CN101955894A - Saccharothrix used for preventing and controlling tomato leaf mold and preparation method thereof - Google Patents
Saccharothrix used for preventing and controlling tomato leaf mold and preparation method thereof Download PDFInfo
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Abstract
The invention discloses Saccharothrix yanglingensis Hhs.015. The bacterial strain has been preserved in a China General Microbiological Culture Collection Center in June, 2009 with a collection number of CGMCC NO.4.5627; and the bacterial strain also has been preserved in a Type Culture Collection Center of the Korea Institute of Bioscience and Biotechnology in October, 2009 with a collection number of Saccharothrix yanglingensis Hhs.015T/KCTC 19722T. The invention also discloses a separation and identification method for the bacterial strain, a preparation method for thallus and a preparation method for crude antibacterial active extracts, and also discloses application of the Hhs.015 bacterial strain and the thallus and antibacterial active substances thereof in preventing and controlling the tomato leaf mold. The Hhs.015 bacterial strain has the advantages of stable natural raw material source, simple screening method, convenient operation, stable product quality, no toxic or side effect, environmental protection, obvious effect of preventing and controlling the tomato leaf mold, and convenient popularization and application.
Description
Technical field
The invention belongs to biological pesticide technical field, relate to a kind of new bacterial strain that is used to prevent and treat leaf muld of tomato, particularly a kind of saccharothrix of preventing and treating leaf muld of tomato and preparation method thereof that is used to.
Background technology
Leaf muld of tomato (Fulvia fulva (Cook) Ciferri) is commonly called as " black wool disease ", is the disease that generally takes place in the tomato production, and especially harm is even more serious in protection ground, and in a single day this disease takes place, and rapid spread has brought tremendous loss to tomato production.Leaf muld of tomato is the typical epizootic disease evil that is caused by the Deuteromycotina fungi, has the intermittently characteristics of outburst.Germ also can be infected stem and fruit except that causing that leaf spot lesion influences the photosynthesis, influences fruit quality, reduces edibleness, causes financial loss.Because the expansion of protection ground tomato planting area, protection ground planting environment conditions favouring was popular in the generation of leaf mold in addition in recent years, so leaf mold has the trend that increases the weight of year by year, had become on the tomato production one of severe diseases so far.One of its reason is unreasonable relevant with prophylactico-therapeutic measures.
For controlling the harm of leaf mold effectively, China's researcher has carried out extensive studies to aspects such as the disease regularty of epidemic of leaf mold germ and integrated controls, has made a series of prophylactico-therapeutic measuress, comprises chemical control, biological control.A large amount of uses of chemical pesticide cause serious environmental to pollute the pesticide residue of agricultural-food, germ resistance, resistance.These problems have produced people's life and have had a strong impact on, and biological control can reduce the consumption of chemical pesticide to greatest extent, to improve the ecological environment.
Domestic antagonism bacterium is more common to antagonistic effect in the ware of leaf muld of tomato bacterium, but utilizes the antagonism bacterium to prevent and treat the example of leaf muld of tomato and few, and the biological prevention and control agent of preventing and treating leaf muld of tomato just still less.
Endophyte of plant is a kind of can growing surely in plant tissue and running, the microorganism harmless even useful to plant, because it has stable living space in plant materials, and can produce and the same or analogous physiologically active substance of host plant metabolism, thereby can suppress the disease resistance that infects or improve host plant of pathogenic bacteria effectively.In recent years, endophyte had more application potential aspect the biological control than microorganisms such as rhizosphere, Ye Wei because of its unique advantage, thereby became the research focus of biological control, for the new world has been opened up in the application of biological control.The present invention will report living actinomycetes---the discovery and the prophylaxis effect thereof of Yang Ling saccharothrix Hhs.015 bacterial strain in the strain.
Summary of the invention
Purpose of the present invention is intended to seek the new source of biological pesticide, but utilize the endophyte of plant resource and develop the actinomycetes strain that a kind of efficient industrialization that can be used for the graw mold of tomato control is produced, screening, the authentication method of Yang Ling saccharothrix (Saccharothrix yanglingensis) Hhs.015 bacterial strain is provided.
