KR101677513B1 - Paenibacillus polymyxa strain effective against Root Rot Pathogen of ginseng and Use Thereof - Google Patents

Abstract

The present invention relates to Penny Bacillus polymyxester ES-17 having antimicrobial activity against ginseng root rot disease and its use, and the antibiotic activity of the antimicrobial activity against the ginseng root rot disease bacteria of the present invention can be used for other ginseng root rot disease bacteria The antimicrobial activity against ginseng roots rot rot disease bacteria is superior to that of microorganisms having antimicrobial activity against ginseng root rot disease bacteria and has a high antimicrobial activity against various ginseng root rot disease bacteria. There is provided an effect of providing a microorganism preparation for controlling germplasm of root rot caused by ginseng as an active ingredient, and a method for controlling germ root rot rot disease bacteria using the microorganism preparation.

Description

[0001] The present invention relates to a polynucleotide of the genus Phenybacillus having antimicrobial activity against ginseng root rot,

The present invention relates to a Pennibacillus polymicus strain having antimicrobial activity against ginseng root rot disease and a use thereof, and more particularly to a strain of Pennibacillus polichmicus having antimicrobial activity against various pathogens known to cause ginseng root rot disease, A microorganism preparation for controlling germplasm of root rot of ginseng containing the above culture medium as an active ingredient and a method for controlling ginseng root rot disease bacteria using the microorganism preparation

KOREA INSAM is an important natural resource of Korea. It is a representative herb medicine of Korea, which has been widely used for medical purposes as well as oriental medicine for mankind for many millennia. With the adverse effects of synthetic medicines and the overcoming of intractable diseases, western medicine is showing its limit, and the value of ginseng and other herbal medicines and natural products is newly emerging. It is also reported that ginseng prevents cancer, inhibits aging, protects the liver, strengthens fatigue and brain function, and recently prevents environmental hormone damage. Ginseng, the first of all herbal medicines, is a plant that grows in an extremely limited environment. It has a longer growth period than other crops, and the cultivation method is also special and difficult.

Korea is a ginseng suzerain producing climatic ginseng, which is excellent in its medicinal effect due to its natural climate and cultivation area. However, many countries are increasingly interested in the ideal condition of ginseng as a health food or health care drug. In addition, existing ginseng producers as well as non-producers are actively promoting policies that encourage research and production of ginseng. In Korea, China, Japan, the United States, and some parts of Europe, ginseng has been cultivated in Korea, China, Japan, Taiwan and Southeast Asia. Is expanding. However, the international competitiveness of Korean ginseng has weakened in terms of price. In order to enhance international competitiveness, it is necessary to study the development of cultivation technology to improve the quality of ginseng as a raw material. Especially, the most urgent problems in cultivation of ginseng in recent years are the absolute lack of proper place, high incidence due to worsening of soil conditions in the country, and pesticide residue due to increase of pesticide usage. There is an urgent need for intensive research on

On the other hand, ginseng root rot symptoms (ginseng root symptoms), which are important in ginseng cultivation, are observed not only in aged paddy field but also in superpowder. Root symptoms are caused by dropping of the emergence and residual rate . ≪ / RTI > In fact, it is common for the seedlings that have been consumed at the farmhouse to have a survival rate of about 50% or less at harvest time after 5 years. This is because the root pathogens live in the area where the ginseng root rotates during the primary cultivation. When ginseng was cultivated as a result of ginseng damage after root damage caused by root pathogens in the supercollade, the thick spores of residual root pathogens in the soil were uniformly distributed in the soil by 7-10 times tillage and rotary work It is thought that it will cause damage to ginseng. Actually root rot is caused by Cylindrocarpon destructans), Fusarium small Lani (Fusarium solani), Fish ium (Pythium sp.), pie Saratov Torah kaek toreom it (the disease caused by Phytophthora cactorum), Air Winiah Caro sat Bora (Erwinia carotovora) are reported, alone or in combination significantly involved (Ginseng, standard Farming Manual p 103, Rural Development Administration, 2000). In addition, the cause of the giant ginseng topoisomerase is Rhizoctonia solani and Pythium sp. and Alternaria panax ) and so on. In order to reduce the density of pathogens in the soil or to control the disease, chemical control using soil fumigant is being tried through the use of organic materials to control the disease. .

