CN105754870A - Trichoderma cyanodichotomus and application thereof - Google Patents
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Abstract
The invention relates to trichoderma cyanodichotomus and an application thereof. The trichoderma cyanodichotomus is named as TW21990-1, the preservation number of the biological strain of the trichoderma cyanodichotomus is CGMCC No.12161, the preservation date is February 23th, 2016, and the preservation authority is CGMCC (China General Microbiological Culture Collection Center) in No.3, No.1 courtyard on the west Beichen road in Chaoyang District in Beijing. The salt tolerance rate is 90.2%. The control effects of a preparation containing two kinds of conidiospores on tomato damping off and cucumber blight are 87.2% and 85.7% respectively, and the yield increasing effects on tomatoes and cucumbers in saline-alkali soil are 67.5% and 67.3% respectively.
Description
Technical field
The present invention relates to a kind of to blue trichoderma strain and application thereof, be specifically related to a kind of to blue trichoderma strain
(Trichoderma cyanodichotomus) TW21990-1 and this bacterial strain are prevented in salt-soda soil and disease thereof
Application in controlling, belongs to microbe application field.
Background technology
Salt-soda soil is a kind of salt aggregation, refers to that salt contained inside soil has influence on crop
Normal growth, according to UNESCO and food and agricultural organization's incomplete statistics, saline and alkaline ground, the whole world
Amassing is 9.5438 hundred million hm2, China's salt-soda soil area is 0.9913 hundred million hm2.China's alkaline earth and alkali-affected soil
Formation, accumulative relevant with Carbonates In Soil of major part, thus basicity is universal the highest, serious
Salt affected soil regional plant is little to existence.China's saline-alkali soil areal area be soil types according to it and
Weather conditions change determines, is divided into salting district, strand, Huang-Huai-Hai plain salting district, desert and desert grass
Crude salt stain district, the big type in four, salting district, grassland.Salt-soda soil preventing and treating and exploitation are that irrigated area agricultural development is not allowed
The problem ignored, for promoting agricultural sustainable development, Guarantee Grain Production has important function.
For improvement and the utilization in salt-soda soil, 19th century gutter drainage Europe prevailing, mid-term in 20th century each reality
Test room to study for moisture and salt regime, be basically divided into physical model, deterministic models etc., 20th century
End abroad starts to sound out biological modification method.
Domestic alkaline land improving research is started late, and generally speaking, defines with physically improved, water conservancy
Improvement, biological modification and the 4 big improvement systems that chemical modifying is core.At present, Drainage of salt technology is
Use relatively broad a kind of improvement salt-soda soil measure, but have that construction investment is big, land occupation is many, water
The shortcomings such as resource consumption is many and operation and maintenance cost is high.
Recent two decades starts to possess some special knowledge on crops and cultivated plant, and biological salt discharge is to saline and alkaline land productivity
With and salination improvement significant, as Chinese Academy of Sciences's Shanghai plant physiology is cultivated the most anti-successful salt
The most cold-resistant plants Shen series High-Yield Wheat Cultivar, shows the anti-salt of excellence, cold performance.Microbial bacteria
Fertilizer becomes reclamation of salinep-alkali soil new tool in recent years, as Trichoderma preparation is used for the kind of salt-soda soil Semen arachidis hypogaeae by Chen Jianai etc.
Plant, improve planting percent.Cui Xiling etc. filter out salt tolerant, the trichoderma strain of preventing and treating black shank.
But this field basic research is the weakest, mostly rests on the starting stage.From now on, saline alkali tolerant plant and
The screening of microbe species, the selection etc. of the soil conditioner that comprehensive value is high will become alkaline land improving
Research emphasis.
Summary of the invention
For above-mentioned technical problem, the invention provides a kind of to blue trichoderma strain and application thereof, specifically relate to
And to blue trichoderma strain (Trichoderma cyanodichotomus) TW21990-1 and this bacterial strain at salt
Alkali ground and application therein.
