KR20180116172A - Burkholderia ambifaria CJ4 strain Possessing antifungal activity against Root Rot Pathogen of ginseng and Use Thereof - Google Patents

Abstract

The present invention relates to Burkholderia ambifaria CJ4 having antibacterial functions against pathogen causing root rot in ginseng, and a use thereof. More specifically, the present invention relates to a microbial preparation for controlling pathogen causing root rot in ginseng, and a method for controlling pathogen causing root rot in ginseng using the microbial agent. The Burkholderia ambifaria CJ4 has excellent antibacterial functions against pathogen causing root rot in ginseng as compared with microorganisms having antibacterial functions against other pathogen causing root rot in ginseng, while showing high antibacterial activities against various pathogen causing root rot in ginseng, thereby enabling production of the Burkholderia ambifaria strain, a culture solution of the strain, a microbial preparation for controlling pathogen causing root rot in ginseng containing a mixture thereof as an active ingredient, and a method for controlling pathogen causing root rot in ginseng using the microbial preparation.

Description

Burkholderia ambifaria CJ4 strain Possessing antifungal activity against Root Rot Pathogen of ginseng and Use Thereof}

The present invention relates to Buckholderia ambifaria CJ4 having antibacterial activity against ginseng root rot and its use, and more particularly, to Buckholderia cancer having antimicrobial activity against various pathogens known to cause ginseng root rot. It relates to a microbial agent for controlling ginseng root rot pathogens containing the biparia CJ4 strain, the strain culture medium as an active ingredient, and a method for controlling ginseng root rot pathogens using the microbial agent.

Korean ginseng is an important natural resource unique to Korea. It is a representative herbal medicine in Korea that has been widely used in oriental medicine as well as for the health of mankind from thousands of years ago. As Western medicine shows its limitations in overcoming side effects and incurable diseases of synthetic drugs, the value of herbal and natural products including ginseng is emerging. Ginseng prevents cancer, inhibits aging, protects the liver, recovers from fatigue and strengthens brain function, and has recently been reported to prevent damage from environmental hormones. Ginseng, the first among all herbal medicines, is a plant that grows in an extremely limited environment. It has a longer growing period compared to other crops, and its cultivation method is also special and difficult, so it is only produced in a few countries around the world.

Korea is a ginseng breeding country that produces Korean ginseng with excellent medicinal properties due to its climate climate being suitable for its natural growth and cultivation. However, many countries are increasingly interested in the ideal conditions that ginseng has as a health food or health medicine, and not only the existing ginseng producing countries but also non-producing countries are actively promoting policies to encourage research and production of ginseng competitively. Countries that grow ginseng are China, Japan, the United States, and parts of Europe, including Korea. In the past, most of the countries that consume ginseng were Confucian cultures, such as Korea, China, Japan, Taiwan, and Southeast Asia, but ginseng consumers in many countries around the world. Is expanding. However, Korean ginseng's international competitiveness in terms of price is weakening, and in order to strengthen international competitiveness, research on cultivation technology development to improve the quality of fresh ginseng, the raw material ginseng, is required. In particular, the most urgent problem in cultivation of ginseng in recent years is the absolute shortage of proper planned land, a high incidence rate due to deterioration in mountainous soil conditions, and there are many cases in which exports are not possible due to residual pesticides due to the increase in pesticide use. Intensive research is urgently required. On the other hand, ginseng root rot symptom (ginseng root symptom), which is considered important in ginseng cultivation, has been observed not only in cropland but also in grassland, and root symptom is one of the causes of lowering the germination and survival rate as it is cultivated as aging root after seedling planting. It is thought to be one of the. In fact, it is common for seedlings planted in farms to show a survival rate of less than 50% at the time of harvest after 5 years. The reason is that root pathogens live in areas where the roots of ginseng rot during cultivation. In this way, when ginseng is cultivated by root pathogens after being damaged by root pathogens in the grassland, the thick film spores of the root pathogens remaining in the soil are uniformly distributed in the soil through 7-10 cycles and rotary work when managing the planned site. If it does, it is believed that it will cause further damage to ginseng. In fact, the cause of root rot is Cylindrocarpon destructance. destructans ), Fusarium solani ), Pythium sp., Phytophthora cactorum ) and Erwinia carotovora have been reported to be largely involved alone or in combination (ginseng cultivation, standard farming textbook p 103, Rural Development Administration, 2000). In addition, the cause of the above-ground disease of ginseng is Rhizoctonia sorani (Rhizoctonia), which is the cause of Mojalok disease, which occurs on the above-ground part of seedlings. solani) and fish ium (Pythium sp.), and the cause of jeommunuibyeong Alterna Ria Panax (Alternaria panax ), etc., so it is a problem. As a control method for these diseases, the planned site management through the use of organic substances to reduce the density of pathogens in the soil or to suppress the outbreak, and chemical control using soil fumigants are being attempted. Has become.