However, the above-mentioned control method is difficult to obtain a clear effect, and soil-infectious pathogens such as ginseng root rot disease cause a disease by forming a thick spore in the soil, so that it is almost impossible to control. The use of soil fumigants has the potential to dominate the pathogen in five years due to the destruction of the soil ecosystem. In particular, the major soil diseases of ginseng are infected at various temperature ranges, so once infected, the disease progresses slowly but it is damaged throughout the year. Therefore, for the effective control of soil disease, biological control of ginseng disease is considered to be the most appropriate method for the control of ginseng disease, which is a continuous and environmentally stable disease control effect due to the characteristics of ginseng cultivation. In this regard, Cylindrocarpon destructans) it has been reported about the Streptomyces species (sympathy and antagonism of Streptomyces species) representing the road drag (Proceedings of the Second World Ginseng Symposium, jeonghuseop, gimchunghui, p 67-74, 1978, the Ginseng Research Institute) .

So far, a method of biological control using antagonistic microorganisms has been used as a method to control ginseng root rot disease, and in particular, a chemical control method using a soil fumigant such as a sand theory, a rosemide, and a rice paddy cultivation method The method is not systematized. In particular, studies on biological control of ginseng soils have been conducted mainly on the selection of antagonistic microorganisms and on the effect of the treatment of pesticides. However, there is a lack of understanding of various rhizospheric soil environments, the antimicrobial activity of the selected strain against the target pathogen is not high, The antimicrobial activity of the ginseng root rot disease is not observed due to the inferiority of the bacterial fixing ability in the soil or the seedling field due to insufficient method or treatment method.

As a result of intensive efforts to select microorganisms having high antimicrobial activity against pathogenic microorganisms causing ginseng root rot disease, the present inventors selected the strains of Pennibacillus policlinicum having high antimicrobial activity against ginseng root rot fungi, Cylindrocarpon < / RTI > destructans ), as well as Botrytis cinerea Cinerea , Fusarium solani , Rhizoctonia < RTI ID = 0.0 > The present invention has been accomplished based on the findings that it has a high antimicrobial activity against various ginseng roots rot fungi such as solani .

Disclosure of Invention Technical Problem [8] Accordingly, the present invention has been made in an effort to solve the above-mentioned problems, and an object of the present invention is to provide a strain of Penny Bacillus polymyx with antimicrobial activity against ginseng roots rot fungi and a culture solution thereof.

A second problem to be solved by the present invention is to provide a microorganism preparation containing the culture of the strain or strain and a method of controlling the germplasm of the ginseng root rot by using the microorganism preparation.

In order to solve the first problem of the present invention described above, the present invention includes a strain of Pennibacillus polymyx of Accession No. KACC92000P.

According to a preferred embodiment of the present invention, the strain may have antimicrobial activity against pathogenic bacteria causing ginseng roots rot disease.

According to another preferred embodiment of the present invention, the pathogenic bacteria causing the ginseng root rot disease are Cylindrocarpon destructans , Botrytis < RTI ID = 0.0 > Cinerea), Fusarium small Lani (Fusarium solani , and Rhizoctonia solani . In the present invention,

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

According to another preferred embodiment of the present invention, the strain comprises at least one lytic enzyme selected from the group consisting of cellulase, pectate lyase and xylanase, Can be generated.

According to another embodiment of the present invention, the strain may have at least one active mechanism selected from the group consisting of auxin production activity, siderophore production activity and phosphate availability activity.

In order to solve the second problem, the present invention provides a microorganism preparation for controlling ginseng roots rot disease comprising the Pennibacillus polymyx strain of the deposit number KACC92000P, a culture thereof or a mixture thereof as an active ingredient, and the microorganism preparation Includes ginseng root rot disease control method.