The present invention is isolatable from Hunan Province's Dongting Lake Bridge to blue trichoderma strain TW21990-1 in April, 2015
Soil sample, locality longitude 113 ° 06 ' 09 ", 29 ° 24 ' 3.03 of latitude ".To blue trichoderma strain TW21990-1
Biological inoculum deposit number be: CGMCC No.12161, preservation date is: on February 23rd, 2016,
Depositary institution is China Committee for Culture Collection of Microorganisms's common micro-organisms center, and address is Beijing
North Star West Road, Chaoyang District 1 institute 3.Classification And Nomenclature of the present invention is Trichoderma cyanodichotomus.
Present invention additionally comprises containing to blue trichoderma strain TW21990-1, prepared by TW21990-1 bacterial strain
Two kinds of conidium or TW21990-1 bacterial strain fermentation liquid at least any one system prepared
Agent.
As follows to blue trichoderma strain TW21990-1 sweat:
(1) slant culture: use the PDA culture medium of 0.4mol/L NaCl, by inoculation at test tube
In culture medium, cultivate 4～5d for 28 DEG C；
(2) triangular flask is cultivated: use the PDB culture medium of 0.4mol/L NaCl, test tube strains is inoculated
In liquid triangular flask, it is placed in 28 DEG C of shaken cultivation 2d on shaking table；
(3) liquid culture: use liquid fermentation medium, 115 DEG C of sterilizing 30min, strain is inoculated
To fermentation tank, inoculum concentration 0.2% (v/v), 28 DEG C of cultivations, dissolved oxygen amount 40～50%, ventilation 1:0.6～0.8,
Mixing speed is 160r/min, cultivates 2d；
(4) solid culture: using solid fermentation culture medium, 121 DEG C of sterilizing 30min, in mixing tank
Being mixed homogeneously with solid medium by liquid spawn, inoculum concentration is 5～10% (v/v), inoculates complete transferring to
Solid culture room, culture medium thickness is 5cm, and material temperature controls at 28 ± 2 DEG C, and room temperature controls at 25～30 DEG C,
The relative water content of culturing room's air is 95～100%, and incubation time is 4～5d；
(5) spore is collected: with normal saline eluting conidium after cultivation, 5000r/min is centrifuged 10min,
Collect precipitation, obtain two kinds and concentrate conidium；
In step (3), liquid fermentation medium is, Semen Maydis powder 2%, glucose 0.5%, soybean cake powder 4%,
K2HPO40.2%, KH2PO40.3%, NaCl 2.3%, surplus is water, pH 6.0～6.5.
In step (4), solid fermentation culture medium is, glucose 0.5%, NaCl 2.3%, wheat bran 42.2%,
Rice husk 5.0%, surplus is water.
The two kinds of conidial preparations prepared containing the present invention, the most composed of the following components:
Two kinds concentrate conidium 10～15 parts
Maifanitum (300 mesh) 950 parts
10 parts of gelatin
NaCl 23 parts
SDS 1 part.
The present invention compared with prior art has the advantage that
Efficient salt-tolerant of the present invention, on the PDA plate of 0.4mol/L NaCl, growing way is preferable, and salt tolerant rate is
90.2%, suppression ratio mould to melon and fruit corruption, fusarium oxysporum is respectively 100.0%, 95.8%.The present invention can produce
Raw two kinds of conidium varied in size, by count plate, after solid fermentation, microgonidium content is about
Being 22.5 hundred million/g, macroconidium content is about 46.7 hundred million/g.Plot experiment shows, containing the present invention two
Plant conidial preparation to Fructus Lycopersici esculenti damping off (Pythium aphanidermatum) and cucumber fusarium axysporum
The prevention effect of (Fusarium oxysporum) is respectively 87.2% and 85.7%, on salt-soda soil to kind
The effect of increasing production of eggplant and Fructus Cucumidis sativi is respectively 67.5% and 67.3%.
