KR100849407B1 - Method for improving plant growth using the self-circulation system of soil-favorable microorganisms - Google Patents

Abstract

The present invention relates to a method for promoting plant growth using a self-circulating system of soil-friendly microorganisms, specifically 1) separating the soil-friendly microorganisms by culturing the soil sample collected from the farmland in a medium containing the soil of the farmland. ; 2) preparing a complex culture of isolated soil-friendly microorganisms; 3) predominantly spraying the soil-friendly microorganisms by spraying the complex culture on the arable land; And 4) cyclically repeating steps 1), 2) and 3). Plant growth promotion method according to the present invention can promote the growth of various plants without causing environmental pollution because it uses its own circulation system of soil-friendly microorganisms to increase productivity and improve agricultural technology for organic farming, soil pollution It can be usefully used for restoration.

Description

METHODS FOR IMPROVING PLANT GROWTH USING THE SELF-CIRCULATION SYSTEM OF SOIL-FAVORABLE MICROORGANISMS}

1 shows a conceptual diagram of a self-circulating system of soil-friendly microorganisms,

Figure 2 is a result of incubating the soil sample collected from the arable land in a minimum medium for separation,

Figures 3a , 3b and 3c respectively show three soil-friendly microorganisms PY-01, PY-02 and PY-03 isolated purely after the incubation of soil samples according to the present invention,

FIG. 4 shows the growth of kidney beans in culture soils mixed with soil cultures of soil-friendly microorganisms PY-01, PY-02, and PY-03 isolated in accordance with the present invention mixed with soil of the corresponding cropland. The photo is a comparison.

5a to 5c is a photograph showing that the self-circulation system of the soil-friendly microorganism according to the present invention promotes the growth of plants even in the actual farmland environment.

The present invention comprises the steps of: 1) separating the soil-friendly microorganisms by incubating the soil sample collected from the arable land in a medium containing the soil of the arable land; 2) preparing a complex culture of isolated soil-friendly microorganisms; 3) predominantly spraying the soil-friendly microorganisms by spraying the complex culture on the arable land; And 4) repeating the steps 1), 2) and 3), and a method of promoting plant growth using a self-circulating system of soil-friendly microorganisms.

In modern agriculture, chemical fertilizers and organic synthetic pesticides are used in large quantities. They are very useful as protection means of crops, and have played an important role in increasing crop production, stable supply, and reducing the labor force of agricultural workers. Recently, however, it has been pointed out that the use of repeated pesticides causes the emergence of drug-resistant bacteria, contamination of groundwater, soil erosion, adverse effects on the environment such as reduction of wildlife habitat, and also harmful to human body. Gradually, their use is limited. As a result, the promotion of environmentally-friendly agriculture is required, and interest in organic farming is increasing for the safety of agricultural products.

As an alternative to solve this problem, much attention has been focused on the development of biological pesticides, which are pests such as insects, mites and nematodes that harm crops by directly using microorganisms or by using microorganism-containing preparations. It is a pesticide used to control prophylaxis or weeds of various plant diseases. In particular, as interest in biological control to use microorganisms themselves or secondary metabolites produced by microorganisms as an alternative means to increase the productivity of crops by controlling only the various diseases of crops, research in the field of microbial preparations in developed countries It is actively underway.

As described above, biological control using microorganisms has attempted to suppress plant pathogens using many kinds of bacteria and fungi since the early 1900s. Most of the microbial agents reported so far are Agrobacterium sp. And Bacillus. Bacillus sp., Pseudomonas sp., Streptomyces sp., And bacteria such as Trichoderma sp.

On the other hand, with the recent emphasis on the importance of eco-friendly agriculture, the necessity of restoring soils acidified and contaminated by the use of chemical fertilizers and pesticides is expected to increase the demand for microbial preparations for soil improvement as well as biological control purposes. .

However, most of the microbial preparations for soil improvement currently used in Korea are dependent on imports, and the method is also limited to the application of only a few kinds of specific microorganisms to areas where necessary. However, this method does not take into account the characteristics of the distribution of microorganisms in each region, which may be a factor that destroys the ecosystem of microorganisms in the soil of a specific region. Therefore, in order to promote soil improvement and plant growth in a specific area, only a soil-friendly microorganism distributed in the area is required to be separated and mass cultured and then sprayed on the area again.