According to a preferred embodiment of the present invention, the culture solution is prepared by culturing the strain in one or more medium selected from the group consisting of LB medium, NB medium, BHI medium, KB (King's B) or TSB medium at 20 ° C to 30 ° C And then cultured.

The antimicrobial activity against the root rot of Ginseng according to the present invention is superior to that of other microorganisms having antimicrobial activity against other ginseng roots rot fungi, It is possible to provide a microbial preparation for controlling ginseng rot root rot disease comprising a Pennibacillus polymyx strain, a culture solution of the strain, and a mixture thereof as an active ingredient, a microbial agent for controlling ginseng root rot disease using the microorganism preparation, There is an effect of providing a method.

1 is data showing the isolation and culture characteristics of ES-17 strain having antimicrobial activity against ginseng root rot disease bacteria selected in the present invention.
Fig. 2 shows data on inhibition of mycelial growth of ES-17 strain ginseng root rot disease selected in the present invention (1: Bacillus spp., 2: Bacillus spp, 3: Bacillus spp, 4: ES17)
3: LB medium, 4: NB medium, 5: KB medium, and 3: LB medium, respectively. FIG. 3 is data showing the degree of antibacterial activity of ES- TSB medium, 6: no treatment group).
FIG. 4 shows 16S rRNA nucleotide sequence information of ES-17 strains selected in the present invention.

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

As described above, a biological control method using an antagonistic microorganism as a method for controlling ginseng root rot disease, and a vigorous method such as a method of chemical control using a soil fumigant such as a sand theory, , The method has not been systematized to date, lack of understanding of rhizosphere soil environment, lack of antimicrobial activity of selected strains against target pathogens, and lack of bacteria preparation method or treatment method, There is a problem that an antagonistic microorganism having an excellent control effect against ginseng root rot disease has not been found.

The present invention has been made to solve the above-mentioned problems by providing a strain of Pennibacillus polymyx with antimicrobial activity against ginseng roots rot fungi. The present invention provides a microorganism preparation for controlling ginseng root rot disease and a method for controlling ginseng root rot disease, which comprises as an effective ingredient of a culture of Penny Bacillus polymyx strains, a culture solution thereof, and a mixture thereof, which can effectively inhibit the growth of various ginseng roots rot rot disease bacteria It is effective.

Accordingly, the present invention includes a strain of Penny Bacillus polymyx of Accession No. KACC92000P (National Institute of Agricultural Science and Technology, National Institute of Agricultural Science and Technology).

In the present invention, the strain may have antimicrobial activity against pathogenic bacteria causing ginseng root rot disease, and the pathogenic bacteria causing ginseng root rot disease may be Cylindrocarpon destructans), Botrytis cine Leah (Botrytis Cinerea), Fusarium small Lani (Fusarium solani , and Rhizoctonia solani. In the present invention,

In one embodiment of the present invention, a ginseng roots rot fungus, Cylindrocarpon destructans ) was isolated from about 200 strains of rhizosphere soil. As shown in FIG. 1, the strains were specifically isolated against Cylindrocarpon destructans ES-17 strains were selected.

In order to confirm the antibacterial activity of the selected strain ES-17, the degree of inhibition of mycelial growth of KACC44657, KACC44661 and KACC44662, which are the published strains of the Cylindrocone disuntant, was checked. As a result, as shown in Table 1 and FIG. 2 It was confirmed that the inhibition of the growth of all the Cylindrocapon disintensis strains was inhibited and the other pathogens causing ginseng root rot disease, Botrytis Cinerea , Fusarium solani and Rhizoctonia solani were also found to be effective in inhibiting mycelial growth.

In the present invention, the antimicrobial activity according to the culture medium of the selected strain was examined in order to establish the optimum culture condition maintaining the high antimicrobial activity of the ES-17 strain selected in the present invention. The culture medium was LB, NB, BHI, KB (King's B), and TSB medium were used.

