CN105462854B - Application of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P in prevention and treatment Radix Notoginseng anthracnose - Google Patents

Abstract

The invention discloses a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P, the classification naming of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is Penicillium citrinum (Penicillium citrinum) SDTE-P, the ITS sequence of bacterial strain is as described in SEQ ID NO.1, depositary institution: China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 Institute of Microorganism, Academia Sinica, preservation date: on May 13rd, 2015, deposit number: CGMCC No.10808.The present invention for the first time from the root of medicinal plant sophora tonkinensis Gapnep separation screening to one plant of endogenetic fungus (Penicillium citrinum) SDTE-P, the bacterium has very strong inhibiting effect to Radix Notoginseng anthrax bacteria, brings wide application prospect for the field of biological control of Radix Notoginseng fungal disease.

Description

Application of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P in prevention and treatment Radix Notoginseng anthracnose

Technical field

The present invention relates to field of biotechnology, in particular to a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is in prevention and treatment Radix Notoginseng anthrax Application in disease.

Background technique

Modern agriculture is overly dependent upon the use of chemical pesticide to the prevention and treatment of plant disease, and the release of a large amount of chemical pesticides is not Only there is long-term destruction to ecological environment, also quality of agricultural product is caused to decline, excessive pesticide residues, the drug resistance of pathogen Property and to problems such as people and animals' nocuousness.It finds safer, effective pest control method to be of great significance, utilize The method of biology, which carrys out controlling plant diseases, effectively to solve the above problems.

Radix Notoginseng (Panax notoginseng F.H.chen) also known as pseudo-ginseng, invaluable etc., have significant activating microcirculation and removing stasis medicinal, Detumescence ding-tong effect is a kind of Chinese tradition rare medicinal herbs.Using Radix Notoginseng as Chinese patent drug made of primary raw material such as " Yunnan Baiyao " " Pien Tze Huang " etc. is wide-spread.In recent years, the demand to Radix Notoginseng raw material is growing.But the nosomycosis in cultivation Evil has seriously affected Panax notoginseng Growth.Currently, prevention and treatment fungal disease depends on chemical pesticide unduly on notoginseng planting, a large amount of chemistry The use of pesticide not only affects the quality of Radix Notoginseng, also results in a large amount of pesticide residues of pseudo-ginseng.

Radix Notoginseng anthracnose is one of Panax notoginseng Growth common disease, and seedling stage, Adult plant can fall ill, and blade early stage is shown as Blade gives birth to yellowish-brown spot, and later period disease portion easily perforates rupture.It will form yellowish-brown recess spot, fruit infection after petiole and stem infection Browning color rots afterwards, and the cause of disease of the disease is Colletotriehum gloeosporioides (abbreviation C.gloeosporioides)。

This fungal disease is one of the main reason for causing Radix Notoginseng product quality for many years and yield to decline.In Guangxi three Seven traditional Genuine producing area, due to low altitude area and the meteorological condition of high temperature and humidity, this disease is often simultaneous, causes The a large amount of underproduction of Radix Notoginseng are even had no harvest, and are had brought tremendous economic losses to farmer.In order to control this fungal disease, greatly The chemical pesticide of amount is used, and a large amount of pesticide residues of Radix Notoginseng product are caused, serious to polluted environment, is greatly threatened People's health.Therefore, development biological control Radix Notoginseng fungal disease is very urgent to the sustainable development of Radix Notoginseng industry and must It wants.The Antagonistic Fungi for filtering out energy while this disease of antagonism thus is of great significance, and screens and then develops and uses Antagonistic Fungi It is also effectively to control one of most potential control measure of Radix Notoginseng fungal disease.

The information disclosed in the background technology section is intended only to increase the understanding to general background of the invention, without answering When being considered as recognizing or imply that the information constitutes the prior art already known to those of ordinary skill in the art in any form.

Summary of the invention

The purpose of the present invention is to provide a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P to prevent and treat the application in Radix Notoginseng anthracnose, So as to the Radix Notoginseng anthracnose occurred during effectively preventing notoginseng planting, to improve the yield and quality of Radix Notoginseng.

