has been preserved in China General Microbiological culture presevation administrative center, preservation date at present：On December 09th, 2016, deposit number is CGMCC No. 13199.The invention also discloses microbial bacterial agent containing the bacterial strain and preparation method thereof.Experiment shows, the characteristics of bacterial strain has fast growth, dissolving P capacity is strong.Microbial bacterial agent prepared by the bacterial strain can improve phosphate fertilizer utilization efficiency, improve soil texture, and then reach the effect of crop yield.

Description

One plant of gibberella and its application
Technical field
The invention belongs to phosphate solubilizing microorganism fertilizer field.Specifically, the present invention relates to one plantGibberella intermediaAnd application thereof.
Background technology
P elements are essential nutrients in plant growth metabolic process.According to statistics, the arable land soil of China about 75% The phosphorus that earth lacks in phosphorus, and soil more than 90% is difficult to directly be absorbed by plant.Meanwhile, there is utilization rate in the phosphate fertilizer after Relatively low situation, traces it to its cause as Ca in quite a few phosphorus and soil2+、Fe2+、Fe3+With A13+Combined to form Deng metal ion Insoluble phosphate and be difficult to be absorbed by crops utilization.Insoluble phosphate in soil is only in chemical factors and microorganism Under effect, mineralizer, which is broken up and decomposed, progressively discharging P elements, plant growth could be supplied to utilize.Numerous studies show, native Microorganism in earth can improve rhizosphere soil environment, promote the release of titanium pigment in soil, so as to significantly improve soil fertilizer Power.
Gibberella intermediaBelong to for gibberella, domestic correlative study report is less.And phosphorus decomposing fungi is in quantity With, far away from phosphate-solubilizing bacteria, but it is reported that the dissolving P capacity of phosphorus decomposing fungi is typically better than bacterium in species.Fungi phosphorus decomposing mechanism compared with For complexity, main mechanism includes generation, the release of proton and biosynthesis of acid phosphatase of organic acid etc., so that by soil In phosphorus be converted into the form that plant available is utilized.Phosphate solubilizing microorganism microbial inoculum is developed, can not only be made full use of solid in soil Stationary state or the phosphorus of ADSORPTION STATE；And to improving phosphate fertilizer utilization efficiency, reduce phosphate fertilizer and caused environmental pollution, Jin Erti is excessively used High crop yield and quality, all have important practical significance.
The content of the invention
The purpose of the present invention is a kind of phosphorus decomposing fungi of offer ---Gibberella intermediaQJP2.The bacterial strain With fast growth, dissolving P capacity is strong the characteristics of, soil texture can be improved, absorption of the enhancing crop to fertilizer, with good Good application prospect.
Another object of the present invention is a kind of by the bacterial strain to provideGibberella intermediaIt is prepared by QJP2 Phosphate solubilizing microorganism microbial inoculum.
The above-mentioned purpose of the present invention is achieved through the following technical solutions：
Bacterial strain provided by the present invention isGibberella intermediaQJP2, is located away from the fluffy wetland of Jiaozhou Bay of Qingdao alkali and adopts The alkaline land soil sample of collection, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center（Referred to as CGMCC, address：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica）, preservation date：2016 On December 09, in, deposit number is CGMCC No. 13199.
It is provided by the present inventionGibberella intermediaQJP2 cultural methods or propagation method include：
（1）Nostoc commune Vanch preserves and uses PDA culture medium, is formulated：The g of potato 200, the g of glucose 20, the g of agar 12, distilled water 1000 mL, pH are adjusted to 7.0；
（2）Prepared by seed liquor uses PDB culture mediums, formula：The g of potato 200, the g of glucose 20, distilled water 1000 mL, pH tune To 7.0；
（3）Solid medium, formula：Gu material and inorganic salt solution in mass ratio 1:1.8 mixing；The solid material is mass ratio 85:15 rice chaff and wheat bran；The inorganic salt solution is 3.5% potassium dihydrogen phosphate, 0.04% magnesium sulfate and 4% ammonium sulfate；