Accompanying drawing explanation
Fig. 1 is the morphological characteristic figure to blue trichoderma strain TW21990-1；
Note: A-C is respectively at culture medium SNA, PDA, CMD to blue trichoderma strain TW21990-1
The front colonial morphology figure of upper 20 DEG C of cultivation 7d；D-F is to exist blue trichoderma strain TW21990-1 respectively
Culture medium SNA, the reverse side colonial morphology of the upper 20 DEG C of cultivation 7d of PDA, CMD；G, 2 kinds of mitogenetic spores
(arrow 1 is microgonidium, and arrow 2 is macroconidium, arrow 3 for son and the chlamydospore come off
Chlamydospore for coming off)；H, the chlamydospore not fallen off；I, conidiophore；Scale G-I=10 μm.
Fig. 2 is the salt tolerance to blue trichoderma strain TW21990-1
Note: A is big to blue trichoderma strain TW21990-1 25 DEG C of bacterium colonies cultivating 4d on PDA plate
Little；B, to blue trichoderma strain TW21990-1 25 DEG C of cultivations on the PDA plate of 0.4mol/L NaCl
The bacterium colony size of 4d.
Fig. 3 is mould flat board inhibitory action figure rotten to melon and fruit to blue trichoderma strain TW21990-1；
Note: A, the rotten mould 25 DEG C of bacterium colony sizes cultivating 4d on PDA plate of comparison melon and fruit；B, melon
Fruit rotten mould with to blue trichoderma strain TW21990-1 bacterium colony of 25 DEG C of opposite culture 4d on PDA plate
Size.
Fig. 4 is to the blue trichoderma strain TW21990-1 flat board inhibitory action figure to fusarium oxysporum；
Note: A, comparison fusarium oxysporum 25 DEG C of bacterium colony sizes cultivating 4d on PDA plate；B, point
Spore fusarium with to blue trichoderma strain TW21990-1 bacterium colony of 25 DEG C of opposite culture 4d on PDA plate
Size.
Detailed description of the invention
Further describing the present invention, advantages of the present invention and feature below in conjunction with specific embodiment will be with
Description and apparent.But embodiment is only exemplary, the scope of the present invention is not constituted any
Limit.It will be understood by those skilled in the art that under without departing from the spirit and scope of the present invention permissible
Details and form to technical solution of the present invention are modified or replace, but these amendments and replacement each fall within
In protection scope of the present invention.
Embodiment 1 is to blue trichoderma strain TW21990-1
Blue Trichoderma spp. TW21990-1 bacterial strain is isolatable from Hunan Province's Dongting Lake Bridge soil sample in April, 2015,
Locality longitude 113 ° 06 ' 09 ", 29 ° 24 ' 3.03 of latitude ".Biology to blue trichoderma strain TW21990-1
Culture presevation is numbered: CGMCC No.12161, and preservation date is: on February 23rd, 2016, preservation
Unit is China Committee for Culture Collection of Microorganisms's common micro-organisms center, and address is Chaoyang, Beijing
North Star West Road, district 1 institute 3.
The molecular sequences analysis of embodiment 2 bacterial strain TW21990-1
ITS, tef1 and rpb2 sequence results in bacterial strain TW21990-1 is compared.
The amplimer of ITS sequence:
ITS1-F:5 '-CTTGGTCATTTAGAGGAAGTAA-3 '
ITS4:5 '-TCCTCCGCTTATTGATATGC-3 '
The ITS sequence total length of bacterial strain TW21990-1 is 665bp, sequence as shown in SEQ ID No.1,
TrichOKEY ITS sequence being committed in www.isth.info compares, result respectively 110,
132, it is found that 5 DNA oligonucleotide bar codings of Trichoderma spp. at 309,467,560, shows bacterium
Strain TW21990-1 is trichoderma, but can not find the sequence higher with this sequence homology in this data base
Information, tentatively shows that this bacterial strain may be for novel species.Www.ncbi.nlm.nih.gov is submitted to enter ITS sequence
Row comparison, TW21990-1 bacterial strain and Trichoderma sp.4TMS-2011vocher MSbale50-9
[HQ630962.1] homology is the highest, is 94%.