However, it is difficult to achieve a distinct effect with the above control method, and because soil infectious pathogens such as ginseng root rot form thick film spores in the soil and cause disease, it is almost impossible to control. The use of soil fumigants also poses a risk of predominance of pathogens in 5 years due to the destruction of the soil ecosystem. In particular, the major soil diseases of ginseng are infected in various temperature ranges, so once infected, the disease progresses slowly, but damages throughout the year. Therefore, for effective soil disease control, it is believed that biological control is most suitable as a control measure for ginseng disease, which has a consistent and stable environment for ginseng cultivation characteristics, and shows antagonism against Cylindrocarpon destructans in Korea. The identification and antagonism of Streptomyces species have been reported (2nd World Ginseng Symposium Journal, Hu-Seop Jung, Choong-Hee Kim, p. 67-74, 1978, Korea Ginseng Research Institute).

Until now, biological control methods using antagonistic microorganisms have been used as measures to control ginseng root rot, chemical control methods using soil fumigants such as Cyron and Basamide, and non-ginseng cultivation methods of Dapjeon Yun-hwan have been used. The method is not systematic. In particular, until now, biological control studies on ginseng soil disease have been mainly conducted on the selection of antagonistic microorganisms and testing of the effect of field treatment, but lack of understanding of the various rhizosphere soil environments, and the antibacterial activity of the selected bacteria against the target pathogens is not high or fungicides. No antagonistic microorganisms with excellent control effect against ginseng root rot have been found, such as poor ability to settle bacteria in soil or seedlings due to inadequate methods or treatment methods.

Accordingly, as a result of the present inventors making diligent efforts to screen microorganisms having high antimicrobial activity against pathogens that cause ginseng root rot, the present inventors selected a strain of Berkholderia ambiparia having high antibacterial activity against pathogens of ginseng root rot, and ginseng Cylindrocarpon destructans pathogen causing root rot, as well as Botrytis cinerea (Botrytis) Cinerea ), Fusarium solani , Rhizoctonia solani ), etc. It was confirmed that it has high antibacterial activity against various ginseng root rot pathogens, and the present invention was completed.

The present invention has been conceived to solve the above problems, and the first problem to be solved of the present invention is Burke having antibacterial activity against pathogens causing ginseng root rot, pathogens causing ginseng mojalock disease, and pathogens causing ginseng gray mold disease. It is to provide a strain of Holderia ambiparia.

The second object of the present invention is to provide a microbial preparation comprising the strain, a culture medium of the strain, or a mixture thereof, and a method for controlling ginseng root rot pathogens using the microbial preparation.

In order to solve the first problem of the present invention described above, the present invention includes a strain of Buckholderia ambiparia of accession number KACC 92097P. According to a preferred embodiment of the present invention, the strain may have antimicrobial activity against pathogens causing ginseng root rot, pathogens causing ginseng mojaloc disease, and pathogens causing ginseng gray mold disease.

According to another preferred embodiment of the present invention, the pathogens causing the ginseng root rot are Cylindrocarpon destructans , Botrytis cinerea. Cinerea ), Fusarium solani (Fusarium solani) and Rhizoctonia solani (Rhizoctonia solani) It may be one or more selected from the group consisting of.

According to another preferred embodiment of the present invention, the strain may include the 16S rRNA nucleotide sequence of SEQ ID NO: 1.

