CN107937283A - Intend trichodermaharzianum fluid nutrient medium - Google Patents
Intend trichodermaharzianum fluid nutrient medium Download PDFInfo
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- CN107937283A CN107937283A CN201711292160.8A CN201711292160A CN107937283A CN 107937283 A CN107937283 A CN 107937283A CN 201711292160 A CN201711292160 A CN 201711292160A CN 107937283 A CN107937283 A CN 107937283A
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- trichodermaharzianum
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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
- C12N1/14—Fungi; Culture media therefor
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Abstract
Intend trichodermaharzianum fluid nutrient medium, it is related to a kind of fungi liquid culture medium.Solve the defects of existing plan trichodermaharzianum fluid nutrient medium spore amplification is undesirable.Intending trichodermaharzianum fluid nutrient medium includes KNO3、KH2PO4、MgSO4、FeCl3And sucrose.The present invention intends trichodermaharzianum fluid nutrient medium can improve about 10 times of spore output for intending trichodermaharzianum under same culture conditions.Fluid present invention culture medium has the advantages that component is simple, cheap, can also improve the quality for intending trichodermaharzianum microbial inoculum.
Description
Technical field
The present invention relates to a kind of fungi liquid culture medium.
Background technology
Fungus Trichoderma belongs to Fungi Imperfecti door, Hyphomycetes, Moniliales, Moniliaceae；Perfect stage is Ascomycota
Hypocrea (Hypocrea Er.).Fungus Trichoderma be widely present in vegetable seeds, rhizosphere, bulb, leaf enclose with soil etc. from
In right ecological environment, and there is antagonism to most plants pathogen.
1932, Weindling had found Trichoderma lignorum (T.lignorum) and Rhizoctonia solani Kuhn (Rhizoctonia
When solani) co-culturing, Rhizoctonia solani Kuhn is parasitic and dead by Trichoderma lignorum, and Weindling considers that the trichoderma can be answered
In the biological control for using plant soil-borne diseases, trichoderma as biological prevents anti-research work from that point on.At present in the world containing wood
The microbial bacterial agent of mould component reaches kind more than 100, is used by more than 60 countries, and significant effect, to being drawn by pathogen
The control of plant disease rate risen is up to 70%.End data in 2014, have been subjected to molecular sequences both at home and abroad and be analyzed to identify in the presence of system
Trichoderma/meat seat bacterium (Trichoderma/Hypocrea) species of system development relation (or forming holotype) reaches 212, and China is
Know trichoderma/91 kinds of Hypocrea fungi.Green trichoderma (T.virid), Trichoderma harzianum (T.harzianum), hook trichoderma
(T.hamatum) had proved with koning trichoderma (T.koningir) etc. with Biocontrol Effect.
Application mode of the trichoderma in biological control mainly has：(1) soil treatment：Reesei spores and soil are mixed
Close, for preventing droop and damping-off, prevention effect is up to more than 80%；(2) seed treatment：With reesei spores powder to seed into
Row parcel, the prevention of the samping off before being unearthed for nursery stock and after being unearthed have a special efficacy, and the germination percentage of seed will be apparently higher than
It is coated without trichoderma；(3) blade face fruit sprays：The blade face fruit of strawberry is carried out using the spore suspension of trichoderma TS bacterial strains
Sprinkling, for preventing the mould gray mold of grass, its preventive effect is more than 80%；(4) it is used in combination with fungicide：Fungicide and trichoderma
Combination application can be remarkably reinforced biocontrol effect, can not only reduce the dosage of fungicide, but also reduce persticide residue, reduce
Environmental pollution, mitigates pesticide and beneficial microorganism in environment is destroyed.
As engineered strain in order to meet the needs of industrialized production, a large amount of amplification breedings are essential in production application
The step of.Trichoderma intends trichodermaharzianum and all shows excellent effect in water process, heavy metal processing, pathogen prevention；
But plan trichodermaharzianum fluid nutrient medium expansion culture effect is not fully up to expectations at present, spore content still only has 1.0 after cultivating 7 days
×108A/ml or so.
The content of the invention
The object of the present invention is to provide a kind of suitable fluid nutrient medium intended trichodermaharzianum and expand culture, to solve existing plan
The defects of amplification of trichodermaharzianum fluid nutrient medium spore is undesirable.
The present invention, which intends every liter of trichodermaharzianum fluid nutrient medium, includes 10 ± 0.2g KNO3、5±0.1g KH2PO4、2.5±
0.1g MgSO4、0.02g FeCl3With 50 ± 0.5g sucrose.