As a result, as shown in Table 2 and FIG. 3, when the selected ES-17 strain was cultured in the TSB medium, the antibacterial activity against the Cylindrocapone disintensitization strain and the Bortritis cinerea strain was the highest , Followed by LB and KB medium, respectively. In addition, when the ES-17 strain was cultured in BHI medium, it was confirmed that the antimicrobial activity of the Fusarium saurani strain and the L. typhimurium strain was the highest.

In the present invention, the 16S rRNA sequence was analyzed to identify the selected strains. As a result, the ES-17 strain had the 16S rRNA sequence (SEQ ID NO: 1) (Fig. 4) and Paenibacillus polymyxa ). The strain was named Penny Bacillus polymyxester ES-17 (hereinafter referred to as ES-17 strain) and was deposited on October 29, 2014 (Accession No. KACC92000P).

In the present invention, the Staphylococcus aureus strain ES-17 is a lytic enzyme selected from the group consisting of cellulase, pectate lyase and xylanase, And the strain may have at least one activity mechanism selected from the group consisting of auxin production activity, siderophore production activity and phosphate availability activity.

In one embodiment of the present invention, the ability to produce a soluble enzyme was confirmed to confirm the characteristics of the ES-17 strain. As a result, as shown in Table 4, the ES-17 strain had no activity against polygalacturonase and protease, but the cellulase, pectate lyase, And xylanase activity, as compared with the control.

As shown in Table 5, the ES-17 strain of the present invention was found to have auxin production ability, siderophore production ability, and siderophore production ability, It was confirmed that it had the ability to produce and the phosphate.

The present invention also includes a microorganism preparation for controlling ginseng root rot disease comprising a Pennibacillus polymyx strain of deposit number KACC92000P, a culture of said strain or a mixture thereof as an active ingredient, and a method for controlling ginseng root rot disease using said microorganism preparation .

In the present invention, the culture solution can be cultured in at least one culture medium selected from the group consisting of LB medium, NB medium, BHI medium, KB (King's B) or TSB medium, Any medium capable of maintaining high antimicrobial activity against the ginseng root rot fungus of the present invention can be used without limitation.

It is preferable that the culture liquid is a strain of Penny Bacillus polymyxester ES-17 at 15 ° C to 35 ° C, preferably 20 ° C to 30 ° C, but not always limited thereto, Any condition can be used without limitation as long as the antimicrobial activity against ginseng roots rot fungus can be maintained high. The culture can be performed at pH 5.0 to 7.0, and more preferably at pH 5.7 to 6.8.

However, when cultivated at a temperature lower than 15 캜, mycelial growth is very slow, which may cause problems of long-term cultivation. When the culture is performed at a temperature exceeding 35 캜, rapid mycelial growth is inhibited, Problems can arise.

In addition, when cultured at a pH lower than pH 5.0, there is a problem that the growth is slow and the cultivation time becomes long, and when cultured at a pH exceeding 7.0, the growth is drastically slowed, Lt; / RTI >

The antiseptic agent comprising the Pennibacillus polymyx strains of the present invention is not particularly limited as long as it is a conventional anticancer agent used for prevention and treatment of ginseng root rot disease. Preferably, the anticancer agent, seed coating agent, ginseng root rot disease control A microbial pesticide for ginseng cultivation, an additive for producing ginseng-free microbicide, a bio-soil, a microbial nutrient, a soil improving agent, a composting homework, a leaf spreading agent, or a root spraying agent.

Particularly, when the ginseng root rot germ, which is a problem in culturing ginseng, Cylindrocarpon destructans , Botrytis < RTI ID = 0.0 > Cinerea), Fusarium small Lani (Fusarium solani and Rhizoctonia solani, and can be used for the production of microbial pesticides for controlling ginseng root rot disease, additives for the production of ginseng-free microbicides, and bio-soil .

The microbial pesticide is not particularly limited as long as it is produced by an ordinary method including the Pennibacillus polymyx strain of the present invention.

For example, the biomatrix may include a Penny Bacillus polymyx strain and a biomatrix as a microorganism carrier, and the biomatrix may include a plant resource polymer material, a microbial biomaterial, a nutrient, an auxiliary material, and an additive.