（4）The same solid medium of scale fermentation culture medium.
Phosphate solubilizing microorganism bacterial preparation process provided by the present invention includes：
（1）By the bacterial strain activatedGibberella intermediaQJP2 is inoculated into the triangular flask equipped with PDB culture mediums, 30 DEG C, 120 r/min shaken cultivation 3-5 d；
（2）By step（1）The seed liquor prepared in mass ratio 10% is inoculated into solid medium, and 28-30 DEG C incubated 5-7 d；
（3）By step（2）The culture prepared and sterilized water in mass ratio 1:15 mixing, filtering.By filtrate by volume 1:6 are inoculated into scale fermentation culture medium, the scale fermentation 8-10 d in 28-30 DEG C, the fermenting cellar of relative humidity more than 80%.
Experiment shows, the characteristics of bacterial strain has fast growth, dissolving P capacity is strong.The zymotic fluid of the bacterial strain or bacterium Contain phosphate solubilizing bacteria in agent, with phosphorus decomposing function, soil texture, absorption of the enhancing crop to fertilizer can be improved.
Embodiment
The invention will now be further described with reference to specific embodiments, advantages of the present invention and feature will be with description and It is apparent.But these embodiments are only exemplary, do not constitute any limitation to the scope of the present invention.People in the art Member to the details and form of technical solution of the present invention it should be understood that can enter without departing from the spirit and scope of the invention Row modifications or substitutions, but these are changed and replacement is each fallen within protection scope of the present invention.
Embodiment 1：Bacterial strainGibberella intermediaQJP2 separation and identification
（1）The separation of bacterial strain
The bacterial strain of the present inventionGibberella intermediaQJP2 is using plating dilutions coating and line etc. from soil Method separation is obtained, separation method：（1）Culture medium：Meng Jinna Phos solid mediums；（2）Collecting soil sample：Locality Point is the fluffy wetland of Jiaozhou Bay of Qingdao alkali, using 5 point samplings；（3）Separating step：1 g soil samples are weighed in 100 mL sterilized waters, 30 DEG C, 150 r/min, 10 min of concussion are placed in shaking table, 10 are diluted to successively-2、10-3、10-4Concentration.100 μ L are drawn respectively Above-mentioned dilution, is spread evenly across Meng Jinna Phos solid medium planar surfaces, and each gradient coating three is parallel, 30 DEG C 7 d are cultivated, observation whether there is Soluble phosphorus circle, according to Soluble phosphorus loop diameter（D）With colony diameter（d）Ratio size primarily determines that the solution of bacterial strain Phosphorus ability.
By the bacterial strain screening and mask work, bacterial strain is obtainedGibberella intermediaQJP2, tests table Bright, the characteristics of bacterial strain has fast growth, dissolving P capacity is strong has a good application prospect.
（2）The identification of bacterial strain
WillGibberella intermediaQJP2 inoculations are to PDA plate surface, observation after 28 DEG C of cultures 5-7 days.Bacterium Fall rounded, mycelia grows vigorous in cotton-shaped, and initial stage is white, and later stage bacterium colony center is changed into orange red.Microscope inspection is found should Bacterial strain can produce spore, and macroconidium is in avette or spindle, typically has 3 ~ 5 barrier films.
Molecular biological characteristic：The rDNA gene sequencing results of the bacterial strain（ITS1-5.8S-ITS4 areas）It is as follows：
TCCGTAGGTGAACCTTGCGGGGGGGTTTCGAGGTTCCTCCCAACCCCTGTGACATACCAATTGTTGCCTCGGC GGATCAGCCCGCTCCCGGTAAAACGGGACGGCCCGCCAGAGGACCCCTAAACTCTGTTTCTATATGTAACTTCTGAG TAAAACCATAAATAAATCAAAACTTTCAACAACGGATCTCTTGGTTCTGGCATCGATGAAGAACGCAGCAAAATGCG ATAAGTAATGTGAATTGCAGAATTCAGTGAATCATCGAATCTTTGAACGCACATTGCGCCCGCCAGTATTCTGGCGG GCATGCCTGTTCGAGCGTCATTTCAACCCTCAAGCCCCCGGGTTTGGTGTTGGGGATCGGCGAGCCCTTGCGGCAAG CCGGCCCCGAAATCTAGTGGCGGTCTCGCTGCAGCTTCCATTGCGTAGTAGTAAAACCCTCGCAACTGGTACGCGGC GCGGCCAAGCCGTTAAACCCCCAACTTCTGAATGTTGACCTCGGATCAGGTAGGAATACCCGCTGAACTTAAGCATA TCAATAAGCGGAGGACCGCAAGGTTCACCTACGGA
Said gene sequence is compared with known bacterial strain corresponding sequence information in Genbank databases, itself and bacterial strainGibberella intermedia(Accession number:JQ846048.1 similarity) is 99%.
Embodiment 2：Bacterial strainGibberella intermediaQJP2 scale fermentation
