CN105462850A - Application of sophora tonkinensis endophytic fungus SDTE-P in preventing and controlling panax notoginseng root rot - Google Patents

Application of sophora tonkinensis endophytic fungus SDTE-P in preventing and controlling panax notoginseng root rot Download PDF

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CN105462850A
CN105462850A CN201510979167.1A CN201510979167A CN105462850A CN 105462850 A CN105462850 A CN 105462850A CN 201510979167 A CN201510979167 A CN 201510979167A CN 105462850 A CN105462850 A CN 105462850A
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sdte
sophora tonkinensis
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黄荣韶
姚裕群
蓝芳
李良波
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Guangxi University
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Abstract

The invention discloses a sophora tonkinensis endophytic fungus SDTE-P. The sophora tonkinensis endophytic fungus SDTE-P is classified and named as Penicillium citrinum SDTE-P, wherein the ITS sequence of the strain is as described by SEQ ID NO.1, the depository authority is the General Microbiological Collection Center of China Committee for Culture Collection of Microorganisms, the depository address is Institute of Microbiology, Building 3, No.1, Beichen West Road, Chaoyang District, Beijing, the depository date is 13 May 2015, and the depository number is CGMCC No.10808. According to the sophora tonkinensis endophytic fungus SDTE-P provided by the invention, a strain of endophytic fungus Penicillium citrinum SDTE-P is separated and screened from the root of a medicinal plant sophora tonkinensis for the first time, the Penicillium citrinum SDTE-P has a strong inhibiting effect on panax notoginseng fusarium solani, and a wide application prospect is created for the biological prevention and control field of panax notoginseng fungal diseases.

Description

The application of sophora tonkinensis Gapnep endogenetic fungus SDTE-P in control notoginseng root rot
Technical field
The present invention relates to biological technical field, the particularly application of a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P in control notoginseng root rot.
Background technology
The control of modern agriculture to Plant diseases is overly dependent upon the use of chemical pesticide, discharging of a large amount of chemical pesticide not only has long-term destruction to ecotope, also cause quality of agricultural product to decline, pesticide residue exceed standard, the resistance of pathogenic bacteria and to problems such as people and animals are harmful.Find safer, effective pest control method to be significant, utilizing biological method to carry out controlling plant diseases can effectively solve the problem.
Pseudo-ginseng (PanaxnotoginsengF.H.chen) has another name called pseudo-ginseng, invaluable etc., and having promoting blood circulation and removing blood stasis, subduing swelling and relieving pain effect significantly, is a kind of Chinese tradition rare medicinal herbs.Be that the Chinese patent medicine that main raw material is made is as wide-spread in " Yunnan white powder " and " Pien Tze Huang " etc. with pseudo-ginseng.In recent years, growing to the raw-material demand of pseudo-ginseng.But the fungal disease in cultivation has had a strong impact on Panax notoginseng Growth.At present, on notoginseng planting, control fungal disease depends on chemical pesticide unduly, and the use of a large amount of chemical pesticide not only have impact on the quality of pseudo-ginseng, also result in a large amount of pesticide residue of pseudo-ginseng.
Notoginseng root rot is one of pseudo-ginseng Major Diseases, is commonly called as " green smelly " in producing region, and cardinal symptom is that the bud-rot in seedling stage and the brown water stain shape pathology of reed head and bastem junction thereof are until spread to whole cane base portion.Just there is generation in this disease seeding stage, 4, May disease stagnate, the 6-9 month enters rainy season, enters onset peak period.Root rot has broad variety, it is reported that its symptom is mostly caused by notoginseng root rot bacterium (Fusariumsolani) (being called for short F.solani).