According to another preferred embodiment of the present invention, the strain may be one having a pH of 5 to 7 and a temperature of 20 to 40 ℃ optimal growth conditions.

In order to solve the second problem, the present invention is characterized by using a microbial preparation for controlling ginseng root rot and the microbial preparation comprising the Berkholderia ambiparia strain of the deposit number KACC92097P, a culture solution thereof, or a mixture thereof as an active ingredient. It includes a method of controlling ginseng root rot.

According to a preferred embodiment of the present invention, the culture medium is 15 ℃ to 35 ℃ in any one or more medium selected from the group consisting of LB medium, NB medium, BHI medium, KB (King'B) or TSB medium. It can be obtained by culturing in.

Burkholderia ambiparia CJ4, which has antibacterial activity against ginseng root rot pathogens of the present invention, has excellent antimicrobial activity against ginseng root rot pathogens, compared to microorganisms having antibacterial activity against other ginseng root rot pathogens, and has a variety of Since it has a high antimicrobial activity against ginseng root rot pathogens, a microbial formulation for controlling ginseng root rot pathogens, which includes the burgholderia ambiparia strain, the culture medium of the strain and a mixture thereof as an active ingredient, ginseng using the microorganism formulation It has the effect of providing a method for controlling root rot pathogens.

1 is a result showing the separation and culture characteristics of the CJ4 strain having antibacterial activity against the ginseng root rot pathogens selected in the present invention.
2 is a result of confirming the growth characteristics according to the pH condition of the CJ4 strain selected in the present invention.
3 is a result of confirming the growth characteristics according to the temperature conditions of the CJ4 strain selected in the present invention.
4 is a result of confirming the effect of inhibiting the growth of ginseng root rot pathogen mycelium of the CJ4 strain selected in the present invention.
5 is a result of confirming the effect of inhibiting the growth of ginseng root rot pathogen mycelium and the effect of inhibiting spore germination of the CJ4 strain selected in the present invention.
6 is a result of confirming the effect of inhibiting the growth of the ginseng mojaloc pathogen mycelium of the CJ4 strain selected in the present invention.

Hereinafter, the present invention will be described in more detail.

As described above, as a method for controlling ginseng root rot, a biological control method using antagonistic microorganisms, a chemical control method using soil fumigants such as Cyron and Bassamide, and a nonginseng cultivation method of Dapjeon Yun-Hwan are used as methods for controlling ginseng root rot. , Until now, the method has not been systematized, and the ability to settle bacteria in the soil or seedlings is poor due to lack of understanding of the rhizosphere soil environment, the antibacterial activity of the selected strain against the target pathogen, or lack of a fungicide method or treatment method. There is a problem that no antagonistic microorganisms having excellent control effect against ginseng root rot such as falling are found.

The present invention seeks to solve the above-described problem by providing a buckholderia ambiparia strain having antibacterial activity against ginseng root rot pathogens. Through this, a microbial preparation for controlling ginseng root rot and a method for controlling ginseng root rot containing as an active ingredient of the Berkholderia ambiparia strain that can effectively inhibit the growth of various ginseng root rot pathogens, a culture solution thereof, and a mixture thereof. It has the effect it provides.

Accordingly, the present invention includes the Burkholderia ambiparia strain of accession number KACC92097P.

In the present invention, the strain may have antimicrobial activity against any one or more selected from the group consisting of pathogens causing ginseng root rot, pathogens causing ginseng mojaloc disease, and pathogens causing ginseng gray mold disease. The pathogens that cause the ginseng root rot are Cylindrocarpon destructans , Botrytis cinerea. Cinerea ), Fusarium solani (Fusarium solani) and Rhizoctonia solani (Rhizoctonia solani) It may be one or more selected from the group consisting of. In addition, as a pathogen that causes ginseng mojaloc pathogen, Rhizoctonia sorani (Rhizoctonia solani ) or Phythium spp.

In an embodiment of the present invention, Cylindrocarpon destructance, a ginseng root rot pathogen known as a major causative agent of ginseng serial disturbance. destructans ) In order to select antagonists that can effectively control, 200 strains were isolated from 100 ginseng samples, and CJ4 strains having antimicrobial activity specifically against Cylindrocarpon destructans were selected. Were selected.