Therefore, the present inventors have made intensive research to develop a method of using soil-friendly microorganisms that can effectively satisfy two purposes of environmental purification and food crop production. As a result, the soil samples collected from the arable land contain the soil of the arable land. After separating and cultivating soil-friendly microorganisms in one medium, preparing a complex culture of isolated soil-friendly microorganisms, and discovering that the complex culture can be environmentally friendly to improve plant growth, The present invention has been completed.

Accordingly, an object of the present invention is to separate the microorganisms having excellent soil affinity from the cultivated land to increase the productivity of environmentally friendly crops, and to spread the complex cultures on the cultivated land again, and to promote plant growth. To provide.

In order to achieve the above object, the present invention

1) separating the soil-friendly microorganisms by accumulating the soil sample collected from the arable land in a medium containing the soil of the arable land;

2) preparing a complex culture of isolated soil-friendly microorganisms; And

3) predominantly spraying the soil-friendly microorganisms by spraying the complex culture on the arable land; And
4) It provides a method for promoting plant growth using the self-circulation system of the soil-friendly microorganism, including the step of repeating the steps 1), 2) and 3).

In the present invention, the "self-circulating system of soil-friendly microorganisms" refers to soil-sprayed by separating and cultivating soil-friendly microorganisms from the soil of a specific region by the integrated culture method, and then spraying the cultured soil-friendly microorganisms on the soil of the region. By a system that continually circulates the way in which affinity microorganisms dominate (see FIG. 1 ).

Hereinafter, the method will be described in more detail step by step.

Step 1) is a step of separating soil-friendly microorganisms from the arable land. The soil sample collected from the farm is mixed with physiological saline, and then the mixed solution is inoculated into a separation medium containing 1 to 10% by weight of the soil of the arable land. And incubate for 15 to 20 days at 25 to 30 ° C. The diluted solution obtained by diluting the culture solution with physiological saline is plated on a separation medium and incubated at 25 to 30 ° C. for 5 to 7 days, and colonies formed on the medium are separated into soil-friendly microorganisms of the farmland.

The minimum medium for separation that can be used is usually any medium used for the separation of soil microorganisms can be used, in a preferred embodiment of the present invention NH ₄ Cl 0.1 to 0.2% (w / v), K ₂ HPO ₄ 0.44 to 0.54% (w / v), NaH ₂ PO ₄ 2H ₂ O 0.4 to 0.5% (w / v), MgSO ₄ 7H ₂ O 0.05 to 0.06% (w / v), CaCl ₂ · 2H ₂ O 0.0003 to 0.0004% (w / v), FeSO ₄ .7H ₂ O 0.0001 to 0.0002% (w / v), MnCl ₂ · 4H ₂ O 0.0001 to 0.0002% (w / v), CoCl ₂ · 6H ₂ O 0.00001 A minimum medium comprising from 0.00002% (w / v), Na ₂ MoO ₄ .2H ₂ O 0.00001 to 0.00002% (w / v) and 0.0004 to 0.0005% (w / v) monohydrogen citric acid.

In addition, the separation medium may be any medium used for cultivation of soil microorganisms in general, in a preferred embodiment of the present invention 0.4 to 0.5% by weight of peptone, 0.2 to 0.3% by weight of the juice (Beef extract) and agar Separation medium containing 1.0 to 1.5% by weight is used.

In a preferred embodiment of the present invention, three strains of PY-01, PY-02, and PY-03 are separated from the farmland soil as a strain having excellent soil affinity.

The strains, by physiological characterization, PY-01 can reduce nitrate to nitrite, break down glucose, gluconate, caprate, adipate, malate, citrate, phenyl-acetate to produce acid , Alcaligenes xylosoxidans , which exhibits cytochrome oxidase activity, PY-02 can hydrolyze gelatin, decompose caprate to produce an acid, and exhibits cytochrome oxidase activity Cellar birosa is identified as (Weeksella virosa), PY-03 will have an Esculin be hydrolyzed to break down glucose and maltose, and formation of the acid, Breda beundi Pseudomonas Bezier Temecula less (Brevundimonas vesicularis) representing the cytochrome oxidase activity (See Table 1 ).