Tef1 sequence amplification primer:
EF1-728F:5 '-CATCGAGAAGTTCGAGAAGG-3 '
TEF1LLErev:5 '-AACTTGCAGGCAATGTGG-3 '
The tef1 sequence of bacterial strain TW21990-1 is 1346bp, sequence as shown in SEQ ID No.2,
Tef1 sequence submission NCBI is compared, TW21990-1 bacterial strain and Trichoderma estonicum
Strain CBS111147 [FJ860638.1] homology is the highest, is 91%.
Rpb2 sequence amplification primer:
FRPB2-5F:5 '-GA (T/C) GA (T/C) (A/C) G (A/T) GATCA (T/C) TT (T/C) GG-3 '
FRPB2-7cR:5 '-CCCAT (A/G) GCTTG (T/C) TT (A/G) CCCAT-3 '
The rpb2 sequence of bacterial strain TW21990-1 is 1234bp, sequence as shown in SEQ ID No.3,
Rpb2 sequence submission NCBI is compared, TW21990-1 bacterial strain and Hypocrea strictipilosa
Strain C.P.K.1601 [FJ860594.1] homology is the highest, is 94%.
By above-mentioned analysis, it can be seen that bacterial strain TW21990-1 is trichoderma, its ITS, tef1 and rpb2
Three kinds of sequences are all not more than 94% with the Trichoderma spp. homology of other kind in NCBI, therefore conclude bacterial strain
TW21990-1 is trichoderma novel species.
Embodiment 3 bacterial strain TW21990-1 morphological characteristic describes
Bacterial strain TW21990-1 is 20 DEG C, 25 DEG C, 30 DEG C, 35 DEG C of cultivation 72h in SNA culture medium,
Colony radius is respectively 36～43mm, 62～64mm, 67～71mm, 13～14mm, and more than 40 DEG C are not
Growth, optimum growth temperature is 25～30 DEG C.SNA culture medium cultivates 7d, aerial hyphae for 25 DEG C
Becoming radial growth, nearly vaccination is sparse, and remote vaccination is the densest, and spore bunch results from vaccination edge
Wheel stricture of vagina band on, green, become sector distribution, without obvious abnormal smells from the patient, without obvious pigment, such as Figure 1A and 1D
Shown in.
Bacterial strain TW21990-1 is 20 DEG C, 25 DEG C, 30 DEG C, 35 DEG C of cultivation 72h in PDA culture medium,
Colony radius is respectively 33～39mm, 59～61mm, 66～67mm, 10～11mm, and more than 40 DEG C are not
Growth, optimum growth temperature is 25～30 DEG C.Cultivating 7d for 25 DEG C in PDA culture medium, bacterium colony is uniform,
Aerial hyphae is dense, velvet shape, and conidium celadon is covered with whole flat board, without obvious odorlessness,
The back side produces obvious blue pigment, as shown in Figure 1B and 1E.
Bacterial strain TW21990-1 is 25 DEG C of cultivation 7d in CMD culture medium, produce a large amount of green spores bunch,
Graininess, is randomly distributed, and without obvious odorlessness, the back side produces light blue pigment, such as Fig. 1 C and 1F
Shown in.
As shown in Figure 1 I, bacterial strain TW21990-1 mycelia is transparent, sturdy, and cell wall is smooth, diameter 3.9～4.4
μm, have between mycelia every, gap length is 31.1～41.4 μm, and conidiophore has obvious long axis,
The general Dan Sheng of side shoot, minority is to life, sturdy, and branch is longer, and primary branch size is
(19.2～19.6) × (3.9～4.1) μm, secondary branch size is (8.6～10.2) × (3.7～3.9) μm, between branch
Having distinct visible barrier film, branch is at an acute angle, and the barrier film of sporophore bifurcation is " V "-shaped, and bottle stalk is right
Giving birth to, long flask shape, middle part is slightly expanded, and top is relatively thin, and size is (5.8～6.2) × (3.4～3.6) μm.
As shown in Figure 1 G, conidium has 2 kinds, microconidia (arrow 1 direction) generally circle
Shape, size is (1.0～) 1.2～2.0 (～2.3) × (1.0～) 1.1～1.9 (～2.2) μm, length/width than average out to 1.0～1.1,
Green, wall is smooth.Macroconidium (arrow 2 direction) avette or pyriform, is individually oval,
Size is (3.7～) 4.3～5.0 (～5.5) × (3.4～) 3.8～4.4 (～4.6) μm, length/width ratio (1.0～) 1.1～1.2 (～1.3),
Green or yellow green, wall is smooth.