To achieve the above object, technical solution provided by the invention is as follows:

The classification naming of a kind of sophora tonkinensis Gapnep endogenetic fungus SDTE-P, sophora tonkinensis Gapnep endogenetic fungus SDTE-P are Penicillium citrinum (Penicillium citrinum) SDTE-P, the ITS sequence of bacterial strain is as described in SEQ ID NO.1, depositary institution: the micro- life of China Object culture presevation administration committee common micro-organisms center, preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 China Institute of microbiology, the academy of sciences, preservation date: on May 13rd, 2015, deposit number: CGMCC No.10808.

Preferably, the sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite the preparation method comprises the following steps:

(1) sophora tonkinensis Gapnep endogenetic fungus SDTE-P is inoculated in potato dextrose agar plane ware (PDA plate) In, being placed in temperature is to cultivate 10 days at 28 DEG C, and gained culture materials are cut into bulk and are transferred in sterile solid culture medium, are placed in 28 DEG C ferment 40 days；

(2) after fermentation, the methanol soaking fermentation object and ultrasound 40min with 2 times of fermentation material, is then filtered；

(3) step (2) are repeated twice, merges filtrate twice and medicinal extract is concentrated under reduced pressure into, obtains endogenetic fungus SDTE-P The methanol crude extract of fermentation material, as sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite.

Preferably, sterile solid culture medium potato containing 400g described in step (1), the dextrose and 20g sugarcane of 20g Sugar.

Application of the metabolite of sophora tonkinensis Gapnep endogenetic fungus SDTE-P as obtained by above-mentioned preparation in prevention and treatment Radix Notoginseng anthracnose.

Compared with prior art, the invention has the following beneficial effects:

The present invention for the first time from the root of medicinal plant sophora tonkinensis Gapnep separation screening to one plant of endogenetic fungus (Penicillium Citrinum) SDTE-P, the bacterium have very strong inhibiting effect to Radix Notoginseng anthrax bacteria, are the biological control of Radix Notoginseng fungal disease Bring wide application prospect in field.

Detailed description of the invention

Fig. 1 is the plate opposite culture of sophora tonkinensis Gapnep endogenetic fungus SDTE-P of the present invention Yu Radix Notoginseng anthracnose, and wherein F is Radix Notoginseng Anthrax bacteria (Colletotriehum gloeosporioides), a are blank control, and b is experimental group.

Fig. 2 is sophora tonkinensis Gapnep endogenetic fungus SDTE-P strain morphology feature of the present invention, wherein a1 is colonial morphology, and b1 is bacterium Volume morphing.

Fig. 3 is systematic evolution tree of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P bacterial strain based on ITS sequence.

Fig. 4 is mycelia of the metabolite to Radix Notoginseng anthrax bacteria of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P according to the present invention The inhibitory effect of growth；Wherein the 1st, the 5th for blank control be not drug containing PDA plate；2nd, the 3rd, the 4th is positive right According to powder of carbendazim；6th, the 7th, the 8th is the metabolite of bacterial strain SDTE-P, and concentration is respectively 2mg/ml, 4mg/ml, 8mg/ ml；T is Radix Notoginseng anthrax bacteria Colletotriehum gloeosporioides.

Specific embodiment

It is described in detail With reference to embodiment, it is to be understood that protection scope of the present invention is not by specific The limitation of embodiment.Experimental method used in following embodiments is conventional method unless otherwise specified.Following implementations Material, reagent used in example etc., is commercially available unless otherwise specified.Methanol is the commercially available pure methanol of analysis. 2X TagMasterMix is purchased from precious bioengineering Co., Ltd (Takara), and Primer-1, Primer-2 are closed by Hua Da gene At.

Embodiment 1

Separation, the screening and identification of bacterial strain

One, the separation of bacterial strain

Material to be tested: the wild sophora tonkinensis Gapnep of Guangxi Tiandeng County In Limestone Area is picked up from.

Strains tested: it is provided by plant pathology research institute, Guangxi University.

Culture medium: 1000ml potato dextrose agar (PDA culture medium), PDA culture medium 300g containing potato, Glucose 20g, agar 20g and chloramphenicol 0.1g, Medium's PH Value are 6.0 ± 0.2.

Surface sterilization: a length of 6-8cm, width is dry to rush for the sophora tonkinensis Gapnep root flowing water flushing 30min of 1-2cm fresh and healthy Net silt air-dries surface moisture, superclean bench is moved on to, sterile then by clean sophora tonkinensis Gapnep root with rinsed with sterile water 2 times Under the conditions of, by sophora tonkinensis Gapnep root volumetric concentration be 75% ethyl alcohol impregnate 1min, rinsed with sterile water 2 times, sodium hypochlorite (effective chlorine 1%) 2min is impregnated, sterile washing 3 times, it is spare that sterile blotting paper blots surface.