The technical scheme that realizes the foregoing invention purpose is specific as follows:
A kind of Yang Ling saccharothrix Hhs.015 bacterial strain (being designated hereinafter simply as the Hhs.015 bacterial strain), be deposited in China Committee for Culture Collection of Microorganisms common micro-organisms center in June, 2009, its deposit number is CGMCCNO.3023, classification name Yang Ling saccharothrix (Saccharothrix yanglingensis), depositary institution address: Datun Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, postcode: 100101.This bacterial strain also is deposited in Korea S's bio-science and biotechnology research institute typical culture collection center in October, 2009, is numbered Saccharothrix yanglingensis Hhs.015
T/ KCTC 19722
T
The preparation method of Hhs.015 bacterial strain of the present invention, concrete grammar comprises the following steps:
1) Radix Cucumidis sativi to be separated is launched to dry with tap water flushing back, be cut into the thin slice of 0.5 * 0.5mm size after the surface sterilization with the aseptic operation cutter.
2) the Radix Cucumidis sativi tissue block after the surface sterilization is placed contain on the Gause I flat board, cultivate behind the 21d statistics actinomycetes colony number in 28 ± 1 ℃ of incubators and carry out purifying, separable to the Hhs.015 bacterial strain, in-80 ℃ of preservations.
The condition of described surface sterilization is: the root of clip health plant sample soaks 1min after cleaning with clear water in 99% ethanol, soak 6min then in 3.125% NaOCl, puts into 99% ethanol once more and soaks 30sec, uses aseptic water washing at last 5 times.
Whether completely detection surface sterilization method is the sterilized water after the sterilization to be got 200 μ L be coated on the PDA flat board.
Hhs.015 bacterial strain of the present invention through the microscopic examination of bacterium colony and form, cytochemistry compositional analysis, the experiment of conventional physio-biochemical characteristics, 16S rDNA sequential analysis and to similar strain hybrid, the new bacterial strain (Hhs.015) that this identification of strains is has following feature:
(A) morphological specificity: the living mycelia of base is elongated, unfolds prosperity; Multi-branched, do not have every.The fine and closely woven gentle song of aerial hyphae, the top spirrillum, Failure to thrive is contained, easy fracture.Late stage of culture, the aerial hyphae maturation is divided into spore chain, and few spiral is loose.Gram-positive;
(B) cultural characteristic: at the initial stage, bacterial strain is smooth at the substratum upper surface, and apricot does not have tangible aerial hyphae, does not produce soluble pigment.Along with the increase of incubation time, the bacterium colony surface is shallow splitting downwards, produces the white aerial hyphae of chestnut meat, is the unwrapping wire shape.The bacterium colony surface produces a spot of dewdrop shape secretory product.
(C) physiological and biochemical property: milk is solidified, can not peptonize, litmus milk is reddened; Hydrolyzable starch; The Phospholipid hydrolase of laying eggs; Do not produce hydrogen sulfide; Produce melanochrome; Can not make gelatin hydrolysis; Produce cellulase; Can reduce nitrate.15～37 ℃ of growth temperatures, optimum growth temperature are 28 ℃; Can utilize dextrin, glucose, sucrose, D-fructose, semi-lactosi, maltose, glycerine, L-arabinose, L-rhamnosyl, inositol, wood sugar, sorbyl alcohol; Can not utilize seminose, sodium hippurate, raffinose.Can be nitrogenous source with glycine, L-Ala, tryptophane; Can not utilize Methionin, L-halfcystine, Gelucystine, citrulline, urea, D, L-tyrosine, aspartic acid.
(D) 16S rDNA sequential analysis: the result carries out maximum homology relatively in GeneBank, finds that 16s rDNA sequencing result and Saccharothrix 16S rDNA sequence homology reach 99%.
(E) the Hhs.015 bacterial strain shows that to similar strain hybrid result two strain bacterium dna homologies are 33.81%, and homology requires to be lower than 70% between not of the same race, so Hhs.015 is novel species of Saccharothrix.
A further object of the invention provides the thalline of Hhs.015 bacterial strain, it prepares according to following method: with the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder liquid medium of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, fermented liquid is behind the centrifugal 20min of 5000rpm, and the gained throw out is the thalline of Yang Ling saccharothrix Hhs.015 bacterial strain.
Described analysis for soybean powder liquid medium is by soyflour 20g (boiling water boils 30min, filters to keep filtrate), sucrose 10g, Zulkovsky starch 5g, peptone 2g, yeast extract paste 2g, NaCl2g, CaCO
31g, MgSO
4-7H
2O0.5g, KH
2PO
40.5g, agar 15g, H
2O 1000mL forms, and the pH value is 7.2.