As shown in Fig. 1 G and 1H, chlamydospore is the abundantest, pyriform or sub-spherical, end is raw, concatenate or
Between raw, (1G arrow 3 direction) easy to fall off, size is (7.3～) 8.4～11.5 (～12.6) × (5.3～) 6.8～7.7 (～9.7)
μm, length/width ratio (1.0～) 1.1～1.2 (～1.6).
By plate morphology and conidiophore branch feature, bacterial strain TW21990-1 should belong to trichoderma
In meat seat group (Hypocreanum), in this group, majority is the asexual generation of meat seat bacterium (Hypocrea),
Trichoderma spp. kind in this group identified is less, bacterial strain TW21990-1 and other Trichoderma spp. kind in this group
Feature differs greatly, and concludes that this bacterial strain is trichoderma novel species further.The present invention is by bacterial strain TW21990-1
Named to blue Trichoderma spp. (Trichoderma cyanodichotomus).
Embodiment 4: the salt tolerance of bacterial strain TW21990-1 and the experiment of flat board opposite culture
In PDA culture medium, add the NaCl of 0.4mol/L NaCl, inoculate trichoderma strain TW21990-1,
With PDA for comparison, 25 DEG C of cultivations, as in figure 2 it is shown, the growth radius of record Trichoderma spp., calculate salt tolerant rate.
Salt tolerant rate=100% × process Trichoderma spp. radius/comparison Trichoderma spp. half warp
At PDA plate diameter two ends at the 35mm of center respectively inoculation diameter 5mm trichoderma strain and
Pathogenic fungi, with only inoculate pathogenic fungi for comparison, 25 DEG C of cultivation, record pathogenic fungi stand facing each other radius,
Calculate suppression ratio.
Suppression ratio=100% × (comparison pathogenic fungi radius-process pathogenic fungi radius)/comparison pathogenic fungi half warp
By above-mentioned experiment, this trichoderma strain TW21990-1 is on the PDA plate of 0.4mol/L NaCl
Salt tolerant rate be 90.2%.
Mould flat board inhibitory action rotten to melon and fruit to blue Trichoderma spp. TW21990-1 as it is shown on figure 3, wherein, A
For the rotten mould 25 DEG C of bacterium colony sizes cultivating 4d on PDA plate of comparison melon and fruit；B is that melon and fruit is rotten mould and right
Blue Trichoderma spp. TW21990-1 bacterial strain bacterium colony size of 25 DEG C of opposite culture 4d on PDA plate.To indigo plant
To the flat board inhibitory action of fusarium oxysporum as shown in Figure 4, wherein, A is right to Trichoderma spp. TW21990-1 bacterial strain
According to fusarium oxysporum 25 DEG C of bacterium colony sizes cultivating 4d on PDA plate；B is that fusarium oxysporum is wooden with to indigo plant
Mould TW21990-1 bacterial strain is the bacterium colony size of 25 DEG C of opposite culture 4d on PDA plate.Through meter
The calculation present invention is mould to melon and fruit corruption to blue Trichoderma spp. TW21990-1 bacterial strain, the flat board suppression ratio difference of fusarium oxysporum
It is 100.0%, 95.8%.