The present invention intends trichodermaharzianum fluid nutrient medium can improve the spore production for intending trichodermaharzianum under same culture conditions
About 10 times of amount.Fluid present invention culture medium has the advantages that component is simple, cheap, can also improve and intend trichodermaharzianum microbial inoculum
Quality.
Embodiment
Technical solution of the present invention is not limited to act embodiment set forth below, further includes between each embodiment
Any combination.
Embodiment one：Every liter of present embodiment intends trichodermaharzianum fluid nutrient medium by 10g KNO3、5g KH2PO4、
2.5g MgSO4、0.02g FeCl3, 50g sucrose and surplus distilled water composition, pH=6.
Embodiment 1
Potato dextrose broth：1L is mainly made of 200g potatos and 20g glucose；Peeling potatoes are cut
Into the fritter of about 0.5mm square, take 200g to be put into the beaker of 1500mL and boil 30min, pay attention to being stirred to prevent paste with glass bar
Bottom, is then filtered with double gauze, takes its filtrate to add 20g glucose, then supplies distilled water to 1000mL.
Embodiment 2
Common carbon source nitrogen source fluid nutrient medium：1L is mainly made of 20g sucrose and 3g dusty yeasts；By 20g sucrose and 3g ferment
Female powder is added in distilled water, and heating makes to be completely dissolved, and is supplied distilled water to 1000mL, is adjusted pH to 7.0~7.2,121 DEG C of sterilizings
15min。
Embodiment 3
Fluid present invention culture medium：By 10g KNO3、5g KH2PO4、2.5g MgSO4、0.02g FeCl3, 50g sucrose and
The distilled water composition of surplus, pH=6.By 10g KNO3、5g KH2PO4、2.5g MgSO4、0.02g FeCl3, 50g sucrose add
Into distilled water, stirring and dissolving, then distilled water is supplied to 1000mL, and pH to 6.0 is adjusted, 121 DEG C of sterilizing 15min.
Contrast experiment：
Plan trichodermaharzianum bacterium opportunistic pathogen kind is continuously transferred, and it is vigorous to growing to activate：Intend trichodermaharzianum bacterium actication of culture, will intend
Trichodermaharzianum is inoculated in 25~27 DEG C of activation culture 6d~9d of PDA culture medium；Then the plan trichodermaharzianum bacterium after activation is inoculated with
Into PD fluid nutrient mediums, in 25~27 DEG C of Shaking culture 36h~48h, liquid seeds are made；Then liquid seeds are seeded to
In liquid fermentation medium, the fermented and cultured 5d under conditions of 25~27 DEG C；Calcium carbonate is added in the ratio of zymotic fluid 3% again,
Mix and use sterile filter-cloth filtering, air-dry, both obtain intending trichodermaharzianum microbial inoculum；
Wherein, liquid seeds are transferred to one grade fermemtation tank culture, and inoculum concentration is 1%~5%, fermented incubation time for 36h~
48h；It is 3%~8% to be inoculated with by one grade fermemtation tank into secondary liquid fermentation tank, inoculum concentration, and secondary liquid fermented incubation time is
30h~40h, inoculum concentration 3%~8%；It is inoculated with by second order fermentation tank into tertiary liquid fermentation tank, inoculum concentration is 5%~10%, is led to
Tolerance 1:0.6~0.8 (V/Vmin), mixing speed 200r/min, tertiary liquid fermentation incubation time are 30h~40h.Its
In, the Liquid Culture based component in level-one, two level and tertiary liquid fermentation tank is identical, and fermentation time amounts to 5d.
Respectively using the fluid nutrient medium of embodiment 1~3 as fluid nutrient medium, above-mentioned experiment is carried out.Experimental result shows,
At the end of using 1 fluid nutrient medium culture of embodiment, the spore content for intending trichodermaharzianum in culture medium is about 1.0 × 108A/
ml；At the end of using 2 fluid nutrient medium culture of embodiment, the spore content for intending trichodermaharzianum in culture medium only has 0.6 × 108
A/ml；At the end of using 3 fluid present invention culture medium of embodiment, the spore content of intending trichodermaharzianum in culture medium reaches 1.0 ×
109A/ml.It is healthy and free from worry that the results show present invention plan trichodermaharzianum fluid nutrient medium can improve plan under same culture conditions
At least more than 10 times of the spore output of trichoderma, spore concentration is obviously improved.