A further object of the invention provides the antibacterial substance of Hhs.015 bacterial strain, it prepares according to following method: with the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder nutrient solution of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, gained bacterium liquid is through the centrifugal 20min of 5000rpm, and the material after the gained supernatant liquor concentrates is slightly carries actives.
The antibacterial substance composition of Hhs.015 bacterial strain of the present invention is accredited as the antibiotics material by analysis.
A further object of the invention provides the application of Hhs.015 bacterial strain in preventing and treating leaf muld of tomato.
A further object of the invention provides the application of thalline in preventing and treating leaf muld of tomato of Hhs.015 bacterial strain.
A further object of the invention provides the application of antibacterial substance in preventing and treating leaf muld of tomato of Hhs.015 bacterial strain.
Hhs.015 bacterial strain of the present invention has stronger anti-leaf muld of tomato bacterium activity, has possessed the dual nature of antagonism bacterium as the biological and ecological methods to prevent plant disease, pests, and erosion factor, is a kind of biological pesticide that potentiality to be exploited is arranged very much, can be used for the control of leaf muld of tomato.
Hhs.015 bacterial strain of the present invention has stable natural matter source, and bacterial classification derives from the cucumber root, and screening method is simple; The preparation of saccharothrix thalline and antibacterial substance is simple, easy to operate, constant product quality, and low toxicity to people and animals' toxicological harmless, does not have poisoning to farm crop, environmental protection, it is remarkable to prevent and treat leaf muld of tomato disease effect.
Description of drawings
Fig. 1 is that saccharothrix bacterial strain Hhs.015 thalline and antibacterial substance thereof are to restraining effect figure in the ware of tomato leaf mold.
Fig. 2 is the thin-layer chromatogram that saccharothrix bacterial strain Hhs.015 active substance is formed.
Fig. 3 is the contrast figure of saccharothrix bacterial strain Hhs.015 field control effect.
Embodiment
The preparation embodiment and the result of use test example that provide below by the contriver are described in further detail the present invention, but content of the present invention is not limited thereto.
Separation, the evaluation of embodiment 1:Hhs.015 bacterial strain
Radix Cucumidis sativi picks up from Yang Ling new and high technology demonstration area, Shaanxi green house of vegetables, launch to dry with tap water flushing back, the root segment sample of getting plant respectively takes by weighing 1g to carry out surface sterilization and (soaks 1min in 99% ethanol, in 3.125% NaOCl, soak 6min then, put into 99% ethanol once more and soak 30sec, use aseptic water washing at last 5 times).Whether the sterilized water that washed is for the last time got 200 μ L be coated on the Gause I flat board, it is thorough to be used to detect surface sterilization.Material behind the cancellation poison is cut into the thin slice of 0.5 * 0.5mm size with the aseptic operation cutter, Radix Cucumidis sativi tissue block after the surface sterilization placed contain on the Gause I flat board, add up the saccharothrix colony number and carry out purifying after cultivating 21d in 28 ± 1 ℃ of incubators, separable to the Hhs.015 bacterial strain, obtained strains adopts the glycerine method to be preserved in-80 ℃ of refrigerators.
With the streak inoculation of saccharothrix bacterial strain on the analysis for soybean powder solid medium, cultivating behind the 5d bacterium piece of beating cut-off footpath 6mm with punch tool for 28 ℃ is inoculated in the saccharothrix fermentation culture, 28 ℃, after the 150r/min shaking table is cultivated 2d, inoculum size with 10% inserts in the saccharothrix fermentation culture carries out Secondary Fermentation, 28 ℃, the 150r/min shaking table is cultivated 5d.Fermented liquid is at the centrifugal 20min of 5000r/min, and it is standby to get supernatant liquor.The sterilized water suspension of precipitation with supernatant liquor equivalent is thallus suspension liquid, and concentration is 10
8～10
11Cfu/mL.
Brush leaf muld of tomato bacterium spore down from the tomato leaf of field morbidity, be made into concentration and be about 1 * 10
7～10 * 10
7The spore suspension of cfu/mL, the spray inoculation tomato seedling.Behind the natural air drying, spray saccharothrix supernatant liquor and thallus suspension liquid respectively, every processing 1 strain tomato seedling repeats 3 times, is contrast with the clear water.Place the greenhouse to preserve moisture and cultivate 48h under the condition, the sick number of sheets of 15d " Invest, Then Investigate " is calculated disease index and relative control effect.Investigation and method of calculation are with reference to " pesticide field efficacy medicine test criterion ".