Embodiment 5: the present invention contains the industrialized production of two kinds of conidium preparations
(1) slant culture: use the PDA culture medium of 0.4mol/L NaCl, by inoculation at test tube
In culture medium, cultivate 4～5d for 28 DEG C；
(2) triangular flask is cultivated: use the PDB culture medium of 0.4mol/L NaCl, test tube strains is inoculated
In liquid triangular flask, it is placed in 28 DEG C of shaken cultivation 2d on shaking table；
(3) liquid culture: use liquid fermentation medium, 115 DEG C of sterilizing 30min, strain is inoculated
To fermentation tank, inoculum concentration 0.2% (v/v), 28 DEG C of cultivations, dissolved oxygen amount 40～50%, ventilation 1:0.6～0.8,
Mixing speed is 160r/min, cultivates 2d；
(4) solid culture: using solid fermentation culture medium, 121 DEG C of sterilizing 30min, in mixing tank
Being mixed homogeneously with solid medium by liquid spawn, inoculum concentration is 5～10% (v/v), inoculates complete transferring to
Solid culture room, culture medium thickness is 5cm, and material temperature controls at 28 ± 2 DEG C, and room temperature controls at 25～30 DEG C,
The relative water content of culturing room's air is 95～100%, and incubation time is 4～5d；
(5) spore is collected: with normal saline eluting conidium after cultivation, 5000r/min is centrifuged 10min,
Collect precipitation, obtain two kinds and concentrate conidium；
In step (3), liquid fermentation medium (mass percent) is, Semen Maydis powder 2%, glucose
0.5%, soybean cake powder 4%, K2HPO40.2%, KH2PO40.3%, NaCl 2.3%, surplus is water, pH
6.0～6.5.
In step (4), solid fermentation culture medium (mass percent) is, glucose 0.5%, NaCl 2.3%,
Wheat bran 42.2%, rice husk 5.0%, surplus is water.
Described preparation, consists of the following composition:
In preparation, effective active composition is two kinds of conidium, and total content is 500,000,000/g, its middle-size and small-size mitogenetic spore
Sub-ratio is 32.5%, and macroconidium ratio is 67.5%.Due to the distinctive generation of this Trichoderma spp. two kinds points
Sporogenic ability, makes this Trichoderma spp. have Efficient salt-tolerant and disease-controlling effect.
Embodiment 6: the present invention contains two kinds of conidium preparations prevention effect to salt-soda soil Fructus Lycopersici esculenti damping off
And effect of increasing production
Test and be located at Kenli County, Dongying city, experimental field salinity 0.6%, after testing, mainly
Pathogenic bacteria is that melon and fruit corruption is mould, and tomato variety is Tian Bao one, great river, Jinan seed company limited.Test sets 3
Individual process: processing 1 is preparation, processing 2 is medicament (metalaxyl), and processing 3 is comparison, often processes and sets
4 communities are repeated, plot area 16m2, random distribution.
After seed routine accelerating germination, the seed that picking form is consistent is divided into 3 parts, process 1 with this
Bright preparation is mixed thoroughly, and about 500 seeds use 1g preparations to process, and makes the conidium number of absorption on seed be
106Cfu/ grain, processes 2 and soaks 30min with the metalaxyl of 100 μ g/mL, processes 3 and rinses well with clear water.
Sow according to a conventional method, manage.After emerging, 2 weeks investigation planting percents, calculate prevention effect.Unite after results
Meter yield, calculates effect of increasing production.
Prevention effect %=100% × (process group planting percent-matched group planting percent)/(1-matched group planting percent)
Effect of increasing production %=100% × (process group yield-matched group yield)/matched group yield
The results are shown in Table 1, invention formulation can be effectively improved the planting percent of Fructus Lycopersici esculenti in salt-soda soil, and can be the most anti-
Controlling the generation of Fructus Lycopersici esculenti damping off, prevention effect has reached 87.2%, and effect of increasing production has reached 67.5%, effect
Significantly (p ＜ 0.01).
Table 1 invention formulation is to the prevention effect of salt-soda soil Fructus Lycopersici esculenti damping off and effect of increasing production
(note: the Duncan method in data acquisition SPSS 10.0 statistical software is analyzed, result is 4 weights
Multiple mean+SD, represents with different capitalizations and there is significance in 0.01 level between each process
Difference.)
Embodiment 7: the present invention contains two kinds of conidium preparations prevention effect to salt-soda soil cucumber fusarium axysporum
And effect of increasing production
Test and be located at Kenli County, Dongying city, experimental field salinity 0.8%, after testing, mainly
Pathogenic bacteria is fusarium oxysporum, and cucumber variety is the most excellent No. one, great river, Jinan seed company limited.Test sets 3
Individual process: processing 1 is preparation, processing 2 is medicament (probenazole), and processing 3 is comparison, often processes and sets
4 communities are repeated, plot area 16m2, random distribution.