In addition, the plant resource polymer material may include potato powder, soy flour, starch powder and the like. The microbial biopolymer may include mucopolysaccharide, poly-beta hydroxy-alkanoate, and coagulant. Further, the nutrient may include glucose, peptone, and inorganic salts, and the auxiliary material may include vermiculite, rice hull powder, and the like, and the additive may include an extender, a UV blocker, and a surfactant.

The microbial pesticide may be used as an antifungal microbial pesticide, a seed coating agent, a microbial nutrient, a soil improving agent, a compostant homework, a leaf surface spraying agent, or a root spraying agent.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. The present invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

Ginseng root rot disease Won Kyun  Antagonistic separation of

In the present invention, the main pathogenic bacteria causing ginseng root rot disease In order to isolate antagonistic bacteria effective for the control of Cylindrocarpon destructans , 200 antioxidant samples were collected from ginseng cultivation area, planned sites and forests from March to September,

First, 5 g of the soil sample was diluted with 45 ml of sterilized physiological saline, shaken at 200 rpm for 30 minutes in a shaking incubator, diluted stepwise by decidual dilution method, and 200 μl each was dispensed on a KB's agar plate. After incubation for 3 days at 20 ℃ incubation, pure colonies were isolated from colonies which showed morphological differences. The strains were inoculated into LB medium and cultured at 30 ° C and 200 rpm for 1 day. Then, sterilized 20% glycerol was added to the culture medium to prepare a stock strain.

In this way, more than 1,000 kinds of strains were isolated from 200 soil samples. Cylindrocarpon ( Cylindrocarpon) The ES-17 strain, which is a positive bacterium having antimicrobial activity specifically for the ginseng roots rot fungi, was selected by examining the antimicrobial activity against the destructans strain (Fig. 1).

Identification of antimicrobial activity of selective strains

In the present invention, in order to confirm the antibacterial activity of the ES-17 strain selected in Example 1, the antimicrobial activity of Cylindrocarpon (ginseng roots rot) destructans) identified the type strain three kinds of strain (KACC44657, KACC44661, KACC44662) to inhibit mycelial growth of the ES-17 strain in the received pre-sale Agricultural Microbiology bank (KACC) degree.

In addition, ginseng is Botrytis cine Leah (Botrytis Cinerea) pathogens known to cause root rot, Fusarium small Lani (Fusarium solani and Rhizoctonia solani were also measured.

First, ES-17 strain selected in Example 1 was inoculated into 20 ml of LB medium, cultured at 30 DEG C and 180 rpm for 1 day, and centrifuged at 13,000 rpm for 10 minutes to prepare culture medium and supernatant. Each standard strain was cultured at 20 ° C for 10 days. Mycelial fragments were prepared using a 8 mm Cork borer and placed on the center of the PDA medium. 20 μl of the culture supernatant was inoculated on 8 mm filter paper And then incubated for 1 week at 20 ° C. ( Bacillus spp. Culture was firstly inoculated on 1st, 2nd, 3rd filter paper, and ES17 strain was inoculated on 4th filter paper.)

Inhibitory Effect of ES-17 Strain on Growth of Ginseng Root Rot Bacteria Strain Clear zone diameter (mm) KACC44657 KACC44661 KACC44662 B.cinerea R. solani F. solani ES-17 7.13 6.97 10.35 3.36 3.79 3.96

As a result, as shown in Table 1 and Fig. 2, the ES-17 strain was found to be a Cylindrocarpon destructans strain (KACC44657, KACC44661, KACC44662). Especially, KACC44662 strain showed the highest antimicrobial activity by forming 10.35 ㎜ growth support. In addition, Botrytis cinerea, another pathogen causing ginseng root rot disease, Cinerea), Fusarium small Lani (Fusarium solani ) and Rhizoctonia solani , and it was confirmed that these pathogens had an effect of inhibiting mycelial growth by the ES-17 strain of the present invention .

Depending on the type of media ES Identification of antimicrobial activity of strain -17

ES-17 strains were cultured in LB, NB, BHI, KB (King'S B) and TSB medium to investigate the antimicrobial activity of ginseng roots rot fungi according to the cultivation medium of ES-17 strain selected in the present invention The antimicrobial activity of ginseng root rot fungus was measured.