This fungal disease is the one of the main reasons causing pseudo-ginseng quality product and production declining for many years.In traditional Genuine producing area of Guangxi pseudo-ginseng, due to the meteorological conditions of low altitude area and high temperature and humidity, this disease is simultaneous often, and a large amount of underproduction causing pseudo-ginseng are even had no harvest, and brings huge financial loss to plantation family.In order to control this fungal disease, a large amount of chemical pesticides is used, and cause a large amount of pesticide residue of pseudo-ginseng product, serious pollutes environment, has threatened the health of people greatly.Therefore, it is very urgent and necessary for carrying out the Sustainable development of biological control pseudo-ginseng fungal disease to pseudo-ginseng industry.Filter out for this reason and can the Antagonistic Fungi of simultaneously this disease of antagonism be significant, screening and then to develop Antagonistic Fungi be also effectively control one of the most potential prophylactico-therapeutic measures of pseudo-ginseng fungal disease.
The information being disclosed in this background technology part is only intended to increase the understanding to general background of the present invention, and should not be regarded as admitting or imply in any form that this information structure has been prior art that persons skilled in the art are known.
Summary of the invention
The object of the present invention is to provide the application of a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P in control notoginseng root rot, thus the notoginseng root rot occurred in energy effectively preventing notoginseng planting process, thus improve output and the quality of pseudo-ginseng.
For achieving the above object, technical scheme provided by the invention is as follows:
A kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P, the Classification And Nomenclature of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is Penicillium citrinum (Penicilliumcitrinum) SDTE-P, the ITS sequence of bacterial strain is as described in SEQIDNO.1, depositary institution: China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, preservation date: on 05 13rd, 2015, preserving number: CGMCCNo.10808.Preferably, the preparation method of described sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites is:
(1) sophora tonkinensis Gapnep endogenetic fungus SDTE-P is inoculated in potato dextrose agar plane ware (PDA is dull and stereotyped), being placed in temperature is cultivate 10 days at 28 DEG C, gained cultivated material is cut into bulk and transfers in sterile solid substratum, is placed in 28 DEG C of fermentations 40 days;
(2) after having fermented, with the methyl alcohol soaking fermentation thing of fermented product 2 times and ultrasonic 40min, then filter;
(3) repeating step (2) twice, merges twice filtrate and carries out concentrating under reduced pressure and become medicinal extract, obtain the methanol crude extract of endogenetic fungus SDTE-P fermented product, be sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites.
Preferably, the sterile solid substratum described in step (1) contains 400g potato, the dextrose of 20g and 20g sucrose.
The application of meta-bolites in control notoginseng root rot of gained sophora tonkinensis Gapnep endogenetic fungus SDTE-P is prepared as above-mentioned.
Compared with prior art, the present invention has following beneficial effect:
The present invention first from the root of medicinal plant sophora tonkinensis Gapnep separation screening to strain endogenetic fungus (Penicilliumcitrinum) SDTE-P, this bacterium has very strong restraining effect to notoginseng root rot bacterium, and the field of biological control for pseudo-ginseng fungal disease brings wide application prospect.
Accompanying drawing explanation
Fig. 1 is the dull and stereotyped opposite culture of sophora tonkinensis Gapnep endogenetic fungus SDTE-P of the present invention and notoginseng root rot, and wherein F is notoginseng root rot bacterium (Fusariumsolani), a is blank, and b is experimental group.
Fig. 2 is sophora tonkinensis Gapnep endogenetic fungus SDTE-P strain morphology feature of the present invention, and wherein, a1 is colonial morphology, and b1 is thalli morphology.
Fig. 3 is the systematic evolution tree of sophora tonkinensis Gapnep endogenetic fungus SDTE-P bacterial strain based on ITS sequence.
Fig. 4 is to the inhibition of the mycelial growth of notoginseng root rot bacterium according to the meta-bolites of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P of the present invention; Wherein the 1st, the 5th is that namely the PDA of pastille is not dull and stereotyped for blank; 2nd, the 3rd, the 4th is positive control powder of carbendazim; 6th, the 7th, the 8th is the meta-bolites of bacterial strain SDTE-P, and concentration is respectively 2mg/ml, 4mg/ml, 8mg/ml; F is notoginseng root rot bacterium Fusariumsolani.
