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Bacillus subtilis and preparation and application of microbial inoculum thereof

A technology of Bacillus subtilis and microbial inoculum, which is applied in the field of agricultural microorganisms, can solve the problems of host infection and harm, and achieves the effects of short fermentation period, good safety and favorable industrial production.

Active Publication Date: 2014-03-05
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 tests such as the isolation and screening of various bacteria from apple trees in Tai'an City. The specific separation method is: rinse the collected apple tree samples with sterile water, then soak them in 75% alcohol and 5% sodium hypochlorite for 3 minutes, then rinse them with sterile water for 3-5 times; then grind the samples, Add 90ml of sterile water, place in a Erlenmeyer flask and culture at 180r / m for 2h at 25°C; carry out gradient dilution of the culture solution, 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 forms were selected for repeated plate streaking and purification, and then the confrontation test was carried out between Fusarium oxysporum and the purified single colonies. After repeated experiments, a single colony was obtained. The strain has a significant inhibitory effect on Fusari...

Embodiment 2

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

[0048] First, inoculate the Y-1 strain on LB plates with a rifampicin concentration of 0.5 μg / mL. After the colonies were observed to grow significantly with the naked eye, they were transferred to LB plates with a rifampicin concentration of 1 μg / mL. After obvious growth, transfer into LB with rifampicin concentration of 2 μg / mL, 4 μg / mL, 8 μg / mL, 16 μg / mL, 32 μg / mL, 64 μg / mL, 128 μg / mL, 256 μg / mL, 300 μg / mL on the tablet. A Y-1 mutant strain with rifampicin resistance was obtained.

[0049] On the LB plate containing rifampicin 300 μg / mL, the Y-1 mutant strain resistant to kanamycin 300 μg / mL was screened by the same method as above.

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

[0051] Inoculate the Y-1 double-antibiotic mutant strain obtained in the above experiment into LB liquid medium containing rifampin and kanamycin 300 μg / mL, culture at 30°C, 200 r / m for 24 hours, and take 20...

Embodiment 3

[0057] Glucose was selected as the quick-acting carbon source, corn flour and soybean cake powder were used as the slow-acting carbon and nitrogen source, and three inorganic salts, ammonium sulfate, potassium dihydrogen phosphate, and calcium carbonate, were used to conduct an orthogonal experimental design. The settings of the experimental factors and levels are shown in Table 4 , the formula of each medium is shown in Table 5, and the best spore-forming fermentation medium of Bacillus subtilis Y-1 is optimized.

[0058] Table 4 Experimental factors and level settings

[0059]

[0060] Table 5 Formulas of 18 kinds of fermentation media

[0061]

[0062]

[0063] Pick the colony of Bacillus subtilis Y-1 with an inoculation loop, inoculate it into a 250ml Erlenmeyer flask containing 50ml of LB medium, cultivate it at 30°C and 200r / m for 10h to obtain its seed liquid, and then inoculate the seed liquid with a 2% inoculation amount Inoculate into the above-mentioned 18...

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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 strain of Bacillus subtilis and its bacterial agent, belonging to the field of agricultural microorganisms. Background technique [0002] Chemical pesticides have played an important role in preventing or mitigating the harm caused by pests and diseases and protecting agricultural production. However, chemical pesticides have high toxicity, stable structure, and are difficult to degrade. After being used in large quantities, they will remain in crops, enrich after animals eat, and will penetrate into soil, rivers or groundwater, causing environmental pollution and affecting human health. Moreover, due to the long-term use of some chemical pesticides, the resistance of pests and bacteria has been caused. In order to control chemical pollution and protect the environment, countries all over the world regard the development of ecological agriculture, reducing the use of chemical pesticides, re...

Claims

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Application Information

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Patent Type & Authority Patents(China)
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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