The grade scale of leaf muld of tomato (is unit with the blade) adopts 9 grades of state of an illness grade scales of present domestic unification.That is:
0 grade: asymptomatic;
1 grade: the inoculation leaf chlorisis occurs to macula lutea occurring;
3 grades: produce thin sparse mould layer on the inoculation leaf disease spot;
5 grades: produce obvious mould layer on the inoculation leaf disease spot;
7 grades: produce dense mould layer on the inoculation leaf disease spot, upper blade is also infected;
9 grades: on inoculation leaf disease spot, have mould layer of the dense upper blade of mould layer also clearly.
According to the live test investigation result of Hhs.015 bacterial strain to leaf muld of tomato, screen a kind of active bacterial strain of efficient diseases prevention, through the microscopic examination of bacterium colony and form, cytochemistry compositional analysis, the experiment of conventional physio-biochemical characteristics, 16S rDNA sequential analysis and to similar strain hybrid, this identification of strains is new kind Saccharothrix yanglingensis Hhs.015 of Saccharothrix.
The preparation method of embodiment 2:Hhs.015 bacterial strain thalline
With the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder liquid medium of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, fermented liquid is behind the centrifugal 20min of 5000rpm, and the gained throw out is the thalline of Yang Ling saccharothrix Hhs.015 bacterial strain.
Described analysis for soybean powder liquid medium is by soyflour 20g (boiling water boils 30min, filters to keep filtrate), sucrose 10g, Zulkovsky starch 5g, peptone 2g, yeast extract paste 2g, NaCl2g, CaCO
31g, MgSO
47H
2O0.5g, KH
2PO
40.5g, agar 15g, H
2O 1000mL forms, and the pH value is 7.2.
The preparation method of the antibacterial substance of embodiment 3:Hhs.015 bacterial strain
With the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder nutrient solution of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, gained bacterium liquid gained supernatant liquor behind the centrifugal 20min of 5000rpm concentrates the material that obtains.
Test routine 1:Hhs.015 bacterial strain thalline and slightly propose the stripped bacteriostatic activity experiment of antibacterial substance
Get leaf muld of tomato bacterium PDA inclined-plane, be made into suspension, draw 100 μ L and be uniformly coated on the culture dish that contains 15mL PDA with the 10mL sterilized water.Hhs.015 bacterial strain thalline described in the embodiment 2 is inoculated in the Gause I flat board, grows 5 days; Block hitting; The evenly anti-saccharothrix bacterium piece that pastes 3 diameter 6mm in the distance culture dish is cultivated down for 25 ℃.Treat to measure when the target bacterium is covered with culture dish antibacterial circle diameter.
The leaf muld of tomato bacterium is seeded on the PDA inclined-plane, cultivates 3d for 25 ℃.Target bacterial strain mixing with on 10mL sterilized water and the inclined-plane is made into thallus suspension liquid, and behind 50mL PDA substratum mixing, every ware 15mL is tiled in the culture dish of diameter 9mm.In culture dish, evenly place the Oxford cup of 3 sterilizations, draw the antibacterial substance liquid described in the 200 μ L embodiment 3 in the cup of Oxford.Culture dish is cultivated 3～5d in 25 ℃ of incubators after, measure antibacterial circle diameter.
The result shows, Hhs.015 bacterial strain thalline and antibacterial substance are to remarkable (Fig. 1 of restraining effect of leaf muld of tomato bacteria growing, a left side: Hhs.015 bacterial strain thalline is to the restraining effect of leaf muld of tomato bacteria growing, and the right side: antibacterial substance is to the restraining effect of leaf muld of tomato bacteria growing).
Test example 2: the analysis of active substance
Antibacterial substance described in the embodiment 3 carries out thin-layer chromatography (Fig. 2), and to wherein several tangible bands recovery carrying out bacteriostatic activities experiment, the result shows that polycomponent has the activity that suppresses the growth of tomato leaf mold.
Test routine 3:Hhs.015 bacterial strain thalline and active substance and prevent and treat the potted plant experiment of leaf muld of tomato
With the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder nutrient solution of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, gained bacterium liquid gained supernatant liquor behind the centrifugal 20min of 5000rpm carries out spissated material.Precipitation is made spore suspension with sterilized water, and concentration is 10
8～10
9Cfu/mL.