Embodiment
Be described in detail below in conjunction with embodiment, but be to be understood that protection scope of the present invention not by the restriction of embodiment.The experimental technique used in following embodiment if no special instructions, is ordinary method.The material used in following embodiment, reagent etc., if no special instructions, all can obtain from commercial channels.Methyl alcohol is commercial analytical pure methyl alcohol.2XTagMasterMix is purchased from precious biotechnology company limited (Takara), and Primer-1, Primer-2 are by Hua Da gene chemical synthesis.
Embodiment 1
The separation of bacterial strain, screening and qualification
One, the separation of bacterial strain
For examination material: the wild sophora tonkinensis Gapnep picking up from In Limestone Area, Tiandeng County, Guangxi.
Strains tested: provided by plant pathology institute of Guangxi University.
Substratum: 1000ml potato dextrose agar (PDA substratum), PDA substratum is containing potato 300g, glucose 20g, agar 20g and paraxin 0.1g, and Medium's PH Value is 6.0 ± 0.2.
Surface sterilization: by long for 6-8cm, the wide sophora tonkinensis Gapnep root running water 30min for 1-2cm fresh and healthy are to rinse silt, then the sophora tonkinensis Gapnep root rinsed with sterile water 2 times will cleaned, air-dry surface-moisture, moving on to Bechtop, aseptically, is 75% alcohol immersion 1min by sophora tonkinensis Gapnep root volumetric concentration, rinsed with sterile water 2 times, clorox (available chlorine 1%) soaks 2min, aseptic washing 3 times, and it is for subsequent use that aseptic thieving paper blots surface.
The separation of bacterial strain, purifying: the sophora tonkinensis Gapnep root that surface sterilization is good, under aseptic condition, epidermis is scraped off with aseptic wood chip, the two ends of sophora tonkinensis Gapnep root are cut off with aseptic secateurs, remaining part aseptic pocket knife and aseptic nipper separate xylem and phloem, and be cut into the long tissue block of 0.5cm with aseptic secateurs, be asepticly forwarded to the PDA substratum (containing paraxin) prepared, 28 DEG C of cultivations.Getting the last rinsing liquid of surface sterilization is coated on PDA flat board, as negative control.Every day is observed, and waits to organize to grow mycelia, the single mycelia of picking around, is connected to the PDA substratum of antibiotic-free.The bacterium colony grown, if colonial morphology is inconsistent, continues picking single mycelia switching PDA substratum, until the formalness of the bacterium colony that flat board newly grows is consistent.
The endogenetic fungus be separated to is inoculated in PDA flat board, cultivates after 20 days stand-by at 28 DEG C.Notoginseng root rot bacterium is inoculated in PDA flat board, cultivates after 3 days stand-by at 28 DEG C.
Two, screen
Adopt dull and stereotyped face-off method to the primary dcreening operation carrying out thalline bacteriostatic activity for examination sophora tonkinensis Gapnep endogenetic fungus.
First, under aseptic condition, with sterilizing punch tool, endogenetic fungus and notoginseng root rot bacterium are made 6mm bacterium cake; In treatment group, endogenetic fungus bacterium cake is transferred to the culture dish central authorities containing PDA, then around the bacterium cake of endogenetic fungus, notoginseng root rot bacterium bacterium cake is placed at isometrical 3cm place, and every strain endogenetic fungus repeats 3 wares; In control group, the culture dish central authorities of PDA do not connect endogenetic fungus bacterium cake, and around the culture dish central authorities of PDA, notoginseng root rot bacterium bacterium cake is placed at isometrical 3cm place, repeats 3 wares.Then cultivate at 28 DEG C, periodic observation.Treat that in control group, mycelia grows to the dull and stereotyped central authorities of culture dish of PDA, in treatment group, measure the growth radius of notoginseng root rot bacterium between notoginseng root rot Jun Junbing center to endogenetic fungus Jun Bing center for process growth radius; In control group, measure notoginseng root rot Jun Junbing center to PDA culture dish center between notoginseng root rot bacterium growth radius for contrast grow radius.Finally, according to following formula, calculate bacteriostasis rate:
Contrast increment=contrast growth radius-bacterium cake radius
Process increment=process growth radius-bacterium cake radius
Result obtains the 4 strains antagonism sophora tonkinensis Gapnep endogenetic fungal bacterial strain all very strong to notoginseng root rot bacterium restraining effect altogether, and wherein a strain name is called SDTE-P, is 60% to the inhibiting rate of notoginseng root rot bacterium.
