Preparation method and application of microbial agent

Abstract

The invention belongs to the technical field of microbial agents, and particularly relates to a preparation method and application of a microbial agent. Microbial agents in the market at present generally have the problems of complex preparation mode, poor overall plant growth promoting effect and the like, and large-scale popularization and application of the microbial agents are restricted. The microbial agent provided by the invention is simple in preparation mode and obvious in overall plant growth promoting effect. The microbial agent provided by the invention is formed by mixing bacillus velezensis and saccharomycetes, and a finished product of the microbial agent can be prepared by carrying out single-bacterium microbial fermentation culture and then mixing according to a mass ratio. According to the invention, the influence of mixing in different proportions and application of the microbial agent on plant growth is researched, and the research on whether the microbial agent is applied or not shows that the roots and leaves of the plant to which the microbial agent is applied grow more vigorously than those of the plant to which the microbial agent is not applied. Researches on application of microbial agents with different mass ratios show that the growth promoting effect of 70%-80% of bacillus velezensis and 20%-30% of saccharomycetes is most obvious.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbial inoculants, and particularly relates to a preparation method and application of a microbial inoculant. Background Art

[0002] The growth and development of plants are affected by soil nutrients, microbial environment and external environmental conditions. At present, the main means to promote plant growth include chemical fertilizers, plant growth regulators and microbial inoculants. Chemical fertilizers (such as nitrogen fertilizers, phosphorus fertilizers, potassium fertilizers) can quickly supplement the nutrients required by plants, but long-term excessive application will lead to soil compaction, salinization and imbalance of microbial communities, and may pollute groundwater resources. Plant growth regulators (such as gibberellin, indole acetic acid, cytokinin) can directly affect the plant hormone level and promote growth, but excessive use may cause imbalance of plant hormones and affect the safety of agricultural products. In contrast, microbial inoculants have gradually become an important development direction in agricultural production due to their environmental protection, safety and sustainability advantages. However, existing microbial inoculants generally have problems such as single strain function, poor stability and limited adaptability, which restrict their large-scale popularization and application.

[0003] Bacillus velezensis is widely used in the field of plant disease resistance, and can secrete antibacterial peptides, volatile organic compounds, etc., inhibit a variety of plant pathogenic bacteria, and improve plant disease resistance. Saccharomyces is mainly used in the fields of food, beverage, feed, industrial ethanol, etc., and there is less research on its application in the agricultural field.

[0004] Based on the combined fermentation of Bacillus velezensis and Saccharomyces, the present invention aims to utilize the synergistic effect of the two to improve the growth promotion effect, stability and environmental adaptability of the microbial inoculant, and provide an efficient and safe new microbial inoculant for the green and sustainable development of agriculture. Summary of the Invention

[0005] The purpose of the present invention is to provide a preparation method and application of a microbial inoculant, which can well promote plant growth.

[0006] In order to achieve the above purpose, the technical solution adopted by the present invention is:

[0007] Select a strain of Bacillus velezensis and a strain of Saccharomyces from the self-owned strains in the laboratory.

[0008] Furthermore, a microbial inoculant is synthesized, and the raw materials are composed according to the following weight ratio: 70% - 80% of Bacillus velezensis and 20% - 30% of Saccharomyces.

[0009] Furthermore, the viable counts of the Bacillus velezensis and Saccharomyces are not less than 10 8 CFU / ml.

[0010] A preparation method of a microbial inoculant, comprising the following steps:

[0011] (1) Prepare each microbial raw material according to the described weight ratio: Bacillus velezensis and yeast liquid.

[0012] (2) Mix the mixed Bacillus velezensis and yeast, and stir well at 25 °C and 50 r / min for 24 h, then let it stand for 36 h to obtain a plant growth-promoting microbial inoculant.

[0013] An application of the described microbial inoculant in promoting plant growth.