（1）Bacterial strain is activated：Bacterial strainGibberella intermediaQJP2 is stored in PDA culture medium inclined-plane, -20 DEG C of preservations. Before use, by bacterial strain streak inoculation on PDA culture medium surface, activation 2 times is standby.
（2）It is prepared by seed liquor：By the bacterial strain activatedGibberella intermediaQJP2 is inoculated into equipped with PDB trainings In the triangular flask for supporting base, 30 DEG C, 120r/min shaken cultivation 3-5 d；
The PDB culture mediums are：The g of potato 200, the g of glucose 20, distilled water 1000 mL, pH are adjusted to 7.0；
（3）It is prepared by solids manufacture strain：By step（2）The seed liquor prepared in mass ratio 10% is inoculated into solid medium In, 28-30 DEG C of incubated 5-7 d；
The solid medium is：Gu material and inorganic salt solution in mass ratio 1:1.8 mixing；The solid material is mass ratio 85: 15 rice chaff and wheat bran；The inorganic salt solution is 3.5% potassium dihydrogen phosphate, 0.04% magnesium sulfate and 4% ammonium sulfate；
（4）Scale solid fermentation：By step（3）The culture prepared and sterilized water in mass ratio 1:15 mixing, sterile yarn Cloth is filtered.By filtrate by volume 1:6 are inoculated into scale fermentation culture medium, in 28-30 DEG C, the fermentation of relative humidity more than 80% Scale fermentation 8-10 d, are obtained in roomGibberella intermediaQJP2 original powder, its living bacteria count is more than 2 × 108 cfu/g.The same step of scale solid fermentation culture medium（3）Described in solid medium.
Embodiment 3：Bacterial strainGibberella intermediaQJP2 plot experiment
Test material：Bacterial strainGibberella intermediaQJP2 zymophyte powder, with phosphorus decomposing effect, living bacteria count More than 2 × 108 cfu/g。
Test period：In April, 2016~2016 year October
Test site：Forest-science academy of Shandong Province Pilot Base
Experimental cultivar：Cotton（Shandong cotton grinds No. 28）
Test soil：Moisture soil
Experimental design：Using clear water seed dressing as control, with different dilution factors（10 times, 20 times, 30 times of dilution）Bacterium powder liquid mix Plant as demonstration, each handle three parallel, m of plot area 202, random distribution.
Dose and fertilizing method：Before sowing, using different dilution factors（10 times, 20 times, 30 times of dilution）Bacterium powder liquid Seed dressing, using clear water seed dressing as control, other measures are uniformly watered, the field management measure such as weeding with locality.
Economical character and yield investigation：In harvest the previous day measurement plant height, begin section position, basal munure and Single boll weight.
From table 1, useGibberella intermediaQJP2 opportunistic pathogen powder dilution seed dressing cotton, its It is better than the cotton of clear water seed dressing in terms of economical character.
After cotton harvesting, each cell production is measured, and compares its significant difference, 2 are the results are shown in Table.
The output of cotton application form of table 2
From table 2, useGibberella intermediaThe cotton of the dilution seed dressing of QJP2 opportunistic pathogen powder, its yield ratio The control dressed seed with clear water is high.Wherein, the cotton of 10 times of dilutions seed dressing of opportunistic pathogen powder, yield and blank control significant difference, Effect of increasing production is obvious.
It can be seen that, bacterial strainGibberella intermediaThe characteristics of QJP2 has fast growth, dissolving P capacity is strong.Contain There are the zymotic fluid or microbial inoculum of the bacterial strain, with phosphorus decomposing function, soil texture, absorption of the enhancing crop to fertilizer can be improved.
SEQUENCE LISTING
<110>Qingdao Agricultural University
<120>One plant of gibberella and its application
<130> 1
<140> 0
<141> 2017-02-22
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 570
<212> DNA
<213>Gibberella intermediaQJP2 rRNA gene orders（ITS1-5.8S-ITS4 areas）
<400> 1
tccgtaggtg aaccttgcgg gggggtttcg aggttcctcc caacccctgt gacataccaa 60
ttgttgcctc ggcggatcag cccgctcccg gtaaaacggg acggcccgcc agaggacccc 120
taaactctgt ttctatatgt aacttctgag taaaaccata aataaatcaa aactttcaac 180
aacggatctc ttggttctgg catcgatgaa gaacgcagca aaatgcgata agtaatgtga 240
attgcagaat tcagtgaatc atcgaatctt tgaacgcaca ttgcgcccgc cagtattctg 300
gcgggcatgc ctgttcgagc gtcatttcaa ccctcaagcc cccgggtttg gtgttgggga 360
tcggcgagcc cttgcggcaa gccggccccg aaatctagtg gcggtctcgc tgcagcttcc 420
attgcgtagt agtaaaaccc tcgcaactgg tacgcggcgc ggccaagccg ttaaaccccc 480
aacttctgaa tgttgacctc ggatcaggta ggaatacccg ctgaacttaa gcatatcaat 540
aagcggagga ccgcaaggtt cacctacgga 570

