KR20150141293A - A novel strain of alcaligenes faecalis and a use of the same - Google Patents

Abstract

The present invention relates to a composition for promoting plant growth and a composition for preventing bacterial pustule disease in soybean, containing one or more kinds selected among the group consisting of: a Alcaligenes faecalis JBCS 1294 strain having an ability to induce salt damage resistance and growth promotion to a plant, which is accessed as accession no. KACC 91943P; a cultured product of the strain; a concentrate of the cultured product; and a dried cultured product.

Description

FIELD OF THE INVENTION [0001] The present invention relates to a novel alkaline zinispicillis strain,

The present invention relates to a novel strain which promotes the growth of crops, induces salting resistance in a salt solution from a plant, and has an effect of controlling crops, specifically, an effective control effect against soybean flour disease, and the use of the strain.

In modern times, it is common to use pesticides and fertilizers when cultivating crops as an important means of preventing plant pests and increasing yields. It is true that chemical pesticides and chemical fertilizers made from organic synthetic materials that have been rapidly developed since the early 1950s have greatly contributed to the increase in global food production, but these abuse of chemical pesticides and chemical fertilizers It is also true that serious damage such as the destruction of the ecosystem is continuing.

Specifically, when ingesting crops cultivated using artificially synthesized pesticides and fertilizers, artificially synthesized chemicals contained in pesticides and fertilizers may accumulate in the body, which may have harmful effects on the human body, It is reported that it may have a bad influence on the soil.

Due to these problems, various regulations on the use of chemical fertilizers and chemical pesticides, which use artificially synthesized chemicals as main materials, have been strengthened day by day in advanced countries, So-called organic farming, which does not use fertilizers or chemical pesticides, or cultivates crops while limiting their use, is on the increase.

However, there is a problem that organic farming requires a lot of effort by human beings and does not yield high yields. Despite these problems, as the demand for organic farming laws increases, government policies to support such organic farming and eco-friendly farming are being prepared.

For example, in recent years, various research and development on green technology, which is a technology to reduce chemical fertilizer and pesticide, has been proceeding in accordance with environment friendly agricultural promotion policy, which is a part of environment friendly policy. Interest in microorganisms is increasing. Specifically, as a substitute for chemical synthetic pesticides, the development of biological pesticides using microorganisms themselves or natural products derived from microorganisms has been actively carried out, and a number of microbial agents have been registered. As the demand for microbial pesticides increases, efforts for R & D and commercialization are increasing .

With the changes in government policy and environment and efforts to commercialize microbial pesticides, the microbial fertilizer market, which is a substitute for chemical fertilizers, is also expanding. In particular, the microbial fertilizer market is expected to expand gradually, considering the increase in salinity and dry areas due to environmental changes related to global warming and the increase of reclaimed land due to technological development. For example, while the growth rate for products based on chemical-based plant products is only 3%, the growth rate for products based on bio-based plant products is 16%, indicating that the market for bio-based crop protection products is growing more rapidly than the market for chemical based crop protection products . This trend of the market change can be confirmed by the fact that the global chemical companies or the global agricultural companies are actively participating in the development of biological products.

Specifically, a global agricultural company, Monsanto, USA, includes a microbial screening process to promote the growth of Agradis R & D centers and large plant-related microbial groups and plants to actively participate in the development of agricultural biological products. (Syngenta, USA) acquired Pasteuria Bioscience, a company that developed a biosecurity to prevent nematodes, and BASF (Germany), a global chemical company, Becker Underwood, a world-class biocontrol producer, has joined the company for $ 1 billion and is actively involved in the development of microbial products.

These microbial agents can be used as microbial fertilizers or microbial pesticides to replace conventional chemical fertilizers or chemical pesticides.

Plant growth promoting rhizobacteria (PGPR) have been used as microorganisms used in these microorganism preparations. The rhizobia promoting plant growth is a rhizosphere bacterium inhabiting the rhizosphere around the root of the plant which has a beneficial effect on the growth of the plant. The utilization of rhizosphere bacterium which promotes the growth of the plant is firstly applied to root vegetables such as radish, potato, beet, But recently it has been expanded to various plants such as soybean, barley, oilseed rape, cotton, peanut, pea, rice etc. Vegetables and apples. Compared with conventional farming methods using chemical pesticides, it is preferred as a method for sustainable agriculture.

The plant growth promoting rhizobial bacterium is widely used at home and abroad. As an example, in overseas, growth promoting bacteria such as Achromobacter piechaudii produced ACC deaminase (1-aminocyclopropane-1-carboxylic acid deaminase), which promoted resistance to drying of pepper and tomatoes and increased the growth of tomatoes in high salt concentrations by 66%. In addition, Paenibacillus polymyxa has been reported to improve resistance to drought in Arabidopsis thaliana , and Bacillus Subtilis has been reported to secrete volatile organic compounds (VOCs) to enhance tolerance to chlorophyll in Arabidopsis thaliana. In addition, the promotion was a typical soil rhizosphere bacteria, Pseudomonas fluorescens is the peanut seedling grown in the experimental results of high salt concentration region using the genus Pseudomonas, Pseudomonas fluorescens and Pseudomonas Putida significantly enhanced the growth of oilseed rape, Pseudomonas It has been reported that putida and Pseudomonascorrugata promote the growth of barley, oat and tall fescue, and increase cell membrane safety and photosynthesis

In addition, 1,330 strains of rhizobia were isolated from rhizosphere in the coastal dune area of Gyeongbuk province in Korea, and the antimicrobial activity against the major plant pathogens and the promotion of plant growth were examined. As a result, Pseudomonas It was reported that the fluorescence IB4-14 strain secreted ACC deaminase, which promoted root growth as well as seed germination performance of P. shrimp.

The plant growth promoting rhizobacteria produce plant growth materials such as auxin, solubilize insoluble phosphorus, directly affect plants through nitrogen fixation, inhibit plant pathogenic microorganisms, interact indirectly with other rhizospheric microorganisms And it is known to have a great influence on plants.

Therefore, in order to prevent the harmful effects of organic synthetic chemical pesticides such as ecosystem destruction, development of biological pesticides, development of biological pesticides using microorganisms including plant growth promoting ability as well as control ability, for example, It is true.

Plants are subject to growth inhibition by various environmental factors, leading to a reduction in yield. Biological stress and abiotic stress are classified into environmental factors that inhibit growth. Of these, abiotic stress is known to cause the most serious effects on plants, such as salinity.

In particular, long-term droughts due to unusual weather conditions and localized heavy rains are changing the cultivation environment to hot and humid climates, and problems such as global warming caused by long-term drought due to global climate change and lack of precipitation, And damage to plants (crops) due to increased salt concentrations in coastal farmland and reclaimed land is serious.

The salt damage is also referred to as a sea storm, and is a disaster mainly caused by intrusion of seawater containing typhoon or monsoon, or extraction of salinity from a reclaimed land. The seawater intrusion occurs when the flow rate of stream is decreased due to drought, Storm surge, tsunami or tsunami, or when the levee collapses due to high waves.

According to previous reports, about 20% of freshwater farmland worldwide is salty, and UN data indicate that about 50% of arable land is likely to be salted. In the case of Korea, there are many reclaimed land such as Sihwa and Seosan. Recently, Saemangeum Reclaimed Land has a total area of 28,300 3, which is 3,650 식. The area of 5,950 생 ecological environment includes 2,000 총 The largest marshland park and 1,000-square-meter wildlife park will be created. It is anticipated that such reclamation sites that are constructed or planned to be built in the country will not be free from salinization.

It is known that the salinity reduces the area of cultivated land and inhibits the growth of surrounding plants or restricts diversity. Specifically, sodium (Na ⁺ ) ion affects enzyme activation and protein synthesis, and chloride (Cl ^- ) ion is known to inhibit photosynthesis if it is overly concentrated at the root.