After seed routine accelerating germination, the seed that picking form is consistent is divided into 3 parts, process 1 with this
Bright preparation is mixed thoroughly, and about 500 seeds use 1g preparations to process, and makes the conidium number of absorption on seed be
106Cfu/ grain, processes 2 and soaks 30min with the probenazole of 100 μ g/mL, processes 3 and rinses well with clear water.
Sow according to a conventional method, manage.After emerging, 4 weeks investigation disease indexs, calculate prevention effect.After results
Statistics yield, calculates effect of increasing production.
Cucumber fusarium axysporum occurring degree:
0 grade is without disease；
1 grade there is mild disease for plumular axis or cotyledon, but growth is normal；
2 grades is plumular axis or the cotyledon obvious ecthyma gangrenosa of appearance, or 1 cotyledon yellow, impact growth；
3 grades is 2 cotyledon yellows, or 1 cotyledon is withered；
4 grades is that 2 cotyledon growths ossify, and plant part is wilted or stops growing；
5 grades is that whole strain is wilted, lodged or withered.
Disease index %=100% × (∑ (state of an illness rank × this grade of diseased plant number)/(5 × investigate total strain number)
Protection effect %=100% × (comparison disease index-process disease index)/comparison disease index)
Effect of increasing production %=100% × (process group yield-matched group yield)/matched group yield
The results are shown in Table 2, invention formulation can effectively reduce the disease index of alkali Viola grypoceras A. Gray, and can effectively prevent and treat
The generation of cucumber fusarium axysporum, prevention effect has reached 85.7%, and effect of increasing production has reached 67.3%, and effect shows
Write (p ＜ 0.01).
Table 2 invention formulation is to the prevention effect of salt-soda soil cucumber fusarium axysporum and effect of increasing production
(note: the Duncan method in data acquisition SPSS 10.0 statistical software is analyzed, result is 4 weights
Multiple mean+SD, represents with different capitalizations and there is significance in 0.01 level between each process
Difference.)
Claims (9)
1. one kind to blue trichoderma strain, it is characterised in that described named to blue trichoderma strain
TW21990-1, its biological inoculum deposit number is: CGMCC No.12161, and preservation date is: 2016
In on February 23, in, depositary institution is China Committee for Culture Collection of Microorganisms's common micro-organisms center,
Address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3.
2. as claimed in claim 1 to blue trichoderma strain, and by prepared by blue trichoderma strain two kinds
At least any one preparation prepared in conidium or fermentation liquid.
3. the fermentation technology to blue trichoderma strain as claimed in claim 1, specifically comprises the following steps that
(1) slant culture: use the PDA culture medium of 0.4mol/L NaCl, by bacterial strain TW21990-1
It is seeded on Tube propagation base, cultivates 4～5d for 28 DEG C；
(2) triangular flask is cultivated: use the PDB culture medium of 0.4mol/L NaCl, test tube strains is inoculated
In liquid triangular flask, it is placed in 28 DEG C of shaken cultivation 2d on shaking table；
(3) liquid culture: use liquid fermentation medium, 115 DEG C of sterilizing 30min, strain is inoculated
To fermentation tank, inoculum concentration 0.2% (v/v), 28 DEG C of cultivations, dissolved oxygen amount 40～50%, ventilation 1:0.6～0.8,
Mixing speed is 160r/min, cultivates 2d；
(4) solid culture: using solid fermentation culture medium, 121 DEG C of sterilizing 30min, in mixing tank
Being mixed homogeneously with solid medium by liquid spawn, inoculum concentration is 5～10% (v/v), inoculates complete transferring to
Solid culture room, culture medium thickness is 5cm, and material temperature controls at 28 ± 2 DEG C, and room temperature controls at 25～30 DEG C,
The relative water content of culturing room's air is 95～100%, and incubation time is 4～5d；
(5) spore is collected: with normal saline eluting conidium after cultivation, 5000r/min is centrifuged 10min,
Collect precipitation, obtain two kinds and concentrate conidium；
In step (3), liquid fermentation medium is, Semen Maydis powder 2%, glucose 0.5%, soybean cake powder 4%,
K2HPO40.2%, KH2PO40.3%, NaCl 2.3%, surplus is water, pH 6.0～6.5；
In step (4), solid fermentation culture medium is, glucose 0.5%, NaCl 2.3%, wheat bran 42.2%,
Rice husk 5.0%, surplus is water.