The separation of bacterial strain purifies: the good sophora tonkinensis Gapnep root of surface sterilization, under aseptic condition, scrapes off epidermis with sterile wood chip, uses Sterile secateurs cuts off the both ends of sophora tonkinensis Gapnep root, and the sterile pocket knife of remaining part and aseptic nipper separate xylem and bast, and The tissue block of 0.5cm long is cut into sterile secateurs, it is sterile to be forwarded to the PDA culture medium (containing chloramphenicol) prepared, 28 DEG C of cultures. Surface sterilization last time rinsing liquid is taken to be coated on PDA plate, as negative control.Observation daily, wait be grown around organizing Mycelia, the single mycelia of picking, is connected to the PDA culture medium of antibiotic-free.The bacterium colony grown continues if colonial morphology is inconsistent The single mycelia switching PDA culture medium of picking, until the formalness of the bacterium colony newly grown on plate is consistent.

The endogenetic fungus being separated to is inoculated in PDA plate, it is stand-by after being cultivated 20 days at 28 DEG C.By Radix Notoginseng anthrax bacteria It is inoculated in PDA plate, it is stand-by after being cultivated 3 days at 28 DEG C.

Two, it screens

Using tablet face-off method to the primary dcreening operation for carrying out thallus bacteriostatic activity for examination sophora tonkinensis Gapnep endogenetic fungus.

Firstly, 6mm bacteria cake is made in endogenetic fungus and Radix Notoginseng anthrax bacteria with sterilization punchers under aseptic condition；Locating In reason group, endogenetic fungus bacteria cake is transferred to the center of the culture dish containing PDA, then the isometrical 3cm around the bacteria cake of endogenetic fungus Radix Notoginseng anthrax bacteria bacteria cake is placed at place, and every plant of endogenetic fungus repeats 3 wares；In control group, the culture dish center of PDA does not connect interior life Fungi bacteria cake places Radix Notoginseng anthrax bacteria bacteria cake at the culture dish center isometrical 3cm of surrounding of PDA, repeats 3 wares.Then 28 DEG C Lower culture, periodic observation.Radix Notoginseng anthrax is measured in processing group to the long culture dish plate center to PDA of mycelia in control group The growth radius of germ bacteria cake center to Radix Notoginseng anthrax bacteria between endogenetic fungus bacteria cake center is processing growth radius；It is compareing In group, the growth radius at measurement Radix Notoginseng anthrax bacteria bacteria cake center to Radix Notoginseng anthrax bacteria between the culture dish center of PDA is pair According to growth radius.Finally, according to the following formula, calculating bacteriostasis rate:

It compares increment=control and grows radius-bacteria cake radius

It handles increment=processing and grows radius-bacteria cake radius

As a result 4 plants are obtained to all very strong antagonism sophora tonkinensis Gapnep endogenetic fungal bacterial strain of Radix Notoginseng anthrax bacteria inhibiting effect, In one plant of entitled SDTE-P, be 59% to the inhibiting rate of Radix Notoginseng anthrax bacteria.

Three, it identifies

(1) strain morphology feature

Morphological features observation: endophyte fungal bacterial strain SDTE-P to be identified is seeded in PDA culture medium, is placed in 28 DEG C of cultures, observed its cultural characteristic and color change at 5,14 and 20 days respectively.Take the color characteristic for stablizing maturation as it Cultural characteristic, the foundation as identification.Observe and record the color of aerial hyphae, the size of bacterium colony, color, tissue profile, surface Shape etc. is used as fixed reference feature.

Spore shape observation of characteristics: endophyte fungal bacterial strain SDTE-P to be identified is made into inserted sheet culture, is cultivated when in PDA Spore is not produced on base, by mycelia switching on the low nutrition culture medium (a quarter concentration PDA) containing sterile sophora tonkinensis Gapnep root to lure Spore is led, by observed form result microphotograph, as shown in Figure 2.