In order to confirm the antibacterial activity of the selected CJ4 strain, as a result of confirming the degree of mycelial growth inhibition for the published strains 12hon 1-6, 12 Poc2-7 and 12Yeol-3 of Cylindrocapone destructance, Table 2 and As shown in FIG. 4, it was confirmed that all the Cylindrocarpon destructance strains inhibited the growth, and Botrytis cinerea, another pathogen causing ginseng root rot. Cinerea ), Fusarium solani ) and Rhizoctonia solani were also confirmed to exhibit an inhibitory effect on mycelial growth.

In the present invention, the culture conditions of the selected strain were investigated in order to establish the optimal culture conditions in which the high antibacterial activity of the CJ4 strain selected in the present invention is maintained.

As a result, as shown in FIG. 2, it was found that the bacteria did not survive at pH 4 or lower or pH 9 or higher, and did not survive well at pH 8 or higher. However, it was confirmed that the bacteria were well cultured at pH 5 to 7.

In addition, as shown in FIG. 3, the bacteria did not grow well at a temperature of 15°C, but it was confirmed that the bacteria grew well at a temperature higher than that. Therefore, the optimum temperature for culturing CJ4 bacteria may be 20 to 40 °C, more preferably 25 to 40 °C.

In the case of culturing at a temperature of less than 20°C, the growth of the mycelium is very slow, which may cause a problem that requires cultivation for a long time.When cultivating at a temperature exceeding 40°C, the problem of rapid mycelial growth inhibition and the death of the strain during long-term cultivation may occur. Can occur.

In the present invention, as a result of analyzing the 16S rRNA sequence to identify the selected strain, the CJ4 strain has the 16S rRNA nucleotide sequence of SEQ ID NO: 1, and Burkholderia ambiparia ambifaria ) was confirmed to correspond to the species, and the strain was named Burkholderia ambiparia CJ4 (hereinafter referred to as'CJ4 strain'), and was deposited with the Agricultural Genetic Resource Center of the National Academy of Agricultural Sciences (accession number KACC92097P).

The present invention also provides a microbial preparation for controlling ginseng root rot comprising the Berkholderia ambiparia strain of deposit number KACC92097P, a culture solution of the strain or a mixture thereof as an active ingredient, and a method for controlling ginseng root rot using the microbial preparation. Includes.

In the present invention, the culture medium may be cultured in any one or more medium selected from the group consisting of LB medium, NB medium, BHI medium, KB (King'B) or TSB medium, but , Any medium capable of maintaining high antimicrobial activity against ginseng root rot pathogens of Buckholderia ambiparia CJ4 of the present invention can be used without limitation.

The culture medium is preferably in the range of 20°C to 40°C, preferably 25°C to 40°C, but is not necessarily limited to the Berkholderia ambiparia CJ4 strain of the present invention. Ginseng root rot can be used without restrictions provided it is a condition that can maintain high antibacterial activity against pathogens. The culture may be cultured at pH 5.0 to 7.0, more preferably at pH 5.7 to 6.8.

However, in the case of culturing at a temperature below 20°C, the growth of the mycelium is very slow, which may cause a problem requiring long-time cultivation, and when cultivating at a temperature exceeding 40°C, rapid inhibition of mycelial growth and death of the strain during long-term cultivation may occur. Problems can arise.

In addition, if cultivated at a pH of less than 5.0, the growth may be poor and the cultivation time may be prolonged. If cultivated at a pH exceeding 7.0, the growth will be rapidly slowed and the problem of long cultivation may occur. Can occur.

The control agent containing the Burkholderia ambiparia strain of the present invention is not particularly limited as long as it is a conventional control agent used for the prevention and treatment of ginseng root rot, but preferably microbial pesticides, seed coating agents, ginseng roots of ginseng It may be one or more of the group consisting of a microbial pesticide for controlling rot disease, an additive for the production of a non-sterile medium for cultivation of ginseng, biotop soil, microbial nutrients, soil conditioners, composting agents, foliar sprayers, or drench sprayers.