Micro- sem observation, have in the plan trichodermaharzianum gone out using fluid present invention medium culture a large amount of chlamydospores (1~
2×107A/ml), and thick wall spore is not observed in the plan trichodermaharzianum turned out using Examples 1 and 2 fluid nutrient medium culture
Son.The present invention, which intends trichodermaharzianum fluid nutrient medium, can significantly improve microbial inoculum quality.
The plan trichodermaharzianum that fluid present invention medium culture goes out is placed under normal temperature condition and is preserved 24 months, viable count is still
It can reach 4~8 × 108A/ml, and the plan trichodermaharzianum normal temperature condition turned out using Examples 1 and 2 fluid nutrient medium culture
Lower to preserve 24 months, viable count is<1.0×106A/ml.Experiment is proved can using present invention plan trichodermaharzianum fluid nutrient medium
To extend the preservation under room temperature time of microbial inoculum.
Field pathogen prevention effect experimental data is directly proportional to intending trichodermaharzianum viable count in fluid nutrient medium.
Claims (3)
1. intend trichodermaharzianum fluid nutrient medium, it is characterised in that intending trichodermaharzianum fluid nutrient medium for every liter includes 10 ± 0.2g
KNO3、5±0.1g KH2PO4、2.5±0.1g MgSO4、0.02g FeCl3With 50 ± 0.5g sucrose.
2. plan trichodermaharzianum fluid nutrient medium according to claim 1, it is characterised in that intend trichodermaharzianum fluid nutrient medium
PH=6.
3. plan trichodermaharzianum fluid nutrient medium according to claim 1, it is characterised in that every liter is intended the training of trichodermaharzianum liquid
Base is supported by 10g KNO3、5g KH2PO4、2.5g MgSO4、0.02g FeCl3, 50g sucrose and surplus distilled water composition, pH=
6。
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| CN201711292160.8A CN107937283A (en) | 2017-12-08 | 2017-12-08 | Intend trichodermaharzianum fluid nutrient medium |
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| CN201711292160.8A CN107937283A (en) | 2017-12-08 | 2017-12-08 | Intend trichodermaharzianum fluid nutrient medium |
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59151888A (en) * | 1983-02-18 | 1984-08-30 | Res Assoc Petroleum Alternat Dev<Rapad> | Production of cellulase |
| DE3823068A1 (en) * | 1988-07-07 | 1990-01-11 | Biotechnolog Forschung Gmbh | Steroids and preparation processes |
| CN102154194A (en) * | 2011-03-31 | 2011-08-17 | 上海交通大学 | Preparation method for high-yield chlamydospore liquid by virtue of fermentation of trichoderma on pilot plant test scale |
| CN106399129A (en) * | 2016-12-01 | 2017-02-15 | 中国热带农业科学院橡胶研究所 | Trichoderma harzianum strain and application thereof |
| CN107043711A (en) * | 2017-04-26 | 2017-08-15 | 山东省农业科学院农产品研究所 | One plant of bacterial strain of trichoderma aureoviride T 321010 and its cultural method and application |
-
2017
- 2017-12-08 CN CN201711292160.8A patent/CN107937283A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59151888A (en) * | 1983-02-18 | 1984-08-30 | Res Assoc Petroleum Alternat Dev<Rapad> | Production of cellulase |
| DE3823068A1 (en) * | 1988-07-07 | 1990-01-11 | Biotechnolog Forschung Gmbh | Steroids and preparation processes |
| CN102154194A (en) * | 2011-03-31 | 2011-08-17 | 上海交通大学 | Preparation method for high-yield chlamydospore liquid by virtue of fermentation of trichoderma on pilot plant test scale |
| CN106399129A (en) * | 2016-12-01 | 2017-02-15 | 中国热带农业科学院橡胶研究所 | Trichoderma harzianum strain and application thereof |
| CN107043711A (en) * | 2017-04-26 | 2017-08-15 | 山东省农业科学院农产品研究所 | One plant of bacterial strain of trichoderma aureoviride T 321010 and its cultural method and application |
Non-Patent Citations (3)
| Title |
|---|
| 孙秀梅等: "《农业生物技术》", 31 January 2006, 中国农业出版社 * |
| 王福源等: "《生物工艺技术》", 30 September 2006, 中国轻工业出版社 * |
| 赵永强: "木霉菌对大豆根腐病菌的生防机制及其制剂的初步研究", 《中国优秀硕士学位论文全文数据库 农业科技辑》 * |
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