The ES-17 strain was inoculated on LB medium, cultured at 30 ° C for 1 day, and inoculated into 20 ml of each LB, NB, BHI, KB (King's B) and TSB medium to obtain 3 log CFU / For one day. Then, each pathogenic bacterium cultured at 20 ° C for 10 days was made into a piece of hypha using a 8 mm cork borer, and then placed in the center of the PDA medium. Escherichia coli (ES-17) was inoculated on 8 mm filter paper, placed on the edge of the PDA, incubated with the pathogen at 20 ° C for one week, and the degree of inhibition of mycelial growth was examined . Plates on which only filter paper which had not been treated as a control group were prepared separately, and the antibacterial activity of ES-17 strain was compared.

Antimicrobial activity according to the culture medium of ES-17 strain Broth Clear zone diameter (mm) B.cinerea R. solani F. solani C.destruntans BHI 3.0 ± 0.23 3.3 ± 0.17 3.8 ± 0.63 5.5 ± 0.49 NB 2.4 ± 0.23 2.1 ± 0.38 3.7 ± 0.15 4.8 ± 0.29 LB 3.1 ± 0.41 3.0 ± 0.12 3.5 ± 0.51 6.0 ± 0.34 KB 2.5 ± 0.23 2.7 ± 0.18 3.4 ± 0.15 5.9 ± 0.56 TSB 3.5 ± 0.17 3.0 ± 0.25 3.5 ± 0.29 6.6 ± 0.28

As a result, as shown in Table 2 and FIG. 3, when the selected ES-17 strain was cultured in the TSB medium, the antibacterial activity against the Cylindrocapone disintensitization strain and the Bortritis cinerea strain was the highest , Followed by LB and KB medium, respectively. In addition, when the ES-17 strain was cultured in BHI medium, it was confirmed that the antimicrobial activity of the Fusarium saurani strain and the L. typhimurium strain was the highest.

Selected ES Identification of -17 strain

The molecular biological method was used to identify ES-17 strains selected in Example 1 of the present invention.

First, ES-17 strains selected in Example 1 were inoculated into 20 ml of LB medium, cultured at 30 DEG C for 1 day, and 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). PCR was performed using the universal primers 27F and 1492R (Macrogen, Korea) shown in Table 3 below to amplify the 16S rRNA gene region.

Primer sequence The base sequence (5 '-> 3') SEQ ID NO: 27F AGAGTTTGATCMTGGCTCAG SEQ ID NO: 2 1492R TACGGYTACCTTGTTACGACTT SEQ ID NO: 3

PCR products were purified using QIAquick PRC purification kit (Qiagen, Germany). Sequencing analysis was performed by Macrogen Co., and analyzed by Blast Search (National Center for Biotechnology Information, NCBI) The isolates were identified through the.

The 16S rRNA gene was analyzed for the identification of the selected strains. As a result, it was confirmed that the ES-17 strain had the 16S rRNA base sequence (903 bp) of SEQ ID NO: 1 (FIG.

As a result of homology analysis on the NCBI web, it was found that Paenibacillus polymyxa CR1 (GeneBank No. CP006941.1), and the ES-17 strain of the present invention was confirmed to be a species of Paenibacillus polymyxa .

Thus, in the present invention, the ES-17 strain was designated as Penny Bacillus polymyxester ES-17 (hereinafter referred to as ES-17 strain) and deposited at the National Institute of Agricultural Science and Technology (National Institute of Agricultural Science and Technology) as accession number KACC92000P.

Penny Bacillus Polyamic acid ES -17 < / RTI > lytic enzyme ) Generation  Confirm

In order to confirm the characteristics of ES-17 strain in the present invention, the ability to produce soluble enzyme was confirmed.