In order to improve the salinity, methods such as removal of the surface of the soil on which the salt is concentrated, continuous exchange of the soil and fresh water have been carried out. However, this method requires a lot of labor and is costly, There are disadvantages.

Given these circumstances, it is necessary to prepare for salt production not only for the cultivation of food crops, but also for the enhancement of biomass for the production of bioenergy and the improvement of urban landscape.

Recently, rhizobia promoting plant growth promotes the growth of plants through the production of plant growth promoting substances such as IAA or the use of phosphoric acid, inhibits the damage to pathogens through the secretion of antibiotics and siderophore, It has been reported to induce resistance to herbivorous animals and the like. In addition, it is expected that the rhizobia promoting plant and plant growth according to the growth of plants can provide soil, organic matter, nutrients, and oxygen through the metabolic process, thereby improving the soil and stabilizing the soil. However, studies on these are concentrated mainly on biological control of pests, and researches and related products for the development of abiotic stress such as salting and drying are limited.

Recently, the demand for environmentally friendly control is increasing due to the emergence of resistant bacteria to chemical pesticides, the destruction of ecosystem by chemical pesticides, and the risk of residual toxicity.

Biological pesticides are products produced by isolating collected pathogens, host resistant microorganisms, natural pesticides and natural enemies in the natural environment in order to control pests, hospital microorganisms and weeds of crops. The biological pesticides are divided into three groups: microbial pesticides, biochemical pesticides including pheromones, and plant pesticides. The microbial pesticide is an effective ingredient and includes microorganisms such as bacteria, fungi, viruses and algae. Among the above pesticides, there is an increasing demand and concern for biological control and microbial fertilizer using useful microorganisms, and development research of microbial pesticides and microbial fertilizers using beneficial microorganisms both domestically and abroad has been progressing actively.

In addition, during the growing period of soybean, various kinds of diseases occur. As soybean infectious diseases, there are many diseases such as anthrax, mummified, mushy, viral, The hospital germ, Xanthomonas axonopodis pv. Glycines are the most common bacterial pathogens in Korea, and the types and effects of these drugs are low. Therefore, there is a need for research for the solution of these problems. .

KR 10-0747701 B KR 10-0800566 B KR 10-1190650 B

According to the above needs, the object of the present invention is to provide a microorganism which is excellent in resistance to salting, that is, resistant to salt, has a plant growth promoting ability, and is capable of controlling flame wasting of soybean.

It is another object of the present invention to provide a method for promoting the growth of plants using the microorganisms according to the above-mentioned necessity.

It is another object of the present invention to provide a composition for controlling plant diseases using the microorganism according to the above-mentioned necessity.

In order to accomplish the above object, the present invention relates to an alkaline zinispicillis JBCS 1294 ( Alcaligenes faecalis JBCS1294, KACC91943P). The strain may have the ability to induce resistance to salt tolerance and growth in plants. In addition, the strain may have a controlling activity against fire blight of soybean.

In order to achieve the above object, the present invention provides an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, A concentrate of the cultured product, and a dried product of the cultured product as an active ingredient.

In order to achieve the above object, the present invention provides an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, And a microbial fertilizer containing at least one selected from the group consisting of a concentrate of the cultured product and a dried product of the cultured product as an active ingredient.

In order to achieve the above object, the present invention provides an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, A concentrate of the cultured product, and a dried product of the cultured product as an active ingredient. Xanthomonas infection is caused by X. a. pv. glycines , or a composition for preventing blight of soybean flour.

The present invention also relates to an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce tolerance to plants and growth promoting ability to plants, Treating the soil, plant or plant seed with at least one selected from the group consisting of water, a concentrate of the culture, and a dried product of the culture. The plant growth promoting method is an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, And a dried product of the culture may be used in a liquid state in accordance with plants, and the seeds of the crops may be immersed or sprayed or coated on seeds.

In order to achieve the above object, the present invention provides an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, Treating the soil, the plant or the plant seed with at least one selected from the group consisting of a concentrate of the cultured product and a dried product of the cultured product. The stress of the plant may be salty.

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

The inventors of the present invention conducted researches on plant control using microorganisms using rhizospheric microorganisms. Among them, JBCS 1294, which is a strain having excellent plant growth promoting ability, . As a result, the strain was identified as a new strain, Alcaligenes faecalis (JBCS 1294) JBCS 1294), and the causative agent of resistance to salt tolerance and plant growth promoted by the strain was identified and the present invention was completed.

In the present invention, a culture medium means a microorganism, a nutrient substance necessary for culturing a microorganism or an animal or animal's body, as a main component, or further includes an additional substance in addition to the main component for a specific purpose. It is also referred to as a culture medium, an incubator or a culture medium. The medium may be a natural medium, a synthetic medium, or a selective medium, and may be, but not limited to, a solid medium or a liquid medium.

In the present invention, a plant means a creature having a cell wall, a chlorophyll, a photosynthetic effect by independent nutrition, and a characteristic of not moving in movement, in order to be distinguished from animals and fungi constituting the earth's biological system. Examples include a southern plant, a reddish plant, a yellowish plant, a yellow bivalve plant, a yellow algae plant, a grazing plant, a noxious plant, a green plant, a tea plant, a fern or a seed plant. The plant is a concept that includes any part, for example, a tissue or organ, derived from living plants as well as living plants. For example, the part derived from the plant may be a fruit, a flower, a pin, a root, a stem, a leaf or a seed.

In the present invention, the term "crop" refers to a plant which was originally grown in the wild state, but cultivated by humans for use in some kind of application. The term "crop" means a plant grown for human use, ) And the like. Crops are classified into crops and horticultural crops, the former being called crops, and the latter being called garden crops. The crops may again be classified as edible crops, feed crops, and crops. The edible crops mainly consist of cereals, potatoes or legumes. The feed crops consist of crops harvested in an immature state, such as corn or wheat, and used as livestock feeds, before harvesting, and grasses used as grazing bases to form permanent grasslands. The above-mentioned artificial crops are made of fiber crops such as cotton, pay crops such as rape seeds, sugarcane crops such as sugar cane, tea, and tobacco.

In addition, in the present invention, the composition for promoting plant growth is used for promoting the growth of a plant, and is also referred to as a plant growth promoter. The plant growth promoting composition is used for promoting plant growth and promoting flowering and elimination Is used.

In addition, in the present invention, promotion of plant growth refers to promotion of germination of plant seeds, promotion of plant length growth, promotion of volumetric growth of plants, promotion of planting of fruit, increase of fruit yield, Enhancing plant leaf activity, promoting the growth of plant roots, promoting the coloring of fruit, or improving fruit quality, including increasing the sugar content of fruit.

The present invention relates to an alkaline zinispicillis JBCS 1294 deposited with Accession No. KACC 91943P ( Alcaligenes faecalis JBCS 1294).

In one example of the present invention, the process of searching for the strain of Alcaligenes faecalis JBCS 1294 ( Alcaligenes faecalis JBCS 1294) of the present invention is carried out by a method of selecting a strain that improves the resistance of a plant to salt damage from rhizosphere organisms and promotes growth Respectively.

Through the above-described search process, the JBCS 1294 strain, which is selected as a new strain selected to be excellent in the resistance to salt tolerance of plant from rhizosphere organisms and to promote growth, as well as the effect of controlling soybean flour blight, was identified. As a result, ( Alcaligenes faecalis ). Specifically, as a result of the 16S rDNA sequence analysis of the selected strains shown in Fig. 1, the selected JBCS 1294 strain of the present invention was identified as belonging to Alcaligenes faecalis , JBCS 1294 ( Alcaligenes faecalis JBCS 1294).

The alkaline zinispicillis JBCS 1294 ( Alcaligenes faecalis JBCS 1294) was deposited on May 9, 2014 with the accession number KACC 91943P to the National Institute of Agricultural Science and Technology, National Institute of Agricultural Science and Technology, Rural Development Administration, 126, Suen-ro, Suwon-gu, Gyeonggi-do.