In particular, Cylindrocarpon destructans and Botrytis cinereas, which are pathogens of ginseng root rot, which are a problem when cultivating ginseng. Cinerea ), Fusarium solani ) and Rhizoctonia solani (Rhizoctonia solani) has an excellent effect on growth inhibition, microbial pesticides for controlling ginseng root rot of ginseng, additives for the production of non-sterile media for cultivation of ginseng, and can be used in the production of biotope .

The microbial pesticide is not particularly limited as long as it is prepared by a conventional method, including the Burkholderia ambiparia strain of the present invention.

For example, it may include a strain of Burkholderia ambiparia and a biomatrix that is a microbial carrier, and the biomatrix may include a plant resource polymer material, a microbial polymer material, a nutrient, an auxiliary material, and an additive. .

In addition, the plant resource polymer material may include potato powder, soybean powder, starch powder, and the like, and the microbial-based biopolymer may include mucopolysaccharide, poly-betahydroxy-alkanoate, and a coagulant. Further, the nutrient may include glucose, peptone and inorganic salts, and the auxiliary material may include vermiculite, rice husk burned powder, and the like, and the additive may include an extender, a UV blocking agent, and a surfactant.

The microbial pesticide may be used as an antifungal microbial pesticide, a seed coating agent, a microbial nutrient, a soil conditioner, a composting agent, a foliar spray agent or a drench spray agent.

Hereinafter, the present invention will be described in detail with reference to preferred embodiments so that those of ordinary skill in the art can easily implement the present invention. However, the present invention may be implemented in various different forms and is not limited to the embodiments described herein.

Isolation of antagonists against ginseng root rot pathogens

In the present invention, the main pathogen causing root rot of ginseng In order to isolate effective antagonists for the control of Cylindrocarpon destructans , about 100 samples of 5-6 year old ginseng were collected from ginseng cultivation and crop fields in 2015 to isolate 200 kinds of endogenous bacteria.

First, the ginseng roots were surface sterilized in 1% NaOCl for 10 minutes, washed twice with sterile distilled water, and the surface was sufficiently dried for 20 minutes on a sterile workbench. The ginseng roots were cut horizontally with a sterilized knife, and the ginseng pieces were placed on King’B medium and cultured for 1 week at 25°C in an incubator. The colonies that showed differences in morphology that grew on King’B medium were purely isolated. Then, at 30° C. temperature conditions, morphological characteristics were confirmed while culturing in LB medium for 2 days. The purely isolated strain was inoculated into LB medium, cultured at 30° C. and 200 rpm for 1 day, and then sterilized 20% glycerol was added to the strain culture solution to prepare a storage strain and then used in the experiment.

In the same way, 200 strains were isolated from 100 ginseng samples, and the antibacterial activity against Cylindrocarpon destructans strain was investigated to have specific antibacterial activity against ginseng root rot pathogens. CJ4 strain, a negative bacterium, was selected. The selected CJ4 strain had a characteristic of showing a dark yellow pigment in the colonies grown on the LB medium (FIG. 1).

Identification of Selected CJ4 Strains

A molecular biological method was used to identify the CJ4 strain selected in Example 1 of the present invention.

First, the CJ4 strain selected in Example 1 was inoculated into 20 ml of LB medium, cultured at 30° C. for 1 day, and then centrifuged at 13,000 rpm for 10 minutes to harvest the cells.

Genomic DNA was isolated from the harvested cells using a Qiagen Genomic DNA Extraction Kit (Qiagen, Germany). In order to amplify the 16S rRNA gene site, PCR was performed using universal primers 27F and 1492R (Macrogen, Korea) shown in Table 1 below.

Primer sequence Base sequence Sequence number 27F AGAGTTTGATCMTGGCTCAG SEQ ID NO: 2 1492R TACGGYTACCTTGTTACGACTT SEQ ID NO: 3

PCR products were purified using a QIAquick PRC purification kit (Qiagen, Germany), and sequence analysis was requested by Macrogen Co., Ltd. and analyzed by Blast Search at the National Center for Biotechnology Information (NCBI). search) to identify the strain.

As a result of analyzing the 16S rRNA gene for identification of the selected strain, it was confirmed that the CJ4 strain had the 16S rRNA nucleotide sequence of SEQ ID NO: 1.