Three, identify
(1) strain morphology feature
Morphological features is observed: be seeded on PDA substratum by endophyte fungal bacterial strain SDTE-P to be identified, be placed in 28 DEG C of cultivations, observe its cultural characteristic and colour-change respectively at 5,14 and 20 days.Get stable ripe color characteristic as its cultural characteristic, as the foundation of qualification.The color of observed and recorded aerial hyphae, the size, color, tissue profile, surface shape etc. of bacterium colony are as reference feature.
Spore shape observation of characteristics: endophyte fungal bacterial strain SDTE-P to be identified is made inserted sheet and cultivate, when not producing spore on PDA substratum, mycelia is transferred on the low nutrition substratum (1/4th concentration PDA) containing aseptic sophora tonkinensis Gapnep root with inducing spore, by the photomicrography of viewed form result, as shown in Figure 2.
(2) bacterial strain ITS sequence and phylogenetic analysis thereof
The preparation of DNA profiling:
Reagent: (1) lysis buffer: Tris-Ac (pH7.8) 40mM, NaAc20mM, EDTA1mM, SDS1% (w/v);
(2) 5MNaCl solution.
DNA extraction:
Add 600 μ L extracting solution (Tris-Ac (pH7.8) 40mM, NaAc20mM, EDTA1mM, SDS1%) to 1.5ml centrifuge tube (EP pipe), scrape with rod the mycelia that takes a morsel and put into the grinding of EP pipe, 65 DEG C of water-bath 30min, centrifugal rotational speed 12000r/min, centrifugal 10min;
Get centrifugal gained supernatant liquor 400 μ L, add 5MNaCL100 μ L, ice bath 10min;
Then at temperature is 4 DEG C, keep centrifugal rotational speed to be 12000r/min, centrifugal 10min, gets supernatant liquor; Mix (or 2 times of dehydrated alcohols) ice bath 1h with supernatant liquor 0.6 times of Virahol and supernatant liquor, keep centrifugal rotational speed 12000r/min, centrifugal 10min, after abandoning supernatant, the remaining material of gained dries, and adds 20 μ L distilled water (ddH 2o), get 1-2 μ L and do DNA profiling.
Pcr amplification ITS sequence:
(1) PCR instrument: ABI3730-XLDNA sequenator (AppliedBiosystems, USA);
(2) amplimer: ITS1 (5 '-TCCGTAGGTGAACCTGCGG-3 ') is as shown in SEQIDNO.2, and ITS4 (5 '-TCCTCCGCTTATTGATATGC-3 ') is as shown in SEQIDNO.3;
(3) amplification system:
ITS sequence PCR amplification system is as shown in table 1: table 1.
Reactant Application of sample amount
2X TagMasterMix 25μL
Primer-1 1μL
Primer-2 1μL
Template DNA 2μL
ddH 2O 21μL
Reaction cumulative volume 50μL
TemplateDNA in table 1 above-mentionedly makes DNA profiling.
PCR reaction conditions is as shown in table 2:
Table 2.
Note: in table 2, step 2 carries out 30 circulations.
The electrophoresis detection of pcr amplification product:
Deposition condition is the sepharose (containing Goldview5 μ l/100ml) of 1%, 1 × TBE electrophoretic buffer, 90V electrophoresis 1 hour, and PCR primer applied sample amount is 3 μ L, point sample after mixing with 1 μ LLoadingdye.