The alkaline zinispicillis JBCS 1294 ( Alcaligenes faecalis JBCS 1294, KACC 91943P) is a newly cultivated rhizome resistant microorganism newly isolated and identified by the inventor of the present invention. The Alcaligenes faecalis JBCS 1294 strain improves the resistance of plants to salt damage, Not only does it promote growth, it also has an excellent anti-blight effect against soybean.

Specifically, it has been confirmed that the salt tolerance is improved in barley, wheat, and Arabidopsis thaliana. The salt-resistant material is a volatile substance, which is associated with hexanedioic acid or butanoic acid and induces an increase in expression of AtHKT1 and NHX1 genes And it was confirmed that it is a microorganism having excellent control effect and growth promoting effect against fire blight of soybean.

The present invention also relates to a strain of Alcaligenes faecalis JBCS1294 deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, And a dried product of the cultured product as an active ingredient.

The culture of the strain means a culture medium obtained by culturing the strain JBCS 1294, and may be a culture medium containing the strain or a cell-free cell-free culture medium. The culture of the above strain is a culture medium in which an alkali jispypical JBCS 1294 ( Alcaligenes faecalis JBCS 1294) may be cultured and then cultured, but the properties of the culture are not limited to liquid or solid.

The culture medium may be a conventional medium for culturing a microorganism or an animal or animal tissue, and may be one containing a carbon source, for example, potato starch, dextrose, or a mixture thereof. .

The concentrate of the culture means that the culture of the strain is concentrated by a conventional method. The concentrate of the culture may be a concentrate obtained by concentrating a liquid culture medium obtained by culturing the above strain of Alcaligenes faecalis JBCS 1294 ( Alcaligenes faecalis JBCS 1294) by a conventional method, but the culture is not limited to the liquid .

The plant may be a plant that is subject to the above-mentioned plant growth promoting composition may be specifically ssangjayeop plants (dicotyledonous plants) or monocotyledonous plants (monocotyledonous plants), an example beans (Phaseolus Vugalis L.), barley ( Hordeum There is vulgare . hexastichon), wheat, lettuce ( Lactuca sativa L.), radish ( Raphanus sativus Var), potatoes ( Solanum tuberosum L.), garlic ( Allium sativum L.), mandarin orange ( Citrus unshiu Marcvich), tomatoes ( Lycopersicon esculentum Mill.), Chinese cabbage ( Brassica campestris L. ssp. pekinensis ) and grass, and the grass may be at least one species selected from the group consisting of Rough bluegrass ( Poa trivialis ).

The composition for promoting plant growth may further comprise a microorganism or a microorganism culture liquid having an effect of promoting plant growth and controlling plant diseases, within the range of maintaining the effect of the present invention.

An example is the plant growth promoting microorganisms and plant disease control effect will be, ah Cine Saturday in bakteo (Acinetobacter sp.) Microorganisms, in Agrobacterium (Agrobacterium sp.) microorganisms, ahsseuro bakteo in (Arthrobacter sp.) microorganisms, azo RY rilreom in (Azospirillum sp.) microorganisms, Bacillus (Bacillus sp.) microorganisms, Bridal Grundig trillion in Away (Bradyrhizobium sp.) microorganisms, Franc Kea in Microorganisms such as Frankia sp., Pseudomonas sp., Rhizobium sp., Serratia sp., Paenibacillus sp. Microorganism and Thiobacillus sp. Thiobacillus sp.) Microorganisms.

The composition for promoting plant growth may further contain, as long as the effect of the present invention is retained, a surfactant used as a fertilizer component, a metal or a nonmetal, an oxide thereof, an absorbent or an electrodeposition agent, Can be added.

The fertilizer component, the metal or the nonmetal and the oxide thereof may be at least one selected from the group consisting of lithium (Li), beryllium (Be), boron (B), sodium (Na), magnesium (Mg), aluminum (Al) P, K, Ca, Sr, Mo, V, Cr, Mn, Fe, Co, Ni, And may be at least one selected from the group consisting of Ni, Cu, Zn, Ga, Ge, Se, Zr and oxides thereof. The fertilizer component, the metal or the nonmetal, and the oxide thereof may be the element, the oxide of the element or the salt of the element, and the salt may be, for example, a carbonate, a hydrochloride, a nitrate or a sulfate. The content of the fertilizer component, the metal or the nonmetal and the oxide thereof may be 0.001% by weight to 15% by weight or 0.1% by weight to 10% by weight based on the total weight of the composition.

The plant growth promoting composition may be prepared into a solid or liquid state by a production method known to those skilled in the art, and in this regard, the plant growth promoting composition The form may be solid or liquid, and may further comprise excipients for plant growth promoters for production in the form of the formulation.

The plant growth promoting composition may be applied by a method known to those skilled in the art and may be carried out by spraying, soil application, surface application, immersion or smoke application, or the like. . The application of the soil may be carried out by spraying, misting, atomizing, powder spraying or granular application. The application of the soil may be carried out by a mixing method or a circulation method. The surface application may be carried out by a coating method, Coating method. In addition, the application amount, that is, the usage amount can be appropriately adjusted according to the conditions such as the formulation, the plant to be applied, the application method, or the application site.

The present invention also relates to a strain of Alcaligenes faecalis JBCS1294 deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, And a dried product of a cultured product as an active ingredient.

The microorganism fertilizer refers to a microorganism preparation having a function of suppressing pests and promoting the growth of crops.

An Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce salt tolerance and growth promoting ability to the plant, a culture of the strain, a concentrate of the culture, and a dried product of the culture One or more selected from the group consisting of the above-mentioned microorganisms may be mixed with a carrier which is usually used in the production of microbial fertilizer if necessary, and the microbial fertilizer composition may be formulated into powders, pellets, granules or solutions or the like.

The carrier includes, but is not limited to, water, celite, zeolite, pearlite, vermiculite, peat moss, talc, kaolinite, bentonite, pig dust, fish dust, chick tone dust, rice dust, .

The present invention also relates to a strain of Alcaligenes faecalis JBCS1294 deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, And a dried product of a culture, as an active ingredient.

The above-mentioned soybean flour is caused by Xanthomonas axonopodis pv. It can be a congenital blight, which is glycines .

The present invention also relates to a strain of Alcaligenes faecalis JBCS1294 deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, And a step of treating the soil, the plant or the plant seed with at least one member selected from the group consisting of the dried product of the cultured product. The strain of the present invention is resistant to high salt stress and can be used to promote the growth of crops.

The method comprises spraying at least one member selected from the group consisting of the strain, the culture of the strain, the concentrate of the culture and the dried product of the culture to a soil around the crop in a solid state, , Seeds of crops, or by coating seeds, but are not limited thereto. In the case of immersion, the strain, its culture or the microorganism preparation may be poured into the soil around the plant or the seed may be immersed in the culture medium and preparation. In the case of spraying, it can be sprayed on the plant in a line by a technique well known in the art.

The plant may be a vegetable crop, for example, but not limited to, barley, wheat, soybeans, Arabidopsis thaliana, tomato, Chinese cabbage, lettuce, cucumber, pepper,

The present invention also relates to a strain of Alcaligenes faecalis JBCS1294 deposited with the deposit number KACC91943P having the ability to induce resistance to salt tolerance and growth in plants, a culture of the strain, a concentrate of the culture, A plant, or a plant seed to a plant, or a plant, a plant, or a plant.

The method for promoting resistance to stress of a plant may include spraying at least one selected from the group consisting of cultures of the strain, the concentrates of the cultures and the cultures of the cultures to a soil around the crop in a solid state, But are not limited to, spraying, spraying, dipping or spraying seeds of crops, or coating seeds. In the case of immersion, the strain, its culture or the microorganism preparation may be poured into the soil around the plant or the seed may be immersed in the culture medium and preparation. In the case of spraying, it can be sprayed on the plant in a line by a technique well known in the art. The stress of the plant may preferably be salty.