As a result of homology analysis on the NCBI web, Burkholderia Ambiparia ambifaria ) IHB B 1065 (GenBank No. KF475832) was confirmed to show a high homology of 99%, and the CJ4 strain of the present invention was confirmed to correspond to the Burkholderia ambifaria species.

Accordingly, in the present invention, the CJ4 strain was named Burkholderia Ambiparia CJ4 (hereinafter referred to as'CJ4 strain'), and was deposited with the Agricultural Genetic Resource Center of the National Academy of Agricultural Sciences (accession number KACC92097P).

3-1. Confirmation of growth characteristics of CJ4 strain according to pH conditions

In order to confirm the growth characteristics of endogenous bacteria, the pH of the LB medium was adjusted from 4 to 9 using 1N KCl and 1N NaOH. CJ4 strain was cultured in LB Agar at 30° C. for 1 day, and then inoculated into 20 ml LB medium and pre-cultured. 20 µl of the culture solution was inoculated into 20 µl of LB medium adjusted for each pH, and cultured at 37°C for 1 day to confirm the growth of the bacteria.

In order to confirm the growth characteristics of the endogenous bacteria B. ambifaria according to the pH conditions, as a result of culturing at 35°C and 200 rpm in LB medium, the bacteria did not grow at all at pH 4 and 9, and the optimum pH condition was pH It was identified as 6. After 24 hours, it was confirmed that the highest growth was 1.52 at _{pH6 and OD 600 nm (FIG. 2).}

3-2. Confirmation of growth characteristics of CJ4 strain according to temperature conditions

In order to investigate the culture characteristics according to the temperature conditions, the culture was incubated in LB Agar at 30°C for 1 day, and then inoculated in 20 ml LB medium and pre-cultured. 20 µl of the culture solution was inoculated into 20 µl of the LB medium, and the growth of the bacteria was confirmed while incubating for each temperature condition.

As a result of investigating the growth of CJ4 bacteria according to the temperature conditions, the appropriate temperature for growth was confirmed to be 35°C, and 8 hours after inoculation, it was grown to 0.822 at _{OD 600 nm.} After 24 hours, it was grown to 1.87 to 10 log CFU/ml. At 25°C, the initial growth rate was low, but there was no difference between the temperature at 35°C and the growth after 24 hours, and it was confirmed that the growth was good even at 25°C (FIG. 3).

4-1. Measurement of antibacterial activity of CJ4 strain against ginseng root rot pathogen

In the present invention, in order to confirm the antibacterial activity of the CJ4 strain selected in Example 1, Cylindrocarpon destructance, the main pathogen of ginseng root rot. destructans ) strains were isolated and identified in the ginseng department of the National Institute of Horticultural Science and were identified as pathogenic strains 12POC 2-7, 12HON1-6 and 12YEO1-3, and Botrytis causing ginseng root rot. cinerea and Fusarium The solani strain was also isolated and identified in the Ginseng Division of the National Institute of Horticultural Science, and a strain with pathogenicity was used for the antibacterial activity assay.

First, the CJ4 strain selected in Example 1 was inoculated into 20 ml of LB medium, cultured for 1 day under conditions of 30°C and 180 rpm, and centrifuged at 13,000 rpm for 10 minutes to prepare a culture solution and a supernatant. 10 µl of the culture solution was dropped on the center of the PDA, and the surface was dried for 10 minutes. Each ginseng root rot pathogen was cultured in a PDA (potato dextrose agar) medium at 20°C for 10 days, and after making a piece of mycelium using an 8mm cork borer, it was placed on the PDA and subjected to replacement culture at 25°C. Then, after one week, the degree of mycelial growth was confirmed.

In addition, in order to investigate the inhibition of spore germination of ginseng root rot pathogens, C. destructans 12POC2-7 mycelium fragments were inoculated in CMC (carboxymethylcellulose) medium to form spores, and then spores were recovered with sterile gauze. After spreading 100 µl of the spore suspension at a concentration of 5 log CFU/ml on the top, 10 µl of the CJ4 strain culture solution was dropped, dried for 10 minutes, and incubated at 25°C for 1 week to investigate the inhibition of spore germination.