Brush leaf muld of tomato bacterium spore down from the tomato leaf of field morbidity, be made into concentration and be about 1 * 10
7～10 * 10
7The spore suspension of cfu/mL, the spray inoculation tomato seedling.Behind the natural air drying, spray saccharothrix supernatant liquor and thallus suspension liquid respectively, every processing 1 strain tomato seedling repeats 3 times, is contrast with the clear water.Place the greenhouse to preserve moisture and cultivate 48h under the condition, the sick number of sheets of 15d " Invest, Then Investigate " is calculated disease index and relative control effect.Investigation and method of calculation are with reference to " pesticide field efficacy medicine test criterion ".
The grade scale of leaf muld of tomato (is unit with the blade) adopts 9 grades of state of an illness grade scales of present domestic unification.That is:
0 grade: asymptomatic;
1 grade: the inoculation leaf chlorisis occurs to macula lutea occurring;
3 grades: produce thin sparse mould layer on the inoculation leaf disease spot;
5 grades: produce obvious mould layer on the inoculation leaf disease spot;
7 grades: produce dense mould layer on the inoculation leaf disease spot, upper blade is also infected;
9 grades: on inoculation leaf disease spot, have mould layer of the dense upper blade of mould layer also clearly.
The result's mensuration of preventing and treating of different spraying times is undertaken by following processing.
Dispenser in advance: get three leaves, the tomato seedling of one heart stage, spraying concentration 1 * 10
7～10 * 10
7The tomato leaf mould spores suspension of cfu/mL behind the 48h that preserves moisture, sprays Hhs.015 bacterial strain fermentation liquor supernatant liquor again.
Postpone dispenser: get three leaves, the tomato seedling of one heart stage, spray bacterial strain Hhs.015 fermented liquid supernatant liquid, behind the cultivation 48h that preserves moisture, spraying concentration 1 * 10
7～10 * 10
7The tomato leaf mould spores suspension of cfu/mL is preserved moisture and is cultivated 48h.
Inoculation simultaneously: first inoculum density 1 * 10
7～10 * 10
7The tomato leaf mould spores suspension of cfu/mL treats to spray after tomato leaf dries bacterial strain Hhs.015 fermented liquid supernatant liquid again.
More than test every processing 1 strain tomato, 5 repetitions.Place the greenhouse to preserve moisture and cultivate under the condition, the sick number of sheets of 15d " Invest, Then Investigate " is calculated disease index and relative control effect.Investigation and method of calculation reference literature carry out.
The result shows that in the pot experiment, saccharothrix strain Hhs.015 supernatant liquor is higher to the relative control effect of leaf muld of tomato, reaches 49.7%, and the bacteria suspension preventive effect reaches 14.8%.And no matter be to shift to an earlier date dispenser or postpone dispenser, antagonism saccharothrix bacterial strain Hhs.015 has the better prevention effect to leaf muld of tomato, its relative control effect reaches 62.6% and 50.6% respectively, and the prevention effect of dispenser in advance is apparently higher than the prevention effect (table 1) of stepping back dispenser.
Table 1 saccharothrix bacterial strain Hhs.015 is to the preventive and therapeutic effect of leaf muld of tomato
Test routine 4:Hhs.015 bacterial strain active substance field efficiency test
Test design, efficacy survey and method of calculation are undertaken by document.Carry out radix investigation and dispenser for the first time earlier, dispenser for the second time behind the 10d.Again through 10d investigation drug effect.4 processing are established in test: bacterial strain BAR1-5 fermented liquid supernatant liquid (B1), and bacterial strain Hhs.015 fermented liquid supernatant liquid dilution 5 times of liquid (B2), 72% gram reveals wettable powder (be Shanghai E.I.Du Pont Company product, main component is a cymoxanil mancozeb), is contrast with the clear water.
The grade scale of leaf muld of tomato (is unit with the blade) grade scale is with test example 3.
Field test results shows that saccharothrix bacterial strain Hhs.015 still keeps preventive effect preferably, and relative control effect reaches 42.5%, and the relative control effect that the dilution of fermented liquid supernatant liquid is 5 times has also reached 33.1%.The relative control effect that chemical pesticide 72% gram reveals wettable powder is 46.8%, and is only high by 9.2% than fermented liquid supernatant liquid (B1) effect.But analyze from effective constituent, used sample is that wherein antibacterial effective constituent is well below chemical pesticide, so its prevention effect is than agricultural chemicals (table 2) on the low side without the fermented liquid of purifying in this field test.