As described above, since the alkaline zinispicillis strain according to the present invention can improve the plant resistance against abiotic stress, especially the saltiness, which is not developed at home and abroad, the strain of alkaline zinispicillis JBCS1294 can be used as a seed or root of plant , It is possible to alleviate salinization, promote the growth of plants, and protect the damage caused by flour blight, which is seriously damaging to soybean, in an environmentally friendly manner. Therefore, the strain of alkali zincpicillus of the present invention And fruit farms, especially soybean cultivators and farmers in reclaimed land, and increase the productivity of related industries.

1 is a nucleotide sequence of 16S rDNA of JBCS 1294 strain according to an embodiment of the present invention.
FIG. 2 is a diagram showing a phylogenetic relationship with another bacterium based on the nucleotide sequence of 16S rDNA of JBCS1294 strain according to an embodiment of the present invention. FIG.
FIG. 3 shows the results of confirming the increase in resistance to barley and wheat for resistance to the treatment of JBCS1294 culture medium according to an embodiment of the present invention, wherein (1) represents the vitality index, and (2) represents the weight before drying mg), Control means control group without any treatment, 1294 means experimental group treated with culture medium of JBCS1294, Fig. 3a is a graph obtained through growth of barley, Fig. 3b shows growth of wheat This is the graph I checked.
FIG. 4 is a graph showing the results of confirming the increase in salt tolerance and growth activity according to the salt concentration of the JBCS 1294 culture medium according to an embodiment of the present invention, wherein □ denotes a control group without any treatment, 1294 strain, and the horizontal axis of each graph represents the treated salt concentration. FIG. 4A shows the result of measuring the root length, 4B shows the result of measuring the length of the stem, and 4C 4d is the result of measuring the vitality index.
FIG. 5 is a graph showing the activity of the JBCS 1294 strain in the presence of JBCS 1294 and the control group Enterobacter The abscissa is the incubation time and the ordinate is the absorbance (OD ₆₀₀ ) under the condition of 600 nm, 1294 is the JBCS 1294 strain, UW4 is the Enterobacter means UW4 cloacae strains, and, ACC means a medium containing 3.0 mM ACC as a nitrogen source, and (NH _{4) 2} SO _4; means a medium containing _{3.0 mM (NH 4) 2 SO} 4 , respectively.
FIG. 6 is a graph showing the relationship between the degree of growth and the degree of growth of the JBCS 1294 strain by culturing the JBCS1294 strain in a culture medium having different salt concentrations, in order to confirm whether or not IAA (indoleacetic acid) is produced in the JBCS 1294 strain according to an embodiment of the present invention. The abscissa is the salt concentration of the culture medium and the ordinate axis of the left side is the degree of growth of the strain measuring the absorbance (OD ₆₀₀ ) under the condition of 600 nm, and the vertical axis of the right side is the absorbance at the condition of 535 nm (OD ₅₃₅ ).
FIG. 7 is a photograph for confirming the growth activity of Arabidopsis thaliana (Col-0) stressed by 100 mM NaCl by a volatile substance produced by the strain JBCS1294 according to an embodiment of the present invention. FIG. 7B is a photograph showing the result of culturing the JBCS1294 strain on the opposite side of the Arabidopsis thaliana, followed by incubation for 14 days This is a photograph showing one result.
FIG. 8 is a graph showing the growth of Arabidopsis thaliana (Col-0) stressed by 100 mM NaCl by a volatile substance produced by JBCS1294 strain and Arabidopsis mutated at its specific signal transduction site according to an embodiment of the present invention Graphs showing the results of measuring the activity, specifically the length of the stem (shoot length, cm), the root length (cm), the number of the lateral root number and the fresh weight (mg) , □ denotes the control group with distilled water, and ■ denotes the experimental group treated with the culture medium of JBCS 1294 strain. The horizontal axis of the graph denotes the type of Arabidopsis thaliana, and the vertical axis of each graph indicates the growth activity of each Results are shown.
FIG. 9 is a graph showing the growth activity of the Arabidopsis thaliana (Col-0) stressed by 100 mM NaCl by the volatile substances produced by the JBCS1294 strain according to the plant hormone inhibitor according to an embodiment of the present invention, (Root length, cm), Lateral root number (number), and fresh weight (mg), and □ indicates that the distilled water is dropped Means a control group, and & cir & indicates an experimental group treated with the culture medium of JBCS 1294 strain. The abscissa of the graph indicates the type of Arabidopsis thaliana, and the vertical axis indicates the results of measuring the respective growth activities.
10 is a graph showing the results of SPME-GC / MS analysis of volatile organic compounds produced by JBCS 1294 strain according to an embodiment of the present invention.
FIG. 11 is a graph showing changes in the content of proline by the volatile substances produced by the JBCS 1294 strain according to an embodiment of the present invention. FIG. (Proline content / Arabidopsis seedling biomass weight) of the Arabidopsis thaliana mutant at the site of signal transduction, and FIG. 11B shows the results of the analysis of the stress of the one-salt stress by the volatile substance produced by the JBCS 1294 strain according to the plant hormone inhibitor (Proline content / Arabidopsis seedling biomass weight) obtained by measuring the content of proline in the Arabidopsis thaliana. In the graph, " □ " means a control group in which distilled water was dropped, " means an experimental group treated with JBCS 1294 culture medium, , The abscissa of the graph represents the species of the Arabidopsis or the plant hormone inhibitor, and the vertical axis of the graph represents the content of proline And it shows the
FIG. 12 is a graph showing the amount of gene expression before and after the treatment of the volatile organic material produced by the strain JBCS 1294 according to an embodiment of the present invention. In the graph, the abscissa is the target gene for the expression level, □, □, □, □, □, and □, respectively.
FIGS. 13 to 16 show the control rates of the soybean flour blight according to the throughput of the JBCS 1294 strain according to an embodiment of the present invention. FIG. 13 is a photograph showing the control of the soybean flour blight according to the throughput of the JBCS 1294 strain FIG. 14 is a graph showing the treatment of soybean flour blight according to the throughput of the strain JBCS1294, wherein the numbers on the horizontal axis represent the throughput of the strain, and the numbers on the vertical axis represent the number of leaves FIG. 15 and FIG. 16 are graphs showing the degree of control of the control of soybean flour blight in the greenhouse grown in the greenhouse when the JBCS1294 strain was treated, The horizontal axis in FIG. 15 indicates whether or not the strain is treated, the vertical axis indicates the incidence rate, and the letters under the photograph in FIG. 16 indicate whether or not the strain is treated.
17 is a graph showing whether or not the growth of soybean is promoted according to the throughput of JBCS 1294 strain according to an embodiment of the present invention. In Fig. 17, □ denotes root portion, □ denotes stem portion, The vertical axis on the left indicates the vitality index of each site, and the vertical axis on the right side indicates the biomass of each site.

Hereinafter, embodiments of the present invention will be described in detail so that those skilled in the art can easily carry out the present invention. The present invention may, however, be embodied in many different forms and should not be construed as being limited to the embodiments set forth herein.

Example  1. Induction and growth of salt tolerance Accelerating ability  Identification of microorganisms with rhizosphere

The strains were harvested from the cultivation area of soybean in Jeonbuk Province in 2013, and the strains were isolated. The strains identified as promoting the growth of Arabidopsis thaliana inducing salting were selected as JBCS1294 strain Respectively.

As a result of confirming the gram-positive reaction to the JBCS1294 strain that best promoted the growth of the Arabidopsis thaliana inducing the salting out, Gram-negative reaction was confirmed and Gram-negative bacteria were mainly detected. In addition, The results of the analysis are shown in Table 1 below. The '+' in Table 1 means that there is a contemporary ability, and '-' means that there is no contemporary competence.