Ginseng root rot pathogen hyphae growth inhibition by CJ4 strain Strain Clear zone diameter(mm) C. destructans
12hon1-6 C. destructans
12 Poc2-7 C. destructans
12Yeo1-3 F. solani B.cinerea CJ4 7.98 8.97 10.82 6.0 12.09

As a result, as shown in Table 2, Figure 4 and Figure 5, the CJ4 strain is antibacterial against all three species (12honl-6, 12Poc2-7, 12Yeol-3) of the Cylindrocarpon destructans strain It showed activity (see Fig. 4), and in particular, it was confirmed that the 12Yeol-3 strain showed the highest antimicrobial activity by forming a growth inhibitory source of 10.82 mm. In addition, as a result of investigating the degree of inhibition of mycelial growth in Botrytis Cinerea and Fusarium solani , which are other pathogens that cause ginseng root rot, they formed growth inhibitors of 12 mm and 6 mm or more, respectively. It was confirmed that there is an effect of inhibiting mycelial growth by the CJ4 strain of the present invention (see Figs. 5a and 5b). In addition, it was confirmed that spore germination of ginseng root rot pathogens was also inhibited (Fig. 5c).

Ginseng Mojalok pathogen Measurement of antibacterial activity of CJ4 strain in Korea

In the present invention, in order to confirm the antibacterial activity of the CJ4 strain selected in Example 1, phythium spp. And Rhizoctonia solani strains were tested for antimicrobial activity.

First, the CJ4 strain selected in Example 1 was inoculated into 20 ml of LB medium, cultured for 1 day under conditions of 30°C and 180 rpm, and centrifuged at 13,000 rpm for 10 minutes to prepare a culture solution and a supernatant. 10 µl of the culture solution was dropped on the center of the PDA, and the surface was dried for 10 minutes. Each ginseng mojalok pathogen was cultured in a PDA (potato dextrose agar) medium at 20°C for 10 days, and after making a piece of mycelium using an 8mm cork borer, it was placed on the PDA and subjected to replacement culture at 25°C. Then, after one week, the degree of mycelial growth was confirmed.

The degree of inhibition of mycelial growth of ginseng mojaloc pathogen by CJ4 strain Strain Clear zone diameter(mm) R. solani Phythium spp. CJ4 9.76 4.11

As a result, as shown in Table 3 and Figure 6, Mojalok pathogen Rhizoctonia Both solani and phythium spp. showed mycelial growth inhibitory activity, and CJ4 strain was Rhizoctonia. solani strain was confirmed to have high antibacterial activity by forming a growth retardation support of 9.76mm (Table 3).

As described above, specific parts of the present invention have been described in detail, and for those of ordinary skill in the art, these specific techniques are only preferred embodiments, and the scope of the present invention is not limited thereby. It will be obvious. Therefore, it will be said that the practical scope of the present invention is defined by the appended claims and their equivalents.