From this, it can be seen that alkali-genes xylososidans, wickella virosa and ravendemonas vesicularis exist as soil-friendly microorganisms in the arable land. The alkali Zenith xylososidans have been previously reported as a microorganism that degrades pesticides, Wickella virosa is known as a microorganism found in the urine, and the Brevendimonas vesicularis is known as a soil microorganism of the general Pseudomonas strain.

Step 2) is a step of preparing a complex culture of the strains separated by soil-friendly microorganisms in step 1), because the strains are excellent in soil affinity, the complex culture obtained by culturing them in a large amount of components that promote plant growth It contains.

The complex culture was inoculated in a culture medium containing 0.4 to 0.5% by weight of peptone and 0.2 to 0.3% by weight of the soil-friendly microorganism isolated from the arable land according to step 1) 25 to 30 ℃, 150 to 200 Obtained by incubating for 48 to 72 hours under the condition of rpm. In a preferred embodiment of the present invention, inoculated with alkaline gene xylososidans, Wickcellar virosa and Bravendemonas vesicularis isolated in soil-friendly microorganisms in the culture medium, and incubated for 48 hours under conditions of 25 ℃, 200 rpm To obtain a complex culture.

Biopolymers, protective agents and / or nutrients may be added to the complex culture, and biopolymers, protective agents and nutrients may be mixed and used in different ratios.

The biopolymer is preferably at least one selected from the group consisting of corn starch, potato starch, soy flour, flour, rice flour and glutinous rice flour. Such biopolymers act as protective agents to protect microorganisms from sunlight as well as the role of an electrodeposition agent and a binder to allow microbial strains in a complex culture to adhere well to the leaves of plants.

In addition, the protective agent acts as an emulsifier to encapsulate the microorganisms to protect them from ultraviolet rays and to mix the components of the formulation well, preferably one or more selected from the group consisting of soybean oil, corn oil and olive oil.

In addition, the nutrient is preferably a sugar as a nutrient source of the microorganism, the sugar may be brown sugar, white sugar or brown sugar.

The complex culture of the present invention can be used in the form of the culture medium of the soil-friendly microorganisms as it is or in various forms, it can be prepared in the form of a hydrate, liquid or emulsion. In addition, the composite culture of the present invention may further include an excipient, a diluent, a diatomaceous earth, a zeolite, a mineral carrier such as white carbon, various additives, or a surfactant.

Step 3) spraying the complex culture of the soil-friendly microorganisms prepared as described above to the soil of the arable land and left for 10 to 15 days so that the soil-friendly microorganisms in the complex culture to adapt to the soil environment and cultivate the crop therein Step. As a result, a population of soil-friendly microorganisms among the indigenous microorganisms may exist as dominant bacteria in the arable land, thereby promoting plant growth. At this time, the microbial complex culture is preferably sprayed on the soil at a concentration of 1 × 10 ⁷ to 1 × 10 ¹⁰ cells / ㎖ per unit area (㎡).

Step 4) is a step of repeating the process of steps 1) to 3) 1 to several times as necessary in consideration of the soil condition of the arable land, the climatic environment, the type of crops, and the like.

In a preferred embodiment of the present invention, a mixed culture of alkaline gene xylososidans, wickella virosa and Bravendemonas vesicularis isolated with soil-friendly microorganisms is mixed with the soil of the arable land from which the microorganisms are separated, Planting the kidney beans confirms the excellent growth compared to the kidney beans planted in the culture soil not mixed with the complex culture (see Figure 4 ). In addition, in experiments targeting actual arable land rather than small environments such as pollen, it is confirmed that the growth of tomatoes, kidney beans, and radish in the arable land sprayed with the microbial complex culture cultured and separated from the arable land is excellent ( FIGS. 5A to 5A ) . 5c ).