Nutritional Sources result Nutritional Sources result Arabitol - Glucopyranoside - Lactose - Esculin + Sucrose - ONPG (beta-galactosidase) - Mannitol - Tryptophan - Dulcitol - Phenylalanine - Adonitol - Potassiumnitrate - Sorbitol - Urea - Cellobiose - Sodium thiosulfate - Raffinose - Lysine + Rhamnose - Arginine + Inositol - Ornithine + Maltose - Glucose -

As shown in Table 1 above, the JBCS 1294 strain was confirmed to use Esculin, Lysine, Arginine, Ornithine as a nutrient source. As a result of the biochemical analysis, it was expected to be an Alcaligenes sp. Strain.

More specifically, in order to identify the JBCS 1294 strain, the 16S rDNA nucleotide sequence of the JBCS 1294 strain was identified and compared with the nucleotide sequences registered in GenBank using the 16S rDNA nucleotide sequence and the BLAST program, and Clustal X And the MEGA5 program were used to analyze the homology of the nucleotide sequences, the phylogenetic relationships are shown in FIG.

As shown in FIG. 2, the JBCS1294 strain was found to have the highest degree of flexibility with Alcaligenes faecalis . As a result of the analysis, the strains were found to be Alcaligenes faecalis ), and according to the result of the analysis, the new bacterial strain was treated with Alcaligenes faecalis JBCS 1294 JBCS 1294). The above-mentioned alkali jinis picolis JBCS 1294 was deposited on May 9, 2014 with the accession number KACC 91943P at the National Institute of Agricultural Science and Technology, National Institute of Agricultural Science, Rural Development Administration, 126, Suen-ro, Suwon-

Example  2. Separated Microbial resistance to salt Induction ability  And plant growth Acceleration ability  Confirm

Example  2-1. JBCS  1294 barley and wheat Salt resistance  Induction confirmation

In order to confirm the induction of salting resistance in barley (brand name new barley) and wheat (field name planted wheat) of JBCS 1294, an experiment using a paper towel was conducted. Specifically, the surface-sterilized barley seeds (control group) and the surface-sterilized barley seeds were inoculated into the LB liquid medium for 12 hours and then centrifuged at 4,000 rpm for 10 minutes to obtain cells , Washed three times with sterilized water, and seeded in a suspension of the above-mentioned culture medium suspension at a concentration of 1 × 10 ⁸ CFU / ml for 1 hour and then sown.

Barley seeds (experimental group) treated with the bacteria were inoculated on a paper towel treated with 150 mM NaCl, put in a zipper bag, and cultured in a thermostat at 25 ° C for 14 days. After the culture, the growth of barley was investigated by measuring the vigor index and fresh weigh. The vitality index means a value obtained by multiplying the germination percentage (%) by the sum of the average root length and the average stem length. After repeating the experiment three times, the average value of the total test results is shown in FIG. 3A. In addition, an experiment was performed on the ground mill using the same experimental method as that of the barley experiment except that paper towel treated with 200 mM NaCl was used, and the average value of the total experimental results was shown in FIG. 3B.

As shown in FIG. 3A, the viability index of the barley barley was 132.9% higher than that of the untreated control group, and the fresh weight was found to be similar to that of the untreated control group compared with the control group treated with the strain of JBCS1294 . In addition, as shown in FIG. 3B, in the case of the landscaping wheat, the vitality index was increased to 824.3% of the untreated and 269.6% of the fresh weight compared to the untreated. In addition, in the case of landscape wheat, the root length and root length of the control group were increased to 212.4% and 863.7%, respectively.

According to the above-mentioned treatments, it was confirmed that, in spite of the induction of salt induction by the salt treatment using the NaCl solution, when the strain of JBCS 1294 was treated, the salt damage was minimized and the growth of the plant was rather promoted. It was confirmed that when the strain of JBCS 1294 was treated, the effect of preventing wheat and preventing growth of wheat was more excellent.

Example  2-2. JBCS  By 1294 salt concentration Salt resistance  Induction confirmation

In order to confirm the degree of salt tolerance induction of JBCS 1294 by salt concentration, the following experiment was conducted. First, as in Example 2-1, the JBCS 1294 was cultivated on the surface sterilized barley seeds (control group) and surface disinfected barley seeds, and then the strain of JBCS 1294 was cultivated at a concentration of 1 × 10 ⁸ CFU / ml The barley seeds (experimental group) treated with the adjusted culture medium for 1 hour were inoculated into a pot without salt treatment and a pot treated with 100 mM or 200 mM NaCl, respectively, and cultured at 25 ° C for 28 days. Then, the germination rate, stem and root length, fresh weight and vitality index of barley seeds were confirmed. After repeating the experiment three times, the average value of the total test results is shown in FIG.

As shown in FIG. 4, in the pot of the general soil without salt treatment, the experimental group was found to increase by 1.9% in the live body and by 13.1% in the vitality index, as compared with the control group. When the strain was treated, it was confirmed that the growth promoting effect was obtained. As shown in FIGS. 4A to 4C, when seeding in a pot treated with 100 mM NaCl, the germination rate, stem length, root length and fresh weight were 2.5%, 5.58%, and 11.6% % And 10.42%, respectively, and it was confirmed that the pots treated with 200 mM NaCl increased by 33.3%, 24.6%, 36.5% and 23.4%, respectively. In addition, as shown in FIG. 4D, in the experimental group treated with the strain of JBCS 1294, the potency index was increased by 16.5% and 73.4%, respectively, in the pot treated with 100 mM and 200 mM.

Example  2-3. JBCS  1294 Salt resistance  Check mechanism

The following experiment was conducted to confirm the salt resistance mechanism of JBCS1294.

First, in DF culture medium (DF medium), including to identify ACCdeaminase (1-aminocyclopropane-1- carboxylic aciddeaminase) generated whether the JBCS1294 strain, a nitrogen source in the 3.0 mM ACC and _{3.0 mM (NH 4) 2 SO} 4 , respectively JBCS 1294 strain was cultured. As a control, Enterobacter producing ACC deaminase cloacae UW4 strain was cultured in the same medium. Each of the bacteria was inoculated in the medium, and cultured at 25 DEG C and 180 rpm. After culturing for 24 hours and 48 hours, the absorbance (OD ₆₀₀ ) were measured. The results of the measurement of the absorbance are shown in Fig.

As it is shown in Figure 5, Enterobacter cloacae UW4 strain when Thereafter strain inoculated, 48 hours, the absorbance in the medium supplemented with ACC (OD ₆₀₀₎ is but reached grew to 0.9 degree, JBCS1294 strain, unlike medium supplemented with (NH _{4) 2} SO ₄ It was confirmed that the JBCS1294 strain could not use ACC as a nitrogen source and that the JBCS1294 strain could not produce ACC deaminase.

In addition, it was confirmed whether the JBCS1294 strain produced IAA, a representative auxin hormone related to plant growth. First, the JBCS1294 strain was cultured in LB medium at a concentration of 1%, 3%, 5%, 7%, 9% and 12% (w / v) (OD ₆₀₀ ) was measured at 600 nm and the degree of IAA production was determined at 535 ° C. The culture was incubated at 25 ° C and 180 rpm for 24 hours. The absorbance (OD ₅₃₅ ) was measured under the condition of a wavelength of 400 nm. The results of the measurement of the absorbance are shown in FIG.

As shown in FIG. 5, the JBCS 1294 strain was found to be most active at a salt concentration of 3%, and it was found to be able to grow to a salt concentration of 5%. However, the strain JBCS 1294 produced almost no IAA at all concentrations . Therefore, it was confirmed that the growth promoting ability and salt tolerance of the strain JBCS1294 were not related to the production of auxin-like hormone.

Example  2-4. JBCS1294 of Salt resistance  Identification of mechanism (1)

In order to more clearly confirm the salt tolerance mechanism of JBCS1294, the following experiment was conducted using Arabidopsis thaliana.