Observations under 254nm ultraviolet, with the DL1000DNAMarker of TaKaRa company for nucleic acid standard molecular weight object of reference, determines expanding fragment length.Amplified production band should on the position of standard substance 400-700bp.
PCR primer purifying and order-checking: undertaken by Shenzhen Huada Genetic Technology Co., Ltd.
The structure of systematic evolution tree:
The ITS sequence of the fungi measured in surveyed ITS sequence and GenBank gene pool is compared, downloads correlated series according to comparison result.Systems analysis is carried out and constructing system evolutionary tree, as shown in Figure 3 by the adjacent algorithm (Neighbor-joiningNJ) of MEGA6.0.
Result:
(1) strain morphology feature
Colony morphology characteristic: on PDA substratum, bacterium colony is circular, and tool obviously takes turns line, neat in edge, and substrate mycelium white, the milky white carpet-like of aerial hyphae, the aleurioconidium group of bacterium colony surface attachment greyish-green, as shown in a1 in Fig. 2.
Spore shape feature: round, diameter 2.50 μm, as shown in b1 in Fig. 2.
(2) bacterial strain ITS sequence and phylogenetic analysis thereof
Utilize primer I TS1 and ITS4, from strain gene group DNA, amplify the fragment of a 400-500bp size, through order-checking and by sequencing result by BLASTn comparison in GenBank.Result shows, fungi during bacterial strain SDTE-P and Penicilliu belongs to has very high base sequence similarity, therefore the reference strain sequence that in GenBank, Penicilliu belongs to is downloaded, for Phylogenetic Analysis, the Monascusaurantiacus (AY629435) in belonging to using Monascus and Monascuspurpureus (AY629416) is as outer group.In the systematic evolution tree built, SDTE-P and Penicilliumcitrinum (KF624801) and Penicilliumcitrinum (KM491892) gets together and forms holding strength is the end branch of 100%.Base similarity system design result shows, SDTE-P and Penicilliumcitrinum (KF624801, KM491892) base sequence does not have difference, and sequence similarity is 100%.Comprehensive morphological and molecular biological characteristics, be initially identified as Penicilliumcitrinum by bacterial strain.
Embodiment 2
The meta-bolites of bacterial strain is to the restraining effect of notoginseng root rot bacterium
One, the fermentation culture of bacterial strain and the extraction of meta-bolites
(1) sophora tonkinensis Gapnep endogenetic fungus SDTE-P is inoculated in potato dextrose agar plane ware (PDA is dull and stereotyped), being placed in temperature is cultivate 10 days at 28 DEG C, gained cultivated material is cut into bulk and transfers to sterile solid substratum (containing 400g potato, the dextrose of 20g and 20g sucrose) the Erlenmeyer flask of 2 liters in, be placed in 28 DEG C of condition bottom fermentations 40 days;
(2) after having fermented, with the methyl alcohol soaking fermentation thing of fermented product 2 times and ultrasonic 40min, then use filtered through gauze, get filtrate;
(3) repeating step (2) twice, merges twice filtrate and carries out concentrating under reduced pressure and become medicinal extract, obtain the methanol crude extract of endogenetic fungus SDTE-P fermented product, be sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites.
