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Bacillus subtilis and preparation and application of microbial inoculum thereof

The technology of Bacillus subtilis and microbial inoculum is applied in the field of agricultural microorganisms, which can solve the problems of host infection and harm, and achieve the effects of short fermentation period, good safety and excellent fermentation properties.

Active Publication Date: 2013-03-13
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At the same time, most soil pathogenic bacteria will parasitize in the residue or soil to form durable living organs in the absence of hosts. Once the hosts appear, they will re-infect the hosts and cause harm.

Method used

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  • Bacillus subtilis and preparation and application of microbial inoculum thereof
  • Bacillus subtilis and preparation and application of microbial inoculum thereof
  • Bacillus subtilis and preparation and application of microbial inoculum thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. Isolation of Y-1 strain

[0031] The Y-1 strain was obtained through a series of experiments such as the isolation and screening of various bacteria in the apple trees in Tai'an. The specific separation method is as follows: Rinse the collected apple tree samples with sterile water, then soak them in 75% alcohol and 5% sodium hypochlorite for 3 minutes, then rinse with sterile water 3-5 times; then grind the samples, Add 90ml of sterile water, place it in a triangular flask and culture with shaking at 180r / m for 2h at 25°C; dilute the culture solution gradually and spread it on the LB plate. Each treatment was repeated 3 times, cultured at 28°C for 3-5 days. Single colonies of different morphologies were selected for repeated plate streaking and purification, and then a confrontation test was performed on Fusarium oxysporum and the purified single colony. After repeated tests, one was obtained. This strain has a significant inhibitory effect on Fusarium oxysporum, name...

Embodiment 2

[0047] (1) Screening of Bacillus subtilis Y-1 double antibiotic mutant strains

[0048] Firstly, the Y-1 strain was inoculated on an LB plate with a rifampicin concentration of 0.5μg / mL. After the colonies were seen to grow significantly with the naked eye, they were transferred to the LB plate with a rifampicin concentration of 1μg / mL. After obvious growth, switch to LB with rifampicin concentrations of 2μg / mL, 4μg / mL, 8μg / mL, 16μg / mL, 32μg / mL, 64μg / mL, 128μg / mL, 256μg / mL, 300μg / mL On the tablet. A Y-1 mutant strain with rifampicin resistance was obtained.

[0049] The Y-1 mutant strain with 300 μg / mL resistance to kanamycin was selected on the LB plate containing 300 μg / mL rifampicin in the same way as above.

[0050] (2) Colonization of Bacillus subtilis Y-1

[0051] The Y-1 double antibiotic mutant strain obtained in the above experiment was inoculated into LB liquid medium containing 300μg / mL rifampicin and kanamycin, and cultured at 30°C and 200r / m shaking for 24h. The test gr...

Embodiment 3

[0057] Select glucose as the quick-acting carbon source, corn flour and soybean meal as the slow-acting carbon and nitrogen sources, and three inorganic salts of ammonium sulfate, potassium dihydrogen phosphate and calcium carbonate. The orthogonal experiment design is carried out. The experimental factors and levels are shown in Table 4. , The formula of each medium is shown in Table 5 to optimize the best spore-producing fermentation medium of Bacillus subtilis Y-1.

[0058] Table 4 Test factors and level settings

[0059]

[0060] Table 5 Formulas of 18 fermentation media

[0061]

[0062]

[0063] Use an inoculating loop to pick out the Bacillus subtilis Y-1 colony and inoculate it into a 250ml Erlenmeyer flask containing 50ml LB medium. After culturing at 30℃ and 200r / m for 10 hours, the seed solution is obtained, and then the seed solution is 2% inoculum Inoculated into the above-mentioned 18 kinds of culture medium, 30℃, 200r / m shaker shaking culture. After fermentation for ...

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Abstract

The invention provides bacillus subtilis and preparation and application of a microbial inoculum thereof. The invention mainly relates to a Bacillus subtilis Y-1 of which the preservation number is CGMCC No.6539, and also relates to the microbial inoculum which is prepared from the bacillus subtilis, and application of the microbial inoculum. The Y-1 strain can be colonized on an apple tree, a polar tree, a cucumber and the rhizosphere of capsicums and in an apple tree, and has efficient and broad-spectrum antibacterial activity on fusarium, rhizoctonia solani, pestallozzia theae, phytophthora and the like; the microbial inoculum of the bacillus subtilis can be used for controlling diseases and continuous cropping obstacles of an apple tree, a polar tree, a cucumber and capsicums caused by those disease fungi. The preparation method of the microbial inoculum is simple in process, short in fermentation cycle, low in cost, and favorable for industrial production.

Description

Technical field [0001] The invention relates to the preparation and application of a Bacillus subtilis strain and its bacterial agent, and belongs to the field of agricultural microorganisms. Background technique [0002] Chemical pesticides play a major role in preventing or reducing the damage caused by plant diseases and insect pests and protecting agricultural production. However, chemical pesticides are highly toxic, stable in structure, and difficult to decompose. After a large amount of use, they remain in crops, are enriched after animals eat, and can penetrate into soil, rivers or groundwater, causing environmental pollution and affecting human health. Moreover, due to the long-term use of certain chemical pesticides, resistance to pests and bacteria has been caused. In order to control chemical pollution and protect the environment, countries in the world are currently developing ecological agriculture, reducing the use of chemical pesticides, and researching and devel...

Claims

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Application Information

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IPC IPC(8): C12N1/20A01N63/00A01N63/02A01P3/00C12R1/125
Inventor 刘训理鞠瑞成毛志泉鲍正宗李金玉赵玉华杨涛姜红霞
Owner SHANDONG AGRICULTURAL UNIVERSITY
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