First, Arabidopsis Columbia-0 (Col-0) ecotype and gai1 (gibberellin insensitive, Koorneef et al ., 1985), etr1 (ethylene-insensitive, Chang et al ., 1993), cbb1 (brassinosteroid deficient mutant dwf1-6, Kauschmann et al al ., 1996), Eir1 (auxin transport deficient and ethylene-insensitive, Luschnig et < RTI ID = al ., 1998) and cre1 (ethylene-insensitive, Inoue et al ., 2001) using mutant Arabidopsis thaliana. First, each Arabidopsis seeds were treated with 70% (v / v) ethanol solution for 2 minutes, treated with 1% (v / v) sodium hypochlorite for 15 minutes, sterilized, and washed 5 times with sterile distilled water. Murashige and Skoog (MS) medium supplemented with 0.8% (w / v) agar, 1.5% sucrose and 100 mM NaCl was added to one side of the above divided I-plates (Fisher Scientific, Pittsburgh, USA) After the seeds were dispensed in duplicate, the surface-sterilized seeds were inoculated on each of 8 plates in a half of the MS medium. The JBCS 1294 strain was cultured at 30 ° C in a Luria-Bertani (LB) medium one day before the experiment. The strain was suspended in distilled water to give a concentration of 1 × 10 ⁷ CFU / mL, Were placed on LB agar discs (4 mm) placed on the other half of I-plates seeded with Arabidopsis and allowed to be exposed to the volatiles produced by the JBCS 1294 strain during the incubation period. As a control group, distilled water was added instead of the culture medium.

The plate was then capped and sealed with parafilm and placed in a plant orientation (16 h light / 8 h dark, 100 m m ^-2 s ^-1 and 23 賊 1 째 C) at an angle of 70 ° Lt; / RTI >

After 14 days, fresh weight, stem and root length, and number of secondary roots of each Arabidopsis were examined. The same experiment as described above was repeated three times each, and the average value thereof was measured. The experimental results are shown in Figs. 7 and 8 and Table 2 below. In the following Table 2, '-' means no increase in live body weight, '+' means less than 30% increase from live body weight before culture, '++' means 30% to 50% increase from live body weight before incubation , &Quot; +++ " means an increase of more than 50% from the live body weight before culturing.

Mutant
lines Description Treatment Control group JBCS1294 Col-0 Wild type - +++ eir1 Auxin transport deficient and ethylene insensitive - - gai1 Gibberellin insensitive - - cre1 Ethylene and cytokinin insensitive - ++ cbb1 Brassinosteroid insensitive - ++ etr1 Ethylene insensitive - +

As shown in Table 2 and FIG. 7 and FIG. 8, it was confirmed that the growth of Arabidopsis thaliana (Col-0) stressed by 100 mM NaCl was increased by the volatile substances produced by the strain JBCS1294, Length and fresh weight were increased by 145.8% and 161.5%, respectively. In addition, as shown in Fig. 7, the number of roots and the number of quadratic roots were increased to 122.9% or 390% in the experimental group (Fig. 7B) in which the JBCS 1294 strain was dropped, compared with the control group .

In addition, as shown in Table 2 and FIG. 8, the fresh weight of cre1 , cbb1 and etr1 mutant Arabidopsis was 133.3%, 131.6% and 117.2%, respectively, and stem lengths were 142.3%, 141.7% and 146.2% , And the number of roots and the number of roots were greatly increased.

On the other hand, as shown in Table 2 and FIG. 8, in the case of the Arabidopsis thaliana, in the case of the eir1 and gai- 1 mutant Arabidopsis, no differences were observed in the fresh weight, stem and root length, It was confirmed that there was no increase by the volatile substances of the JBCS 1294 strain.

From the above results, it was expected that the growth of the JBCS 1294 strain by volatile substances was related to the auxin and gibberellin production signaling pathways and not to the cytokinin, ethylene and bresinosteroid synthesis pathways.

Example  2-5. JBCS1294 of Salt resistance  Identification of mechanism (2)

From the results of the above Example 2-4, Aminoethoxyvinylglycine (AVG; 5 μM, Soeno et al .) Was used as a synthesis inhibitor of auxin, bresinosteroid, cytokinin and gibberellin, which are volatile hormones, al ., 2010), propiconazole (PRZ; 1 nM, Hartwig et . al, 2012), cycloheximide ( CHX;? 1, Elliott & Koltunow, 1983), and daminozide (DZ; 20 μM, Rademacher , 2000) a was used, as the ethylene synthesis inhibitor is aminooxyacetic acid (AOA; 30 μM, Ishii meat al ., 2010) and silver nitrate (SN; 10 μM, Negi et al ., 2008) were used.

Specifically, the experiment using the plant hormone inhibitor was carried out in the same manner as in Example 2-4 except that the plant hormone inhibitor was contained in the medium in which the Arabidopsis seeds were inoculated. The experimental results are shown in Table 3 and FIG. In the following Table 3, '-' means no increase in live body weight, '+' means less than 30% increase from the pre-culture live weight, and '++' means more than 30% increase from live body weight before culturing.

Inhibitor Plant hormones inhibited Treatment Control group JBCS1294 The control (Col-0) Wild type - +++ Aminoethoxyvinylglycine (AVG) Auxin - - Cycloheximide (CHX) Cytokinin - - Propiconazole (PZ) Brassinosteroid - ++ Aminooxy acetic acid (AOA) Ethylene - ++ Silver nitrate (SN) Ethylene - + Daminozide (DZ) Gibberellin

As shown in Table 3 and FIG. 9, in the case of treatment with AVG as the auxin inhibitor or DZ as the gibberellin inhibitor, in the case of the Arabidopsis thaliana Col-0 exposed to the volatile substance of the JBCS 1294 strain, . On the other hand, when treated with CHX, the brassinosteroid inhibitor, PRZ, and the ethylene inhibitors AOA and SN, stem lengths increased by 133.3%, 128.6%, 113.8%, and 134.5%, respectively, Respectively.

From these results it was expected that the volatiles of the strain JBCS1294 were associated with auxin or gibberellins and these results indicate that the growth in cre1 , cbb1 and etrl mutant Arabidopsis thaliana was not increased in Example 2-4 Respectively.

Example  2-6. JBCS  1294 Salt resistance  Identification of mechanism (3)

From the results of Examples 2-4 and 2-5, in order to identify the growth-related volatile organic compounds of the JBCS1294 strain expected to be related to auxin or gibberellins, SPME-GC / MS (Headspace solid phase microextraction gas chromatography / mass spectrometry).

Specifically, 10 μl of the JBCS1294 strain (1 × 10 ⁷ CFU / ml) was added to the LB medium in order to analyze the volatile organic substances produced by the JBCS1294 strain, and the lid was replaced with the bottom portion of another petridish to form parafilm It was tightly sealed and then cultured at 25 ° C. After incubation for 24 hours, the volatile organic compounds in the headspace were adsorbed to SPME fiber (50/30 μm DVB / CARBOXEN-PDMS; Supelco, USA) , 0.25 μm) using a capillary column and analyzed with GC (HP 6890A) / MS (HP 5973; Agilent Technologies, USA). The analyzed material was identified based on the National Institute of Standards and Technology (NIST / EPA / NIH) Mass Spectral Library (Data Version: NIST 05, Software Version 2.0d). The results of the above analysis are shown in Table 4 and FIG. The number of peaks in Table 4 below means the number indicated in the graph of FIG.

Peak Retention time (minutes) % total peak area Compound name One 19.651 6.910 Butanoic acid 2 25.407 1.357 S-Methyl 3-methylbutanethioate 3 25.914 1.580 Propanoic acid 4 32.948 24.048 Butanoic acid 5 41.370 4.030 Benzoic acid 6 51.036 1.040 9,12,15-Octadecatrienoic acid 7 52.575 0.828 9,12,15-Octadecatrienoic acid 8 54.379 0.772 1-Monolinoleoylglycerol trimethylsilyl ether 9 55.978 26.418 Hexanedioic acid 10 57.242 3.452 4-Methyl-6-phenyltetrahydro-1

As shown in Table 4 and FIG. 10, the main constituents of volatile substances were hexanedioic acid (total peak area 26.42%, RT 55.978 min) and butanoic acid (ca. 24.05%, RT 32.948 min) 1.04%), propanoic acid (ca. 1.58%, RT 25.914 min), linolenic acid (ca. 1.04%), benzoic acid (ca. (RT, 51.036 min), 4-Methyl-6-phenyltetrahydro-1 (ca 3.45%, RT 57.242 min) and 1-monolinoleoylglycerol trimethylsilyl ether (ca. 0.77%, RT 54.379 min). From the above results, it is expected that the volatile organic substances exhibiting the salt tolerance and growth promoting ability produced by the JBCS 1294 strain are related to hexanedioic acid or butanoic acid.