4. two kinds of conidium that as claimed in claim 3 prepared by fermentation technology.
Preparation prepared by two kinds of conidium the most as claimed in claim 4.
Preparation prepared by two kinds of conidium the most as claimed in claim 5, according to the mass fraction by following group
It is grouped into:
7. as claimed in claim 1 to blue trichoderma strain in terms of salt tolerant, in terms of disease control or volume increase
The application of aspect；Preferably, described disease refers to Fructus Lycopersici esculenti damping off or cucumber fusarium axysporum.
Two kinds of conidium the most as claimed in claim 2 are in terms of salt tolerant, in terms of disease control or volume increase
The application of aspect；Preferably, described disease refers to Fructus Lycopersici esculenti damping off or cucumber fusarium axysporum.
9. preparation as claimed in claim 3 answering in terms of salt tolerant, in terms of disease control or in terms of volume increase
With；Preferably, described disease refers to Fructus Lycopersici esculenti damping off or cucumber fusarium axysporum.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106434370A (en) * | 2016-09-23 | 2017-02-22 | 山东省科学院生态研究所 | Trichoderma protoplast fusion strain and application thereof |
| CN108076973A (en) * | 2018-01-15 | 2018-05-29 | 石家庄学院 | A kind of production method of mushroom concentrated strain |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100028303A1 (en) * | 2008-07-17 | 2010-02-04 | Bioworks, Inc. | Control of plant diseases and enhancing plant growth using a combination of a trichoderma virens species and a rhizosphere competent trichoderma harzianum species |
| CN103952320A (en) * | 2014-04-28 | 2014-07-30 | 唐山金土生物有机肥有限公司 | Trichoderma harzianum and application thereof in preventing and controlling lawn fusarium blight |
| CN105274009A (en) * | 2015-11-24 | 2016-01-27 | 山东省科学院中日友好生物技术研究中心 | Trichoderma strain and preparation for preventing and treating plant pythium ultinum diseases |
| JP2016202107A (en) * | 2015-04-24 | 2016-12-08 | 佐々木 康晴 | Novel microorganisms having excellent antibacterial action and plant growth promoting action |
-
2016
- 2016-03-22 CN CN201610162215.2A patent/CN105754870B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100028303A1 (en) * | 2008-07-17 | 2010-02-04 | Bioworks, Inc. | Control of plant diseases and enhancing plant growth using a combination of a trichoderma virens species and a rhizosphere competent trichoderma harzianum species |
| CN103952320A (en) * | 2014-04-28 | 2014-07-30 | 唐山金土生物有机肥有限公司 | Trichoderma harzianum and application thereof in preventing and controlling lawn fusarium blight |
| JP2016202107A (en) * | 2015-04-24 | 2016-12-08 | 佐々木 康晴 | Novel microorganisms having excellent antibacterial action and plant growth promoting action |
| CN105274009A (en) * | 2015-11-24 | 2016-01-27 | 山东省科学院中日友好生物技术研究中心 | Trichoderma strain and preparation for preventing and treating plant pythium ultinum diseases |
Non-Patent Citations (1)
| Title |
|---|
| JISHUN LI 等: "Trichoderma cyanodichotomus sp. nov., a new soil-inhabiting species with a potential for biological control", 《CANADIAN JOURNAL OF MICROBIOLOGY》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106434370A (en) * | 2016-09-23 | 2017-02-22 | 山东省科学院生态研究所 | Trichoderma protoplast fusion strain and application thereof |
| CN108076973A (en) * | 2018-01-15 | 2018-05-29 | 石家庄学院 | A kind of production method of mushroom concentrated strain |
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