Institute of Agricultural Biotechnology KACC92097P 20151019

<110> REPUBLIC OF KOREA <120> Burkholderia ambifaria CJ4 strain Possessing antifungal activity against Root Rot Pathogen of ginseng and Use Thereof <130> 1042467 <160> 3 <170> KopatentIn 2.0 <210> 1 <211> 1451 <212> DNA <213> Artificial Sequence <220> <223> Burkholderia ambifaria CJ4 strain 16S rRNA <400> 1 ccggcatgcc ttacacatgc aagtcgaacg gcagcacggg tgcttgcacc tggtggcgag 60 tggcgaacgg gtgagtaata catcggaaca tgtcctgtag tgggggatag cccggcgaaa 120 gccggattaa taccgcatac gatcttcgga tgaaagcggg ggaccttcgg gcctcgcgct 180 atagggttgg ccgatggctg attagctagt tggtggggta aaggcctacc aaggcgacga 240 tcagtagctg gtctgagagg acgaccagcc acactgggac tgagacacgg cccagactcc 300 tacgggaggc agcagtgggg aattttggac aatgggcgaa agcctgatcc agcaatgccg 360 cgtgtgtgaa gaaggccttc gggttgtaaa gcacttttgt ccggaaagaa atccttggtt 420 ctaatatagc cgggggatga cggtaccgga agaataagca ccggctaact acgtgccagc 480 agccgcggta atacgtaggg tgcgagcgtt aatcggaatt actgggcgta aagcgtgcgc 540 aggcggtttg ctaagaccga tgtgaaatcc ccgggctcaa cctgggaact gcattggtga 600 ctggcaggct agagtatggc agaggggggt agaattccac gtgtagcagt gaaatgcgta 660 gagatgtgga ggaataccga tggcgaaggc agccccctgg gccaatactg acgctcatgc 720 acgaaagcgt ggggagcaaa caggattaga taccctggta gtccacgccc taaacgatgt 780 caactagttg ttggggattc atttccttag taacgtagct aacgcgtgaa gttgaccgcc 840 tggggagtac ggtcgcaaga ttaaaactca aaggaattga cggggacccg cacaagcggt 900 ggatgatgtg gattaattcg atgcaacgcg aaaaacctta cctacccttg acatggtcgg 960 aatcctgctg agaggtggga gtgctcgaaa gagaaccggc gcacaggtgc tgcatggctg 1020 tcgtcagctc gtgtcgtgag atgttgggtt aagtcccgca acgagcgcaa cccttgtcct 1080 tagttgctac gcaagagcac tctaaggaga ctgccggtga caaaccggag gaaggtgggg 1140 atgacgtcaa gtcctcatgg cccttatggg tagggcttca cacgtcatac aatggtcgga 1200 acagagggtt gccaacccgc gagggggagc taatcccaga aaaccgatcg tagtccggat 1260 tgcactctgc aactcgagtg catgaagctg gaatcgctag taatcgcgga tcagcatgcc 1320 gcggtgaata cgttcccggg tcttgtacac accgcccgtc acaccatggg agtgggtttt 1380 accagaagtg gctagtctaa ccgcaaggag gacggtcacc acggtaggat tcatgactgg 1440 ggtgaagtct a 1451 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> 27F primer <400> 2 agagtttgat cmtggctcag 20 <210> 3 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> 1492R primer <400> 3 tacggytacc ttgttacgac tt 22

Claims (9)

Burkholderia amphibolite strain of Accession No. KACC92097P.
[Claim 2] The method according to claim 1, wherein the strain has an antimicrobial activity against any one or more selected from the group consisting of pathogens causing ginseng root rot disease, pathogens causing ginseng mosaic disease and pathogenic bacteria causing ginseng gray mold disease Burkholderia strains of Amphibia.
The method according to claim 2, wherein the pathogen causing ginseng root rot disease is selected from the group consisting of Cylindrocarpon destructans , Botrytis Wherein said strain is at least one selected from the group consisting of Cinerea , Fusarium solani and Rhizoctonia solani .
2. The strain of claim 1, wherein the strain comprises the 16S rRNA nucleotide sequence of SEQ ID NO: 1.
The strain of claim 1, wherein the strain has an optimum growth condition at a pH of 5 to 7 and a temperature of 20 to 40 占 폚.
A microorganism preparation for controlling ginseng root rot disease, comprising the strain of Burkholderia ambiopria of any one of claims 1 to 5, a culture thereof or a mixture thereof as an active ingredient.
7. The culture medium according to claim 6, wherein the culture medium comprises the strain selected from the group consisting of LB (Luria Bertani) medium, NB (nutrient broth) medium, BHI (brain heart infusion) medium, KB (King's medium B) medium or TSB Wherein the microorganism is cultured at 30 占 폚 to 40 占 폚 in at least one medium selected from the group consisting of?
A method for controlling ginseng root rot disease, which comprises using a microorganism preparation for controlling ginseng root rot disease, which comprises the Burkholderia ambiparia strain of any one of claims 1 to 5, a culture thereof or a mixture thereof as an active ingredient.
9. The culture medium according to claim 8, wherein the culture medium comprises the strain selected from the group consisting of LB (Luria Bertani) medium, NB (nutrient broth) medium, BHI (brain heart infusion) medium, KB (King's medium B) Wherein the cultivation is carried out at 20 占 폚 to 30 占 폚 in any one or more medium selected from the group consisting of ginseng root rot.

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