Example  2-7. JBCS  1294 Salt resistance  Identification of mechanism (4)

Changes in the proline content in the plant were further confirmed in relation to the mechanism of the volatile organic substances related to the salt tolerance and growth promotion of the JBCS 1294 strain confirmed from the results of Examples 2-4 to 2-6.

Specifically, as in Examples 2-4 and 2-5, Arabidopsis Col-0 and the mutation line were exposed to volatile substances produced by the strain JBCS 1294, and after 14 days, the Arabidopsis thaliana was collected to obtain proline content Respectively.

The measurement of the proline content was carried out in the following manner. First, 300 mg of Arabidopsis seedlings were ground with liquid nitrogen and suspended in 1 ml of 3% sulfosalicylic acid. After the centrifugation, the supernatant was collected and mixed with 0.2 mL of acid ninhydrin and 0.2 mL of glacial acetic acid. The mixture was mixed well and treated at 100 ° C for 1 hour. The reaction was stopped by adding ice to the reaction mixture. The reaction mixture was then quenched with 0.4 mL of toluene. The upper layer was examined for absorbance at 520 nm and its content was compared with a standard curve determined using pure proline. The measurement results are shown in Fig.

As shown in FIG. 11, in the Col-0 seedlings exposed to the volatile substances produced by the JBCS1294 strain, the proline content was increased by 168.6% compared to the untreated ones. The eir1 , cre1 and etr1 mutant Arabidopsis thaliana , And Col-0 Arabidopsis treated with CHX, PRZ and SN (FIG. 11B). Specifically, the eir1 mutation increased by 125% compared to the control group, and the proline content of the Col-0 seedlings treated with AVG was 11.8% higher than that of the control group, and these results were found to be in agreement with each other. In addition, the proline content in the gai1 mutant did not increase significantly (Fig. 11a), and the proline content in the DZ-treated Col-0 seedlings did not increase significantly (Fig. 11b) Was interpreted.

On the other hand, the proline content in the cbb1 mutant decreased by 111.4%, and the proline content in the AOA treated group of the hormone inhibitor treated group was decreased by 107.8%.

Example  2-8. JBCS1294 of Salt resistance  Identification of mechanism (5)

The influence of the volatile substances on gene expression in the plant body was further confirmed with respect to the mechanism of the volatile organic substances related to the salt tolerance and growth promotion of the JBCS 1294 strain confirmed from the results of Examples 2-4 to 2-7 .

Specifically, as in Examples 2-4 and 2-5, Arabidopsis Col-0 was cultivated for 10 days, then exposed to volatile substances produced by the JBCS 1294 strain for 24 hours, seedlings were collected, Stem and root portions were separated, and then subjected to grinding using liquid nitrogen. RNA was isolated using TRI Reagent (Invitrogen, Carlsbad, USA).

The separated RNAs were amplified by SMALL AUXIN UP RNA (SAUR), HISTIDINE KINASE1 (AHK1), ETHYLENE RESPONSE (AHK1) for auxin, cytokinin, cDNA brassinosteroid, cDNA ethylene and gibberellin synthesis pathway using PrimeScript RT reagent kit The expression levels of AtHKT1, AtSOS1, AtNHX1, and AtAVP1 genes were investigated in order to investigate the expression of ion channel and transporter expression, and the expression levels of FACTOR (ERF), BRASSINOSTEROID BIOSYNTHETIC CYTOCHROME P450 (CPD) and GIBBERELLIN 3-BETA-DIOXYGENASE (GA3OX3) The expression level was examined.

The gene expression level was examined by qPCR (Rotor-Gene 6000; Corbet Research, Mortlake, Australia) using SYBR Select Master Mix (Applied Biosystems, USA). The PCR was carried out by using the respective primers described in Table 5 for 15 minutes at 95 ° C, followed by 45 cycles at 95 ° C for 15 seconds and 60 ° C for 60 seconds. Ion channel and transporter expression were measured by Jha et al . (2010), and the primers described in Table 6 below were used. Actin was used as a control and Delta-Delta Ct method (Schmittgen et al . 2000). The same experiment was repeated three times, and the average value was shown in FIG.

Gene name NCBI Accession No Primers Sequence (5'-3 ')
(Forward and Reverse) SAUR-like auxin-responsive protein (SAUR) NM_128001 Forward: 5'-TTCGGTCGGGTTAGAGAAGA-3 '
Reverse: 5'-TCACCAGGTACCCTTCGTTC-3 ' Histidine kinase 1 (AHK1) NM_127335 Forward: 5'-TGCGAGTGCCAAAGCTCTAA-3 '
Reverse: 5'-GGCAGACATGTTCCCGTGTA-3 ' Ethylene response factor AP2 domain transcription factor (At2g31230) (ERF) AY070443 Forward: 5'-TCAAGCCTTCTTTTGCCCTA-3 '
Reverse: 5'-GGCCTTTTCCTCACTCCTCT-3 ' Cytochrome P450 90A1 (CPD) Forward: 5'-GAATGGAGTGATTACAAGTC-3 '
Reverse: 5'-GTGAACACATTAGAAGGGCCTG-3 ' Gibberellin 3-beta-dioxygenase 3 (GA3OX3) NM_118289 Forward: 5'-TCTTCGACGTCACGCTACAC-3 '
Reverse: 5'-CAAACCGGGTAGGAGTTCAA-3 ' Actin Forward: 5'-GAAAAGATCTGGCATCACACTTTCTA-3 '
Reverse: 5'-ACATACATAGCGGGAGAGTTAAAGGT-3 '

Gene name Source Primers
(Forward and Reverse) Sequence (5'-3 ') HIGH-AFFINITY K + TRANSPORTER1 (AtHKT1; 1) Jha et al. (2010) Forward: 5'-TGCAAACTGCGGATTTGTCC-3 '
Reverse: 5'-TGAGCAAAACCAAGAAGCAAGG-3 ' ARABIDOPSIS SALT OVERLY SENSITIVE 1 (AtSOS1) Forward: 5'-CTGCTTGCTACATTTCTGCTG-3 '
Reverse: 5'-TGCTTCCTCTCCTTCCTTTTC-3 ' NA + / H + EXCHANGER 1 (AtNHX1) Forward: 5'-ACTCATAAGCTACCTATTACCG-3 '
Reverse: 5'-GGTCTCGAGTTACTAAGATCAGGAGGGTTTCTC-3 ' ARABIDOPSIS THALIANA V-PPASE 3 (AtAVP1) Forward: 5'-GCAGCTCTTAAGATGGTTGAA-3 '
Reverse: 5'-AGAGGTGTGAGCATGACAAGG-3 '

As shown in FIG. 12, in the Col-0 seedlings exposed to the volatile substances produced by the JBCS1294 strain, the SAUR gene expression was 1.71 ± 0.06 times higher than that of the untreated root, whereas it was 0.13 ± 0.02 Respectively. In addition, the expression levels of AHK1, CPD and ERF in the stem were increased to 1.64 ± 0.02, 11.23 ± 0.34 and 4.14 ± 0.41 times, respectively. The expression level of ERF in the root was also increased similarly to stem, while the expression level of GA3OX3 gene was 0.35 ± 0.07 fold, and the expression in the root was insignificant.