Therefore, as proposed in the present invention, in order to promote soil improvement and plant growth of the arable land, only soil-friendly microorganisms distributed on the arable land are separated and cultured in large quantities, and then the soil-friendly microorganisms are sprayed only on the arable land. The use of a circulatory system will not only prevent the destruction of ecosystems in the soil, but also contribute to improving incomes, improving water quality, and improving the rural environment by restoring contaminated soil and increasing soil crop productivity.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.

Example 1 Isolation of Soil-Friendly Microorganisms

After adding 10 ml of physiological saline to 1 mg of soil samples collected from farmhouses, 1 ml of the mixed solution was mixed with soil (soil 2.0% (w / v), NH ₄ Cl 0.1% (w / v). ), K ₂ HPO ₄ 0.44% (w / v), NaH ₂ PO _{4 ·} 2H ₂ O 0.4% (w / v), MgSO _{4 ·} 7H ₂ O 0.05% (w / v), CaCl _{2 ·} 2H ₂ O 0.0003% (w / v), FeSO ₄ 7H ₂ O 0.0001% (w / v), MnCl ₂ 4H ₂ O 0.0001% (w / v), CoCl ₂ 6H ₂ O 0.00001% (w / v), Na ₂ MoO _{4 ·} 2H ₂ O 0.00001% (w / v) and 0.0004% monohydrogen citric acid ( ₀₄ % (w / v), pH 7.0) was inoculated three times inoculated three times and then incubated at 25 ℃ for 15 days. After diluting the culture solution at a ratio of 1: 1000 using physiological saline, 0.1 ml of the diluted solution was plated on a separation medium (0.5% by weight of peptone, 0.3% by weight of beef extract, 1.5% by weight of agar, and pH 7.0). Incubated for 5 days at ℃ ( Fig. 2 ). Three colonies generated on the medium were isolated and named as PY-01, PY-02 and PY-03, respectively ( FIGS. 3A- 3C ).

Example 2 Identification of Soil-Friendly Microorganisms

As a result of Gram staining of the three strains isolated in Example 1, all of them were found to be Gram-negative. The physiological characteristics of the strains were analyzed by API kit 20NE (Bio Merieux, France). As shown in Table 1 below, PY-01 can reduce nitrate to nitrite, glucose, gluconate, and caprate. The adipate, malate, citrate, and phenyl-acetate were decomposed to produce an acid and exhibited cytochrome oxidase activity. PY-02 was able to hydrolyze gelatin, decompose caprate to produce acid, and exhibited cytochrome oxidase activity. PY-03 can hydrolyze esculin, decompose glucose and maltose to produce acid, and exhibit cytochrome oxidase activity.

Analysis item PY-01 PY-02 PY-03 Reduction of Nitrate to Nitrite + - - Indole productivity - - - Glucose acidification - - - Arginine dehydrolase - - - Urease - - - Esculin hydrolysis (β-glucosidase) - - + Gelatin Hydrolysis (Protease) - + - β-galactosidase - - - Glucose assimilation + - + Arabian fairy tale - - - Mannos Fairy Tale - - - Mannitol fairy tale - - - N-acetyl-glucosamine assimilation - - - Maltose fairy tale - - + Gluconate assimilation + - - Caprate Fairy Tale + + - Adipate fairy tale + - - Malate fairy tale + - - Citrate Fairy Tale + - - Phenyl-acetate assimilation + - - Cytochrome oxidase + + +

From the above analysis results, PY-01 strain isolated as soil-friendly microorganism in the present invention "alkaligenes xylosocydanso", PY-02 strain is "Wickella Virosa", PY-03 strain is "Braven De Monas" Confirmed as "Vegaculis"

Example 3 Confirmation of the Plant Growth Promoting Effect of the Microbial Complex Culture-Pollen Environment

The three soil-friendly microorganisms identified in Example 2 were inoculated together in 250 ml of culture medium (0.5% by weight of peptone, 0.3% by weight of juice), and cultured for 48 hours under conditions of 25 ° C. and 200 rpm. Got it. The complex culture was inoculated in the culture soil (promotional fertilizer, green power) containing 50% of soil (v / v) in the area where soil samples were taken and mixed well in an amount of 5% (v / v) for 25 days. Cultured in a plant growth growth chamber (Growth Chamber, Korea Science) maintained at ℃. Thus, by planting five kidney beans pre-sprouted in the culture soil in which the microorganisms were cultured and the culture soil without the microorganisms (top soil), the kidney beans were grown while incubating in a plant growth capacity chamber maintained at 25 ° C, 80% humidity, and roughness 256. Compared.