[0014] The beneficial effects of the present invention are:

[0015] 1. The plant growth-promoting microbial inoculant provided by the present invention can increase the length and number of plant roots, and can also increase the number and weight of leaves, significantly promoting plant growth.

[0016] 2. The plant growth-promoting microbial inoculant provided by the present invention has the characteristics of being natural, pollution-free, having good and long-lasting fertilizer efficiency, etc. Description of the drawings

[0017] Figure 1 Growth diagram of Arabidopsis thaliana plants treated with the microbial inoculant in Example 1

[0018] Figure 2 Growth diagram of Arabidopsis thaliana plants without treatment with the microbial inoculant

[0019] Figure 3 Growth diagram of Arabidopsis thaliana plants treated with the microbial inoculant in Comparative Example 2 Detailed implementation manners

[0020] To more clearly explain the purpose, methods and advantages of the present invention, specific examples will be combined with the added drawings to further explain the present invention. The examples are only a part of the examples of the present invention, rather than all the examples.

[0021] In the process of implementing the present invention, it was found that the composition and production method of the current plant growth-promoting inoculants are relatively complex, and the overall growth-promoting effect on plants is insufficient. Therefore, preparing a simple and efficient plant growth-promoting microbial inoculant is of great significance for promoting plant growth.

[0022] The plant growth-promoting microbial inoculant provided by the present invention is composed of two plant growth-promoting bacteria. They are Bacillus velezensis and yeast, both of which are self-owned in the laboratory and have not been applied in the field of plant growth promotion.

[0023] Furthermore, the plant growth-promoting microbial inoculant is composed of raw materials in the following weight ratios: 70% - 80% of Bacillus velezensis and 20% - 30% of yeast.

[0024] Furthermore, the viable counts of both Bacillus velezensis and yeast are not less than 10 8 CFU / ml.

[0025] During implementation, Bacillus velezensis and yeast need to be respectively fermented and cultured in corresponding liquid media in advance, and the culture method adopts the conventional microbial culture method.

[0026] The composition of the used media is as follows:

[0027] The mass ratio of the fermentation medium for Bacillus velezensis is: acid hydrolysate casein 4.0 - 6.0 g, peptone 10.0 - 15.0 g, yeast powder 15.0 - 18.0 g, trisodium citrate 4.0 - 7.0 g, potassium chloride 5.0 - 7.0 g, anhydrous calcium chloride 1.0 - 1.5 g, ammonium sulfate: 1.0 - 2.0 g, magnesium sulfate heptahydrate 0.2 - 0.6 g, zinc sulfate heptahydrate 0.5 - 0.7 g, potassium dihydrogen phosphate 1.5 - 2.0 g, sodium chloride 5.0 - 6.0 g, add 1 L of pure water and mix evenly. PH 6.5 - 7.0.

[0028] The mass ratio of the fermentation medium for yeast is: potassium dihydrogen phosphate 1.0 - 1.2 g, sodium bicarbonate 2.0 - 2.5 g, magnesium sulfate heptahydrate 0.5 - 0.8 g, peptone 5.0 - 6.0 g, glucose 10.0 - 12.0 g, agar 20.0 g, NaCl 5.0 - 6.0 g, add 1 L of pure water and mix evenly, pH 6.5 - 7.5.

[0029] The growth medium ratio for Arabidopsis thaliana is: calcium nitrate 1.45 g, ammonium nitrate 1.0 g, magnesium sulfate 0.38 g, iron sulfate 0.004 g, boric acid 0.006 g, zinc sulfate 0.008 g, potassium phosphate 0.17 g, sodium molybdate 0.001 g, citric acid 1.0 g, sucrose 8.0 g, agar 2.8 g, sodium chloride 100 g, add 1 L of pure water and mix evenly, adjust the PH value to 5.7 - 6.0.