Claims (6)

1. one plantGibberellaintermediaQJP2, the deposit number of the bacterial strain is CGMCC No. 13199.
2. the bacterial strain described in claim 1Gibberella intermediaQJP2 can improve phosphate fertilizer utilization efficiency, improve soil Earth structure.
3. a kind of microbial bacterial agent, contains the bacterial strain described in claim 1Gibberella intermedia QJP2。
4. a kind of method of the microbial bacterial agent prepared described in claim 3, it is characterized in that：
（1）PrepareGibberella intermediaQJP2 seed liquors；
（2）By step（1）The seed liquor prepared is inoculated into solid medium, and 28-30 DEG C incubated；
（3）By step（2）The culture prepared is mixed with sterilized water, filtering；Filtrate is inoculated into scale fermentation culture medium In, carry out scale fermentation in 28-30 DEG C, the fermenting cellar of relative humidity more than 80%.
5. microbial bacterial agent preparation method as claimed in claim 4, it is characterized in that：
（1）By the bacterial strain activatedGibberella intermediaQJP2 is inoculated into the triangular flask equipped with PDB culture mediums, 30 DEG C, 120 r/min shaken cultivation 3-5 d；
The PDB culture mediums are：The g of potato 200, the g of glucose 20, distilled water 1000 mL, pH are adjusted to 7.0；
（2）By step（1）The seed liquor prepared in mass ratio 10% is inoculated into solid medium, and 28-30 DEG C incubated 5-7 d；
The solid medium is：Gu material and inorganic salt solution in mass ratio 1:1.8 mixing；The solid material is mass ratio 85: 15 rice chaff and wheat bran；The inorganic salt solution is 3.5% potassium dihydrogen phosphate, 0.04% magnesium sulfate and 4% ammonium sulfate；
（3）By step（2）The culture prepared and sterilized water in mass ratio 1:15 mixing, filtering；By filtrate by volume 1:6 are inoculated into scale fermentation culture medium, the scale fermentation 8-10 d in 28-30 DEG C, the fermenting cellar of relative humidity more than 80%；
The same step of scale fermentation culture medium（2）Described in solid medium.
6. the microbial bacterial agent described in claim 4 can improve phosphate fertilizer utilization efficiency, improve soil texture.
CN201710147806.7A 2017-03-14 2017-03-14 One plant of gibberella and its application Pending CN106967615A (en)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114456949A (en) * 2022-01-13 2022-05-10 贵州民族大学 Beauveria bassiana JSHA-MD912 and application thereof
CN114480141A (en) * 2022-01-19 2022-05-13 青岛中尚奇生物科技有限公司 Gibberellin QJP2 and application thereof in weeding

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1884218A (en) * 2006-07-07 2006-12-27 江西省农业科学院土壤肥料与资源环境研究所 Technology for producing granular organic commercial fertilizer by using waste residue of gibberella culture medium
CN102492750A (en) * 2011-12-26 2012-06-13 江南大学 Method for converting 3-cyanopyridine into nicotinic acid by using gibberella intermedia CA3-1

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1884218A (en) * 2006-07-07 2006-12-27 江西省农业科学院土壤肥料与资源环境研究所 Technology for producing granular organic commercial fertilizer by using waste residue of gibberella culture medium
CN102492750A (en) * 2011-12-26 2012-06-13 江南大学 Method for converting 3-cyanopyridine into nicotinic acid by using gibberella intermedia CA3-1

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
P.W.BRIAN ET AL.,: ""The plant-growth-properties of gibberellic acid, a metabolic product of the fungus gibberella fujikuroi"", 《J.SCI.FOOD AGRIC》 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114456949A (en) * 2022-01-13 2022-05-10 贵州民族大学 Beauveria bassiana JSHA-MD912 and application thereof
CN114456949B (en) * 2022-01-13 2023-06-16 贵州民族大学 Beauveria bassiana JSHA-MD912 and application thereof
CN114480141A (en) * 2022-01-19 2022-05-13 青岛中尚奇生物科技有限公司 Gibberellin QJP2 and application thereof in weeding
CN114480141B (en) * 2022-01-19 2023-08-25 青岛中尚奇生物科技有限公司 Gibberella QJP and application thereof in weeding

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Application publication date: 20170721