Table 2 bacterial strain Hhs.015 is to the field efficacy of leaf muld of tomato
Claims (8)
1. Yang Ling saccharothrix (Saccharothrix yanglingensis) Hhs.015.
2. prepare the method for the described Yang Ling saccharothrix of claim 1 bacterial strain, it is characterized in that, specifically may further comprise the steps:
1) Radix Cucumidis sativi to be separated is launched to dry with tap water flushing back, be cut into the thin slice of 0.5 * 0.5mm size after the surface sterilization with the aseptic operation cutter;
2) the Radix Cucumidis sativi tissue block after the surface sterilization is placed on the Gause I flat board, add up the actinomycetes colony number and carry out purifying behind the cultivation 21d in 28 ± 1 ℃ of incubators, separable to the Hhs.015 bacterial strain, in-80 ℃ of preservations.
3. the method for preparing Yang Ling saccharothrix Hhs.015 bacterial strain according to claim 2, it is characterized in that, the condition of described surface sterilization is: after the root of clip health plant sample is cleaned with clear water, in 99% ethanol, soak 1min, in 3.125% NaOCl, soak 6min then, put into 99% ethanol once more and soak 30sec, use aseptic water washing at last 5 times.
4. the described Yang Ling saccharothrix of claim 1 Hhs.015, it is characterized in that, it prepares its thalline according to following method: with the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder liquid medium of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, fermented liquid is behind the centrifugal 20min of 5000rpm, and the gained throw out is the thalline of Yang Ling saccharothrix Hhs.015 bacterial strain;
Described analysis for soybean powder liquid medium is by soyflour 20g (boiling water boils 30min, filters to keep filtrate), sucrose 10g, Zulkovsky starch 5g, peptone 2g, yeast extract paste 2g, NaCl 2g, CaCO
31g, MgSO
47H
2O0.5g, KH
2PO
40.5g, agar 15g, H
2O 1000mL forms, and the pH value is 7.2.
5. the described Yang Ling saccharothrix of claim 1 Hhs.015, it is characterized in that, it prepares its antibacterial substance according to following method: with the Yang Ling saccharothrix Hhs.015 inoculation of-80 ℃ of preservations on the Gause I substratum, behind 28 ℃ of cultivation 4d, with its single colony inoculation in 100mL analysis for soybean powder liquid medium, in 28 ℃, the dark shaking culture 48h of 150rpm, be inoculated in the analysis for soybean powder nutrient solution of 100mL according to 1% seed liquor then, in 28 ℃, the dark shaking culture 120h of 150rpm, gained bacterium liquid is behind the centrifugal 20min of 5000rpm, and the gained supernatant liquor carries out spissated material and is antibacterial substance.
6. the application of the described Yang Ling saccharothrix of claim 1 Hhs.015 in preventing and treating leaf muld of tomato.
7. the application of the described Yang Ling saccharothrix of claim 4 Hhs.015 thalline in preventing and treating leaf muld of tomato.
8. the application of the antibacterial substance of the described Yang Ling saccharothrix of claim 5 Hhs.015 in control tomato leaf mold.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102643767A (en) * | 2012-04-19 | 2012-08-22 | 郑州大学 | Lactobacillus plantarum and application thereof in fermenting and ensiling sweet potato stem and leaf |
| CN103614488A (en) * | 2013-12-16 | 2014-03-05 | 盎亿泰地质微生物技术（北京）有限公司 | Polymerase chain reaction (PCR) primer for saccharothrix and method and kit for detecting saccharothrix |
| CN103667116A (en) * | 2013-11-13 | 2014-03-26 | 西北农林科技大学 | Saccharothrix yanglingensis for preventing apple valsa canker, and preparation method thereof |
-
2009
- 2009-12-15 CN CN2009102195094A patent/CN101955894A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102643767A (en) * | 2012-04-19 | 2012-08-22 | 郑州大学 | Lactobacillus plantarum and application thereof in fermenting and ensiling sweet potato stem and leaf |
| CN103667116A (en) * | 2013-11-13 | 2014-03-26 | 西北农林科技大学 | Saccharothrix yanglingensis for preventing apple valsa canker, and preparation method thereof |
| CN103614488A (en) * | 2013-12-16 | 2014-03-05 | 盎亿泰地质微生物技术（北京）有限公司 | Polymerase chain reaction (PCR) primer for saccharothrix and method and kit for detecting saccharothrix |
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Application publication date: 20110126 |