[0030] The preparation method of the microbial inoculant of the present invention includes the following steps:

[0031] (1) Prepare each microbial raw material according to the described weight ratios: 70% - 80% of Bacillus velezensis and 20% - 30% of yeast.

[0032] (2) Mix the prepared Bacillus velezensis and yeast, stir well at 25°C and 50 r / min for 24 hours, and then let it stand for 36 h to obtain the plant growth-promoting microbial inoculant.

[0033] The microbial inoculant prepared by the present invention can promote plant growth.

[0034] In the above-mentioned implementation, both strains are owned by the laboratory. The preservation number of Bacillus velezensis is CGMCC No. 21320; the number of the yeast is ATCC 204508.

[0035] Example 1 Preparation of microbial inoculant and plant growth promotion effect

[0036] Prepare the microbial inoculant according to the following steps:

[0037] (1) Prepare each microbial raw material according to the described weight ratio: 80% Bacillus velezensis and 20% yeast.

[0038] (2) Mix the prepared Bacillus velezensis and yeast, and stir well at 25 °C and 50 r / min for 24 hours, then let it stand for 36 h to obtain the plant growth-promoting microbial inoculant.

[0039] Conduct the plant growth promotion effect experiment of the inoculant according to the following steps:

[0040] Select Columbia wild-type Arabidopsis thaliana seeds. Prepare the Arabidopsis thaliana growth medium. Inoculate the pre-treated Arabidopsis thaliana seeds into the above medium, place them in a light incubator at 25 °C to grow, culture in the dark for 16 h, and then culture in the light for 8 h, and Arabidopsis thaliana can germinate;

[0041] Select plants with similar growth conditions from the above germinated Arabidopsis thaliana cultures and transfer them to a new solid medium, with 6 plants evenly distributed horizontally on each plate;

[0042] Inoculate the plant growth-promoting microbial inoculant composed of 80% Bacillus velezensis and 20% yeast by mass ratio onto the above plates, and continue to culture in a light incubator at 25 °C for 7 days. The effect is as Figure 1 shown.

[0043] Control 1

[0044] The Arabidopsis thaliana growth medium is the same as that in Example 2. Select the plate without inoculating the plant growth-promoting microbial inoculant as the blank control CK. The growth effect is as Figure 2 .

[0045] Control 2

[0046] The Arabidopsis thaliana growth medium is the same as that in Example 1. The inoculated microbial inoculant is changed to the following weight ratio: 50% Bacillus velezensis and 50% yeast. The growth result is as Figure 3 .

[0047] Figure 1It is the growth medium of Arabidopsis thaliana inoculated with plant growth-promoting microbial inoculant, compared with the blank control Figure 2 As shown in Table 1, the number of lateral roots increased significantly, the number of leaves increased, and the overall fresh weight increased significantly, indicating that the plant growth-promoting microbial inoculant has an obvious growth-promoting effect on plant growth. The different mass ratios of microbial inoculants used in Comparative Example 2 also showed a significant growth-promoting effect on plants, but the effect was not as good as the mass ratio used in Example 1. Table 1 shows the specific values of the number of lateral roots and fresh weight of Arabidopsis thaliana plants in each example and comparative example. Table 1.

Claims

1. A microbial inoculum, characterized in that: The raw material composition according to the following weight ratios: 70%-80% of Bacillus velezensis and 20%-30% of yeast.

2. The microbial inoculant according to claim 1, characterized in that, The viable counts of the Bacillus velezensis and yeast are not less than 10 8 CFU / mL.

3. The preparation method of the microbial inoculum according to claim 1, characterized in that, The method comprises the following steps: (1) Prepare each microbial raw material according to the weight ratios required in claim 1: Bacillus velezensis and yeast; (2) Mix the prepared Bacillus velezensis and yeast, and fully stir at 25°C and 50 r / min for 24 h, then let it stand for 36 h to obtain the microbial inoculant.

4. The growth-promoting effect of a microbial inoculant as claimed in claim 1 on plants.

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