(2) bacterial strain ITS sequence and its phylogenetic analysis

The preparation of DNA profiling:

Reagent: (1) lysis buffer: 1% (w/ of Tris-Ac (pH 7.8) 40mM, NaAc 20mM, EDTA 1mM, SDS v)；

(2) 5M NaCl solution.

DNA is extracted:

Add 600 μ L extracting solutions (Tris-Ac (pH 7.8) 40mM, NaAc 20mM, EDTA 1mM, SDS 1%) to 1.5ml Centrifuge tube (EP pipe) is put into EP pipe with a small amount of mycelia of stick scraping and grinds, 65 DEG C of water-bath 30min, centrifugal rotational speed 12000r/min, from Heart 10min；

Centrifugation gained 400 μ L of supernatant is taken, 100 μ L of 5M NaCL, ice bath 10min are added；

Then it is 12000r/min that centrifugal rotational speed is kept at being 4 DEG C in temperature, is centrifuged 10min, takes supernatant；In addition clear liquid 0.6 times of isopropanol and supernatant mix (or 2 times of dehydrated alcohols) ice bath 1h, keep centrifugal rotational speed 12000r/min, centrifugation 10min, the remaining substance of gained dries after discarding supernatant liquid, adds 20 μ L distilled water (ddH₂O), 1-2 μ L is taken to do DNA profiling.

PCR amplification ITS sequence:

(1) PCR instrument: ABI 3730-XL DNA sequencer (Applied Biosystems, USA)；

(2) amplimer: ITS1 (5 '-T C C G T A G G T G A A C C T G C G G-3 ') such as SEQ ID Shown in NO.2 and ITS4 (5 '-T C C T C C G C T T A T T G A T A T G C-3 ') is such as SEQ ID NO.3 institute Show；

(3) amplification system:

ITS sequence PCR amplification system is as shown in table 1: table 1.

Reactant	Sample-adding amount
		2X TagMasterMix	25μL
Primer-1	1μL
		Primer-2	1μL
Template DNA	2μL
		ddH2O	21μL
React total volume	50μL

DNA profiling is made to be above-mentioned in Template DNA in table 1.

PCR reaction condition is as shown in table 2:

Table 2.

Note: step 2 carries out 30 circulations in table 2.

The electrophoresis detection of pcr amplification product:

The Ago-Gel (the 5 μ l/100ml containing Goldview) that deposition condition is 1%, 1 × TBE electrophoretic buffer, 90V Electrophoresis 1 hour, PCR product applied sample amount was 3 μ L, point sample after mixing with 1 μ L Loading dye.

In the ultraviolet lower observation of 254nm as a result, using the DL1000DNA Marker of TaKaRa company as nucleic acid standard molecular weight Object of reference determines expanding fragment length.Amplified production band should be on the position of reference substance 400-700bp.

PCR product purifying and sequencing: it is carried out by Shenzhen Huada Genetic Technology Co., Ltd.

The building of systematic evolution tree:

The ITS sequence of the fungi measured in the ITS sequence surveyed and GenBank gene pool is compared, according to than Correlated series are downloaded to result.Network analysis is carried out simultaneously by the adjoining algorithm (Neighbor-joining NJ) of MEGA 6.0 Systematic evolution tree is constructed, as shown in Figure 3.

As a result:

(1) strain morphology feature

Colony morphology characteristic: in PDA culture medium, bacterium colony is round, has obvious wheel line, neat in edge, substrate mycelium white, gas The raw milky white carpet-like of mycelia, the aleurioconidium group of celadon is adhered on bacterium colony surface, as shown in a1 in Fig. 2.

Spore shape feature: it is round, 2.50 μm of diameter, as shown in b1 in Fig. 2.

(2) bacterial strain ITS sequence and its phylogenetic analysis

Using primer I TS1 and ITS4, the segment of a 400-500bp size is amplified from strain gene group DNA, is passed through It is sequenced and compares sequencing result by BLASTn in GenBank.The result shows that in bacterial strain SDTE-P and Penicilliu category Fungi has very high base sequence similitude, therefore downloads the reference strain sequence that Penicilliu belongs in GenBank, for being Unite developmental analysis, with Monascus belong in Monascus aurantiacus (AY629435) and Monascus purpureus (AY629416) it is used as outer group.In the systematic evolution tree of building, SDTE-P and Penicillium citrinum (KF624801) and Penicillium citrinum (KM491892) gets together to form holding strength as 100% end point Branch.Base similarity system design the result shows that, SDTE-P and Penicillium citrinum (KF624801, KM491892) base Sequence does not have difference, sequence similarity 100%.Comprehensive morphological and molecular biological characteristics, bacterial strain is initially identified as Penicillium citrinum。