In the case of Col-0 seedlings exposed to the volatile substances produced by the JBCS1294 strain, the expression level of AtHKT1 gene in roots was increased by 1.7 ± 0.03 times, while that of stem was decreased by 0.27 ± 0.06 times as compared with that of untreated root. The expression levels of SOS1 and AVP1 genes were 3.29 ± 0.14 and 1.414 ± 0.18 and 3.29 ± 0.14 and 1.414 ± 0.18 times higher in the root and stem, respectively. On the other hand, the expression level of NHX1 gene increased 5.61 ± 0.81 times in the roots compared to the control, but decreased by 0.77 ± 0.09 times in the stem. From the above results, it was determined that the expression of AtHKT1 and NHX1 genes in the stem by the volatile substances produced by the JBCS 1294 strain played an important role in the induction of salting resistance.

Example  3. Identification of pathogen resistance of isolated microorganisms

Example  3-1. JBCS  Identification of the control effect of 1294 strain on soybean (1)

In order to confirm the controlling effect of the JBCS1294 strain on the soybean, specifically, the controlling effect on the soybean flour disease, the following experiment was carried out

Specifically, the JBCS 1294 strain was inoculated into an LB culture medium, cultured at 28 ° C and 250 rpm, and centrifuged at 4000 x g for 15 minutes to obtain cells. Thereafter, the cells were diluted to a concentration required for 0.05 M phosphate buffer (pH 6.5) or distilled water.

First, the filter paper was soaked with distilled water in a Petri dish and the leaves were taken from mature beans. Then, the above JBCS 1294 strain was inoculated with a diluted solution adjusted to be 1 x 10 ⁶ , 1 x 10 ^7, and 1 x 10 ⁸ CFU / ml. After 2 hours of inoculating the strain, the X. a. pv. glycines were adjusted to a concentration of 5 × 10 ⁷ CFU / ml and then treated. Petri dishes treated in this manner were stored for 7 days under light condition of 16/8 hr (light / dark), temperature condition of 25 캜 and relative humidity of 95%, and then the leaves were observed to investigate the onset area. The measurement results are shown in Fig. 13 and Fig.

As shown in FIG. 13 and FIG. 14, as the concentration of the antagonist was increased, the control effect was increased. The incidence of the treatment at 1 × 10 ⁶ CFU / ml was 85% , But the incidence was reduced to 60% and 20% at 1 × 10 ⁷ and 1 × 10 ⁸ CFU / ml, respectively. From the above results, it was confirmed that when the strain of JBCS1294 of the present invention was treated with 1 × 10 ⁸ CFU / ml or more, the control effect against the soybean blight was 77.8%.

Example  3-2. JBCS  1294 (2) Determination of Control Effect on Soybean

In order to further confirm the controlling effect of the soybean, the same experiment was carried out in the greenhouse.

Specifically, soybean seeds (Dae-kwang) were sown in pots (1 seedling / pot) containing horticultural soil (Sanglim, Korea) and cultured in a plant growth room (light / dark, 16/8 hr, 28 ° C) The same amount of water was supplied to each treatment as needed every day, and it was used to test the control effect after 5 weeks of cultivation. On the other hand, JBCS 1294 strain (1 × 10 ⁸ CFU / ml) cultivated in the same manner as in Example 3-1 was sprayed on a leaf of bean to form a thin film using an atomizer, and then, in a plant growth room After 1 hour, the soybean flour ( X. a . glycines , 1 x 10 < ⁸ > CFU / ml). The plants inoculated with the pathogens were placed in a transparent plastic box and maintained at a relative humidity of 95% for 24 hours, and then cultivated in a plant growth room for 10 days. The incidence was calculated by the following equation. To mean X ₀ = the number of disease free ranging leaf in the formula, and the X ₁ are yibyeongyeop area (ILA) This means a number of less than 5% leaf, X ₂ is yibyeongyeop area (ILA) is 6% to 10% of leaf X ₃ means the number of leaves with an ILA of 11% to 25%, X ₄ means the number of leaves with an ILA of 26% to 50%, ILA , And X ₅ means the number of leaves having 51% to 100% cross-sectional area (ILA). The results of the incidence measurement by the above calculation are shown in Fig.

[formula]

[Σ (leaf number X1 × 1 + X ₂ × 3 + X ₃ × 5 + X ₄ × 7 + X ₅ × 9) / total leaf number irradiated] × 100

As shown in FIG. 15, when the strain of JBCS 1294 was treated with 1 × 10 ⁸ CFU / ml, it was found to be 32.6, whereas it was found to be 75.8 in the control without treatment, and the strain of JBCS 1294 was found to be 1 × 10 ⁸ CFU / ml, the control effect was confirmed to be about 57.0%.

Example  3-2. JBCS1294 Of growth promotion effect on soybean

In order to confirm the growth promoting effect of the JBCS1294 strain on the soybean, the following experiment was conducted. First, the JBCS1294 strain was adjusted to a concentration of 1 × 10 ⁸ CFU / ml and 0.2% CMC was added thereto. Then, 10 g / 50 ml of soybeans (10 g / 50 ml) was immersed therein and the mixture was incubated at 28 ° C. and 150 rpm for 3 hours And treated to adhere the soybean seeds to the bacteria.

Soybeans with the bacteria were dried at room temperature and seeded on the soil, cultivated under light condition of 16/8 hours (light / dark) and 28 ° C. After 14 days of cultivation, fresh weight was measured, and the length of roots and stem was measured to calculate the vitality index as above. As a control, soybeans without any treatment were grown under the same conditions, and then live weight and vitality index were measured. The measurement results are shown in Fig.

As shown in FIG. 17, when the JBCS 1294 strain was treated, the fresh weight and the vigor index were increased by 21.6% and 38.4%, respectively, and the stem and root lengths increased by 13.1% and 11.5% Thus, it was confirmed that when the JBCS 1294 strain was treated, there was also an effect of promoting growth as well as control of soybean blight.

Agricultural Biotechnology Research Institute KACC91943 20140509

Claims (10)

Alcaligenes faecalis JBCS1294 strain deposited with Deposit No. KACC91943P, which has the ability to induce resistance to salt tolerance and growth in plants. The method according to claim 1,
The strain is an alkaline zinispicillis JBCS 1294 ( Alcaligenes < RTI ID = 0.0 > faecalis JBCS1294) strain. Having the ability to induce resistance to salt tolerance and growth in plants, and an alkaline zinispicillis JBCS1294 deposited with the deposit number KACC91943P ( Alcaligenes faecalis JBCS1294) strain, a culture of the strain, a concentrate of the culture, and a dried product of the culture as an active ingredient. An Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce salt tolerance and growth promoting ability to plants, a culture of the strain, a concentrate of the culture, and a dried product of the culture A microbial fertilizer containing at least one selected from the group as an active ingredient. An Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce salt tolerance and growth promoting ability to plants, a culture of the strain, a concentrate of the culture, and a dried product of the culture A composition for preventing blight of soybean flour containing at least one selected from the group as an active ingredient. 6. The method of claim 5,
The above-mentioned soybean flour is caused by Xanthomonas axonopodis pv. glycines of kongbul blight control composition. An Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce salt tolerance and growth promoting ability to plants, a culture of the strain, a concentrate of the culture, and a dried product of the culture A method for promoting plant growth, comprising treating at least one selected from the crowd into a soil, a plant or a plant seed. 8. The method of claim 7,
The method for promoting the growth of plants is an Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having an ability to induce salt tolerance and growth promoting ability, a culture of the strain, a concentrate of the culture, Wherein the plant is cultivated in a liquid state, and at least one species selected from the group consisting of dry matter of water is used in a plant-like manner, and seeds of crops are dipped or sprayed or coated on seeds. An Alcaligenes faecalis JBCS1294 strain deposited with the deposit number KACC91943P having the ability to induce salt tolerance and growth promoting ability to plants, a culture of the strain, a concentrate of the culture, and a dried product of the culture A method for promoting plant resistance to stress, comprising treating at least one selected from the group to soil, plants or plant seeds. 10. The method of claim 9,
Wherein the stress of the plant is salty.

Images