Two, sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites is to the restraining effect of the mycelial growth of notoginseng root rot bacterium
The meta-bolites of bacterial strain SDTE-P is measured to the inhibit activities of notoginseng root rot bacterium mycelial growth by mycelial growth method.The pastille meta-bolites of bacterial strain SDTE-P and powder of carbendazim (positive control) made respectively containing 2mg/mL, 4mg/mL, 8mg/mL concentration meta-bolites is dull and stereotyped.Aseptically, with punch tool, notoginseng root rot bacterium is made 6mm bacterium cake, it is process that notoginseng root rot bacterium bacterium cake is received the dull and stereotyped central authorities of each pastille, the notoginseng root rot bacterium bacterium cake connect with the dull and stereotyped central authorities of not pastille is for negative control, in triplicate, all flat boards are placed in 28 DEG C of cultivations for all process and contrast.The Field information concentration that drug level presses powder of carbendazim is arranged.When negative control covers with culture dish, measure the growth diameter of contrast bacterium colony and process bacterium colony respectively, and according to following formula, calculate inhibiting rate:
Negative control increment=negative control growth diameter-bacterium cake diameter
Process increment=process growth diameter-bacterium cake diameter
Three, sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites is to the minimal inhibitory concentration of notoginseng root rot bacterium
With the meta-bolites of broth dilution method determination bacterial strain SDTE-P to the minimal inhibitory concentration of notoginseng root rot bacterium.The notoginseng root rot bacterium pure culture biscuits involvng inoculation of 5 days will be cultivated in the PDB liquid nutrient medium containing 0.2% tween 80 (v/v) in PDA substratum, 28 DEG C, 150r/min shaking table cultivates 7 days, then culture is forwarded to triangular flask, and the aseptic double-distilled water poured into containing 0.2% tween 80, stir 30min with magnetism stick, solution sterile gauze is filtered, obtain spores solution, and with blood counting chamber, spore concentration is adjusted to every milliliter 10 4individual spore is for subsequent use.The meta-bolites 1%DMSO of bacterial strain is dissolved into the meta-bolites concentration for the treatment of of 80mg/mL, getting 5 sterile test tube is sequentially arranged on test-tube stand, be numbered 1,2,3,4,5 respectively, often pipe adds the aseptic double-distilled water of 1ml containing 0.2% tween 80 respectively, is then added by the bacterial strain methanol crude extract solution of 1ml80mg/mL in the test tube being numbered 1 and also in each pipe, carries out twice serial dilution successively.The above-mentioned gained spores solution (10 of 1ml 4individual/ml) add in each pipe respectively, thus obtain the concentration for the treatment of of final SDTE-P meta-bolites: 20mg/ml (numbering 1 test tube), 10mg/ml (numbering 2 test tube), 5mg/ml (numbering 3 test tube), 2.5mg/ml (numbering 4 test tube), 1.25mg/ml (numbering 5 test tube).The 8mg/mL Flusilazole of 1%DMSO and 1ml of 1ml replaces meta-bolites to perform above-mentioned treating processes respectively and as negative control and positive control, all process and contrast are in triplicate.Finally, each pipe 28 DEG C, 150r/min shaking table cultivates 5 days.The MIC value of the meta-bolites of SDTE-P is suppress the minimum crude extract concentration of notoginseng root rot bacterium visible growth completely.
Result:
The percent inhibition of meta-bolites to notoginseng root rot bacterium mycelial growth of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P is as shown in table 3:
Table 3.
Note: show positives contrast powder of carbendazim containing 50% derosal; After in table, * represents that LSD that data pass through one-way analysis of variance is relatively, the meta-bolites of bacterial strain SDTE-P and positive control powder of carbendazim under same concentrations, at P 0.05tool significant difference in level.
The suppression result of meta-bolites to notoginseng root rot bacterium mycelial growth of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P shows, the meta-bolites of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P all has extraordinary inhibition to the mycelial growth of notoginseng root rot bacterium, and the percent inhibition of meta-bolites to notoginseng root rot bacterium F.solani mycelial growth of SDTE-P is 100% as can be seen from Table 3.Compare with positive control powder of carbendazim, the meta-bolites of bacterial strain SDTE-P, to the inhibition of notoginseng root rot bacterium F.solani mycelial growth with contrast equivalent.
The minimal inhibitory concentration of sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites to notoginseng root rot bacteria growing is as shown in table 4:
Table 4.
Process MIC(mg/ml)
F.solani
Flusilazole 0.5
SDTE-P 10
Note: showing positives contrast Flusilazole active constituent content is 400mg/mL.
The minimal inhibitory concentration test result of meta-bolites to notoginseng root rot bacteria growing of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P shows, the meta-bolites of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P all has stronger restraining effect to notoginseng root rot bacterium, the minimal inhibitory concentration of meta-bolites to notoginseng root rot bacterium F.solani of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is 10mg/ml as can be seen from Table 4, is 20 times of positive control.