Embodiment 2

Inhibiting effect of the metabolite of bacterial strain to Radix Notoginseng anthrax bacteria

One, the extraction of the fermented and cultured of bacterial strain and metabolite

(1) sophora tonkinensis Gapnep endogenetic fungus SDTE-P is inoculated in potato dextrose agar plane ware (PDA plate) In, being placed in temperature is to cultivate 10 days at 28 DEG C, and gained culture materials are cut into bulk and are transferred to sterile solid culture medium (containing 400g Potato, the dextrose and 20g sucrose of 20g) 2 liters of conical flask in, ferment 40 days under the conditions of being placed in 28 DEG C；

(2) after fermentation, the methanol soaking fermentation object and ultrasound 40min with 2 times of fermentation material, then uses filtered through gauze, Take filtrate；

(3) step (2) are repeated twice, merges filtrate twice and medicinal extract is concentrated under reduced pressure into, obtains endogenetic fungus SDTE-P The methanol crude extract of fermentation material, as sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite.

Two, inhibiting effect of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite to the mycelia growth of Radix Notoginseng anthrax bacteria

The inhibitory activity that Radix Notoginseng anthrax bacteria mycelia is grown with the metabolite of mycelia growth method measurement bacterial strain SDTE-P. The metabolite of bacterial strain SDTE-P and powder of carbendazim (positive control) are respectively prepared containing 2mg/mL, 4mg/mL, 8mg/mL is dense Spend the drug containing tablet of metabolite.Aseptically, 6mm bacteria cake is made in Radix Notoginseng anthrax bacteria with punch, Radix Notoginseng charcoal Subcutaneous ulcer germ bacteria cake is connected to each drug containing tablet center as processing, is feminine gender with the Radix Notoginseng anthrax bacteria bacteria cake that not drug containing tablet center connects Control, in triplicate, all plates are placed in 28 DEG C of cultures for all processing and control.Drug concentration is answered by the field of powder of carbendazim It is arranged with concentration.When negative control covers with culture dish, measurement compares bacterium colony and handles the growth diameter of bacterium colony respectively, and according to Following formula calculates inhibiting rate:

Negative control increment=negative control growth diameter-bacteria cake diameter

Handle increment=processing growth diameter-bacteria cake diameter

Three, minimal inhibitory concentration of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite to Radix Notoginseng anthrax bacteria

With the metabolite of broth dilution method determination bacterial strain SDTE-P to the minimal inhibitory concentration of Radix Notoginseng anthrax bacteria.It will be 5 days Radix Notoginseng anthrax bacteria pure culture biscuits involvng inoculations are cultivated in PDA culture medium in the PDB fluid nutrient medium for containing 0.2% Tween 80 (v/v) In, 28 DEG C, 150r/min shaking table culture 7 days, culture is then gone into triangular flask, and pour into containing the sterile of 0.2% Tween 80 Distilled water stirs 30min with magnetism stick, solution is filtered with sterile gauze, obtains spores solution, and with blood counting chamber by spore Sub- concentration is adjusted to every milliliter 10⁴A spore is spare.The metabolite of bacterial strain is dissolved into the metabolism of 80mg/mL with 1%DMSO Product concentration for the treatment of takes 5 sterile test tubes to be sequentially arranged on rack for test tube, and number is 1,2,3,4,5 respectively, and every pipe is separately added into 1ml contains the aseptic double-distilled water of 0.2% Tween 80, and it is 1 that number then, which is added, in the bacterial strain methanol crude extract solution of 1ml 80mg/mL Test tube in and twice of serial dilution is successively carried out in each pipe.The above-mentioned gained spores solution (10 of 1ml⁴A/ml) it is separately added into In each pipe, to obtain the concentration for the treatment of of final SDTE-P metabolite: 20mg/ml (1 test tube of number), 10mg/ml (number 2 test tubes), 5mg/ml (3 test tube of number), 2.5mg/ml (4 test tube of number), 1.25mg/ml (5 test tube of number).The 1% of 1ml The 8mg/mL Flusilazole of DMSO and 1ml replace metabolite to execute above-mentioned treatment process respectively and as negative control and sun Property control, it is all processing and control in triplicate.Finally, 28 DEG C of each pipe, 150r/min shaking table culture 5 days.The metabolism of SDTE-P The MIC value of product is the minimum crude extract concentration for completely inhibiting Radix Notoginseng anthrax bacteria visible growth.