To investigate the ability of the ES-17 strain to produce soluble enzyme, 0.8% agarose containing 0.2% sodium azide was used. The pectate lyase production ability was determined using 1% polygalacturonic acid, 1% yeast extract (YE), 0.38 μM calcium chloride (CaCl ₂ ) and 100 mM Tris-HCl (pH 8 (PGA), 1% yeast extract, 2.2 mM EDTA (pH 8.0), and 110 mM < RTI ID = 0.0 > The medium containing sodium acetate (pH 5.5) was cultured in a medium containing 1% CMC (carboxymethycellulose sodium salt) and 25 mM sodium phosphate (pH 7.0) as a cellulase producing ability, The xylanase was cultured in a medium containing 1% OSX (Oat Spelts Xylan) and 25 mM sodium phosphate (pH 7.0), the protease was supplemented with 1% skim milk and 0.1% A medium containing yeast extract was used.

Each well was punched out with a sterilized 5 mm cork borer and the agarose pieces were removed and 30 μl of 0.8% agarose containing 0.2% sodium azide was added Wells were made. 20 μl of the supernatant of the ES-17 strain cultured in LB medium for 1 day and centrifuged at 15,000 rpm for 10 minutes were inoculated into a selective medium and cultured for 1 day. Then, the pectate lyase and poly- The polygalacturonase was stained with 4N hydrogen chloride (HCl) and the cellulase and xylanase was congo red to confirm the degradative enzyme activity. (clear zone).

The lytic enzyme production ability of the strain Penny Bacillus polychroma ES-17 Sample Clear zone diameter (mm) Cellulase Pectate lyase Polygalacturonase Protease Xylanase Supernatant 1.91 ± 0.109 2.23 ± 0.156 - - 7.21 ± 0.165 Culture solution 2.15 + 0.123 3.01 + 0.269 - - 8.63 ± 0.299

As a result, as shown in Table 4, the ES-17 strain had no activity against polygalacturonase and protease, but the cellulase, pectate lyase, And xylanase activity, as compared with the control.

Penny Bacillus Polyamic acid ES -17 strain of Auxin production , Sideropore ( siderophore ) Productivity and Phosphorus Availability  Confirm

In order to confirm the characteristics of ES-17 strain in the present invention, auxin production ability, siderophore production ability and phosphate availability were confirmed.

First, the EC-17 strain was plated on R2A medium supplemented with 0.1% L-tryptophan and incubated at 25 ° C for 2 days to examine auxin production. Salkowski reagent was dispersed in the culture medium to observe the degree of appearance of pink around the colony.

Siderophore production was confirmed by CAS (Chrome Azurol sulfonate) assay. To the 50 ml of distilled water, 60 mg of CAS (Sigma-Aldrich, USA) was added and 73 mg of hexadecyltrimethyl ammonium bromide ) Was dissolved in distilled water (40 ml). To 10 ml of 10 mM hydrogen chloride (HCl) was added iron (III) chloride hexahydrate (2.7 ㎎) was added, mixed well and sterilized at high pressure to prepare a blue dye. Then, 750 ml of distilled water, 100 ml of 10X MM9 salt, 15 g of agar, 32.24 g of PIPES (piperazine-N, N'-bis (2-ethanesulfonic acid)), 30 ml of 10 g of casamino acid solution , Vitamin (s) and carbon source were adjusted to pH 6.8, sterilized by autoclaving, cooled to about 50 ° C, and mixed with blue dye to prepare a CAS plate culture medium. After 20 μl of the selected strain was dispensed and cultured at 25 ° C. for 2 days, the degree of orange color formation was confirmed around the culture medium.

The phosphate tolerance was determined by inoculating PVK (Pikovskaya's) medium containing 0.5% insoluble phosphate, tricalcium phosphate, and incubating at 25 ° C for 2 days. Respectively.

Auxin production, siderophore production ability and phosphate availability of Penny Bacillus polyamic acid ES-17 strain Strain IAA
production Siderophore production Phosphate
solubilizing ES-17 + + +

As a result, as shown in Table 5, it was confirmed that the ES-17 strain of the present invention had an auxin production ability, a siderophore production ability and a phosphorus addition ability.