Figure 4 is a photograph of the growth of kidney beans in cultured soil inoculated with microbial complex culture and inoculated microbial complex culture. As shown in Figure 4 , it can be seen that the growth of kidney beans in the culture soil inoculated with a complex culture of microorganisms isolated from the self-circulation system of the soil-friendly microorganism of the present invention.

<Example 4> Confirmation of plant growth promoting effect of the microorganism complex culture-arable land environment

Soil-friendly microorganisms were separated from the soil collected from the plastic house of Dongguk University experimental farm located in Ilsan, Gyeonggi-do in the same manner as in Example 1. The two soil-friendly microorganisms thus separated and identified were inoculated together in 250 ml of culture medium (0.5% by weight of peptone, 0.3% by weight of juice), and cultured for 48 hours under conditions of 25 ° C. and 200 rpm to obtain a complex culture. . 250 ml of this complex culture was inoculated into 2000 ml of the same culture medium, followed by culturing for 3 days at 25 ° C. and 300 rpm using a jar fermenter to obtain a large amount of complex culture. The complex culture was diluted in 100 l of water and incubated for three more days, and the soil sample was collected at a concentration of 5 × 10 ⁹ cells / ml per unit area (㎡), ie, vinyl from an experimental farm at Dongguk University. The house was sprayed. Thus, the growth of the tomato, kidney beans and radish were planted in the soil (experimental group) and the non-sprayed soil (control group) to which the microbial complex culture was sprayed ( FIG. 5A ). The experiment lasted for 53 days from March 19, 2007 to April 23, 2007, and the temperature of the vinyl house was maintained at 25 to 35 ° C throughout the experiment.

5b and 5c are photographs of tomatoes, kidney beans, and radishes grown in arable lands not inoculated with the microbial complex cultures and in the inoculated microbial complex cultures. As a result, the growth of tomatoes, kidney beans, and radish was much better than that of the arable land ( FIG. 5B ) inoculated with the complex culture of microorganisms isolated from the self-circulating system of the soil-friendly microorganism of the present invention ( FIG. 5C ) . It can be seen.

As described above, the method of promoting plant growth using a complex culture of soil-friendly microorganisms isolated from the arable land according to the present invention, unlike chemical fertilizers or pesticides, indigenous microorganisms having a biofertilizer function for safe eco-friendly agriculture, In addition, it is possible to clean up the environment of rivers, rivers, and air, and to use organic materials usefully, not only to improve the productivity and quality of agricultural products, but also to reduce labor.

Claims (7)

Separating the soil-friendly microorganisms by accumulating the soil sample collected from the cropland to be grown in a medium containing the soil of the cropland; 2) preparing a complex culture of isolated soil-friendly microorganisms; 3) predominantly spraying the soil-friendly microorganisms by spraying the complex culture on the arable land; And 4) A method of promoting plant growth using a self-circulating system of soil-friendly microorganisms, comprising repeating steps 1), 2) and 3). delete delete delete The method of claim 1, The complex culture in step 2) was inoculated into a culture medium containing 0.4 to 0.5% by weight of peptone and 0.2 to 0.3% by weight of the soil-friendly microorganism isolated in step 1), and then 25 to 30 ° C. and 150 to 200 rpm. Method characterized in that it is prepared by incubating for 48 to 72 hours under the conditions of. The method of claim 1, In step 3), the complex culture of the soil-friendly microorganisms prepared in step 2) is sprayed on the soil of the arable land and left for 10 to 15 days to allow the soil-friendly microorganisms in the complex culture to adapt to the soil environment. The method of claim 1, In step 3), the complex culture is sprayed onto the soil at a concentration of 1 × 10 ⁷ to 1 × 10 ¹⁰ cells / ml per unit area (m 2).

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