As a result:

The percent inhibition that the metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P grows Radix Notoginseng anthrax bacteria mycelia It is as shown in table 3:

Table 3.

Note: positive control powder of carbendazim contains 50% carbendazim in table；* indicates that data pass through one-way analysis of variance in table LSD relatively after, the metabolite and positive control powder of carbendazim of bacterial strain SDTE-P is under same concentrations, in P_0.05In level Has significant difference.

The suppression result table that the metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P grows Radix Notoginseng anthrax bacteria mycelia Bright, the metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P all has extraordinary suppression to the mycelia growth of Radix Notoginseng anthrax bacteria Effect processed, the metabolite of SDTE-P grows Radix Notoginseng anthrax bacteria C.gloeosporioides mycelia as can be seen from Table 3 Percent inhibition is 100%.Compared with positive control powder of carbendazim, the metabolite of bacterial strain SDTE-P is to Radix Notoginseng anthracnose Bacterium C.gloeosporioides mycelia growth inhibitory effect with compare equally.

Minimal inhibitory concentration such as 4 institute of table that sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite grows Radix Notoginseng anthrax bacteria Show:

Table 4.

Processing	MIC(mg/ml)
			C.gloeosporioides
Flusilazole	0.25
		SDTE-P	2.5

Note: positive control Flusilazole active constituent content is 400mg/mL in table.

The metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P surveys the minimal inhibitory concentration that Radix Notoginseng anthrax bacteria is grown Test result shows that the metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P all has stronger inhibition to Radix Notoginseng anthrax bacteria and makees With the metabolite of sophora tonkinensis Gapnep endogenetic fungus SDTE-P is to Radix Notoginseng anthrax bacteria C.gloeosporioides as can be seen from Table 4 Minimal inhibitory concentration be 2.5mg/ml, be 10 times of positive control.

As may be known from Table 3 and Table 4, in the metabolite of bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P containing very strong restraining epiphyte or Antifungal ingredient, therefore, in the bionomic control of Radix Notoginseng anthrax bacteria, bacterial strain sophora tonkinensis Gapnep endogenetic fungus SDTE-P, which has, to be protruded Potentiality.

The aforementioned description to specific exemplary embodiment of the invention is in order to illustrate and illustration purpose.These descriptions It is not wishing to limit the invention to disclosed precise forms, and it will be apparent that according to the above instruction, can much be changed And variation.The purpose of selecting and describing the exemplary embodiment is that explaining specific principle of the invention and its actually answering With so that those skilled in the art can be realized and utilize a variety of different exemplary implementation schemes of the invention and Various chooses and changes.The scope of the present invention is intended to be limited by claims and its equivalents.

Claims (1)

1. a kind of application of metabolite of sophora tonkinensis Gapnep endogenetic fungus SDTE-P in prevention and treatment Radix Notoginseng anthracnose, it is characterised in that: The classification naming of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P is Penicillium citrinum (Penicillium citrinum) SDTE-P, preservation Number: CGMCC No.10808；

The preparation method of the metabolite of the sophora tonkinensis Gapnep endogenetic fungus SDTE-P, operating procedure are as follows:

(1) sophora tonkinensis Gapnep endogenetic fungus SDTE-P is inoculated in potato dextrose agar plane ware, being placed in temperature is It is cultivated 10 days at 28 DEG C, gained culture materials are cut into bulk and are transferred in sterile solid culture medium, are placed in 28 DEG C and ferment 40 days； The sterile solid culture medium potato containing 400g, the dextrose and 20g sucrose of 20g；

(2) after fermentation, the methanol soaking fermentation object and ultrasound 40min with 2 times of fermentation material, is then filtered；

(3) step (2) are repeated twice, merges filtrate twice and medicinal extract is concentrated under reduced pressure into, obtains endogenetic fungus SDTE-P fermentation The methanol crude extract of object, as sophora tonkinensis Gapnep endogenetic fungus SDTE-P metabolite.