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments, It will be obvious. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

National Institute of Agricultural Science KACC92000P 20141029

<110> REPUBLIC OF KOREA (MANAGEMENT: RURAL DEVELOPMENT ADMINISTRATION) <120> Paenibacillus polymyxa ES-17 strain effective against Root Rot          Pathogen of ginseng and Use Thereof <130> 1040925 <160> 3 <170> Kopatentin 2.0 <210> 1 <211> 999 <212> DNA <213> Paenibacillus polymyxa ES-17 <400> 1 acagggataa ctaccggaaa cggtagctaa tacccgatac atccttttcc tgcatgggag 60 aaggaggaaa gacggagcaa tctgtcactt gtggatgggc ctgcggcgca ttagctagtt 120 ggtggggtaa aggcctacca aggcgacgat gcgtagccga cctgagaggg tgatcggcca 180 cactgggact gagacacggc ccagactcct acgggaggca gcagtaggga atcttccgca 240 atgggcgaaa gcctgacgga gcaacgccgc gtgagtgatg aaggttttcg gatcgtaaag 300 ctctgttgcc agggaagaac gtcttgtaga gtaactgcta caagagtgac ggtacctgag 360 aagaaagccc cggctaacta cgtgccagca gccgcggtaa tacgtagggg gcaagcgttg 420 tccggaatta ttgggcgtaa agcgcgcgca ggcggctctt taagtctggt gtttaatccc 480 gggctcaac ttcgggtcgc actggaaact ggggagcttg agtgcagaag aggagagtgg 540 aattccacgt gtagcggtga aatgcgtaga tatgtggagg aacaccagtg gcgaaggcga 600 ctctctgggc tgtaactgac gctgaggcgc gaaagcgtgg ggagcaaaca ggattagata 660 ccctggtagt ccacgccgta aacgatgaat gctaggtgtt aggggtttcg atacccttgg 720 tgccgaagtt aacacattaa gcattccgcc tggggagtac ggtcgcaaga ctgaaactca 780 aaggaattga cggggacccg cacaagcagt ggagtatgtg gtttaattcg aagcaacgcg 840 aagaacctta ccaggtcttg acatccctct gaccggtcta gagatagacc tttccttcgg 900 gacagaggag acaggtggtg catggttgtc gtcagctcgt gtcgtgagat gttgggttaa 960 gtcccgcaac gagcgcaacc cttatgctta gttgccagc 999 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> 27F <400> 2 agagtttgat cmtggctcag 20 <210> 3 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> 1492R <400> 3 tacggytacc ttgttacgac tt 22

Claims (10)

And one or more members selected from the group consisting of Cylindrocarpon destructans , Botrytis Cinerea , Fusarium solani and Rhizoctonia solani . Which has antimicrobial activity against pathogenic bacteria causing ginseng root rot disease,
Comprising a 16S rRNA nucleotide sequence of SEQ ID NO: 1,
Accession No. KACC92000P (National Institute of Agricultural Science and Technology, National Institute of Agricultural Science and Technology) Paenibacillus polymyxa ES-17 strain.
delete delete delete delete delete A microbial preparation for controlling ginseng root rot disease comprising the strain Penny Bacillus polyamic acid ES-17 of claim 1, a culture thereof or a mixture thereof as an active ingredient.
[Claim 7] The method according to claim 7, wherein the culture solution is obtained by culturing the ES-17 strain at 20 DEG C to 30 DEG C in at least one medium selected from the group consisting of NB medium, BHI medium, KB (King's B) Wherein the microbial agent is used for controlling root rot disease of ginseng.
A method for controlling ginseng root rot disease, which comprises using a microorganism preparation for controlling ginseng root rot disease comprising the strain of Penny Bacillus polymyxester ES-17 of claim 1, a culture thereof or a mixture thereof as an active ingredient.
[Claim 11] The method according to claim 9, wherein the culture solution is obtained by culturing the ES-17 strain at 20 DEG C to 30 DEG C in at least one medium selected from the group consisting of NB medium, BHI medium, KB (King's B) Wherein the roots of the ginseng root rot disease are treated by the method.

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