As may be known from Table 3 and Table 4, contain the composition of very strong antifungal or fungicidal in the meta-bolites of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P, therefore, in the bionomic control of notoginseng root rot bacterium, bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P has outstanding potentiality.
The aforementioned description to concrete exemplary of the present invention is to illustrate and the object of illustration.These descriptions not want the present invention to be defined as disclosed precise forms, and obviously, according to above-mentioned instruction, can much change and change.The object selected exemplary embodiment and describe is to explain certain principles of the present invention and practical application thereof, thus those skilled in the art can be realized and utilize various different exemplary of the present invention and various different selection and change.Scope of the present invention is intended to limited by claims and equivalents thereof.

Claims (4)

1. a sophora tonkinensis Gapnep endogenetic fungus SDTE-P, is characterized in that: the Classification And Nomenclature of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is Penicillium citrinum (Penicilliumcitrinum) SDTE-P, preserving number: CGMCCNo.10808.
2. a preparation method for the meta-bolites of sophora tonkinensis Gapnep endogenetic fungus SDTE-P as claimed in claim 1, it is characterized in that, operation steps is as follows:
(1) be inoculated in potato dextrose agar plane ware by sophora tonkinensis Gapnep endogenetic fungus SDTE-P, being placed in temperature is cultivate 10 days at 28 DEG C, and gained cultivated material is cut into bulk and transfers in sterile solid substratum, is placed in 28 DEG C of fermentations 40 days;
(2) after having fermented, with the methyl alcohol soaking fermentation thing of fermented product 2 times and ultrasonic 40min, then filter;
(3) repeating step (2) twice, merges twice filtrate and carries out concentrating under reduced pressure and become medicinal extract, obtain the methanol crude extract of endogenetic fungus SDTE-P fermented product, be sophora tonkinensis Gapnep endogenetic fungus SDTE-P meta-bolites.
3. the preparation method of meta-bolites according to claim 2, is characterized in that: the sterile solid substratum described in step (1) containing 400g potato, the dextrose of 20g and 20g sucrose.
4. the application of meta-bolites in control notoginseng root rot preparing gained sophora tonkinensis Gapnep endogenetic fungus SDTE-P as claim 2.
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CN105462855A (en) * 2015-12-23 2016-04-06 广西大学 Application of sophora tonkinensis endophytic fungus SDTE-P in preventing and controlling panax notoginseng black spot
CN109112069A (en) * 2017-06-23 2019-01-01 沈阳药科大学 A kind of biological and ecological methods to prevent plant disease, pests, and erosion endogenetic fungus and its application
CN112679272A (en) * 2021-02-20 2021-04-20 长兴县中医院 Biological bacterial fertilizer for saffron crocus and preparation method thereof
CN114134051A (en) * 2021-12-21 2022-03-04 东北林业大学 Penicillium citrinum YW322, culture method, microbial inoculum and application thereof

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CN105462855A (en) * 2015-12-23 2016-04-06 广西大学 Application of sophora tonkinensis endophytic fungus SDTE-P in preventing and controlling panax notoginseng black spot
CN105462855B (en) * 2015-12-23 2019-06-21 广西大学 Application of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P in prevention and treatment Alternaria panax
CN109112069A (en) * 2017-06-23 2019-01-01 沈阳药科大学 A kind of biological and ecological methods to prevent plant disease, pests, and erosion endogenetic fungus and its application
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CN112679272A (en) * 2021-02-20 2021-04-20 长兴县中医院 Biological bacterial fertilizer for saffron crocus and preparation method thereof
CN114134051A (en) * 2021-12-21 2022-03-04 东北林业大学 Penicillium citrinum YW322, culture method, microbial inoculum and application thereof
CN114134051B (en) * 2021-12-21 2023-05-12 东北林业大学 Penicillium citrinum YW322, culture method, microbial inoculum and application thereof

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