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A kind of high-density fermentation method of Bacillus amyloliquefaciens and preparation method thereof

A technology of amylolytic spores and high-density fermentation, which is applied in the field of microbial fermentation, can solve the problems of low-growing and anti-amyloliquefaciens high-density fermentation optimization research, reducing the activity and quality stability of bacterial agents, and being unsuitable for popularization and use. The bacterial agent has a high rate of live bacteria and spores, improves the rate of spore formation, and has good control effect

Active Publication Date: 2019-12-10
湖南慧谷农业生态研究院有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The number of viable bacteria and the yield of spores are very key indicators to measure the Bacillus amyloliquefaciens inoculum. However, the research on the fermentation process of Bacillus amyloliquefaciens at home and abroad is mainly focused on the production of antimicrobial lipopeptides, amylase, protease and other enzymes. Few studies have been conducted on the optimization of high-density fermentation of Bacillus amyloliquefaciens, and most of the high-density fermentation process optimization only increases the number of viable bacteria, while the spore formation rate is low, and the low spore rate will reduce the activity and quality stability of the bacterial agent. Therefore, it is not suitable for popularization and use, and the production cost is high

Method used

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  • A kind of high-density fermentation method of Bacillus amyloliquefaciens and preparation method thereof
  • A kind of high-density fermentation method of Bacillus amyloliquefaciens and preparation method thereof
  • A kind of high-density fermentation method of Bacillus amyloliquefaciens and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Embodiment 1 1000L tank high-density fermentation produces raw control Bacillus amyloliquefaciens bacterial agent

[0044] (1) Strain activation: Streak inoculation of Bacillus amyloliquefaciens on a nutrient agar medium plate for culture, culture at 36°C for 24h, then pick a single colony and transfer it to the slant of nutrient agar, culture at 36°C for 24h Afterwards, the activated bacteria were obtained.

[0045] (2) Preparation of primary seed liquid in shake flasks: 500 mL of seed culture medium containing 5 g of yeast extract, 1 g of peptone, 1 g of glucose, and pH 7.0 ± 0.1 were loaded into two 2000 mL Erlenmeyer flasks respectively. The activated strains were inoculated into the shake flask seed medium with an inoculation loop, and the culture conditions were as follows: temperature 36°C, shaker speed 160r / min, culture time 12h.

[0046] (3) Preparation of the secondary seed solution in the seed tank: the seed solution obtained in step (2) is microscopically c...

Embodiment 2

[0051] Embodiment 2 5000L tank high-density fermentation bacillus amyloliquefaciens prepares its bacterial agent

[0052] (1) Strain activation: same as in Example 1.

[0053] (2) Preparation of primary seed solution in shake flask: same as in Example 1.

[0054] (3) Preparation of the secondary seed liquid in the seed tank: same as in Example 1.

[0055] (4) Fermentation tank high-density fermentation culture: the seed liquid gained in the step (3) is inserted in the 5000L fermentation tank by the inoculum size of 2% after microscopic inspection without assorted bacteria pollution, and the fermentation medium composition in the tank: bran 50000g, soybean meal 37500g, NaCl5000g, MgSO 4 2500g, CaCO 3 2500g, initial pH 6.5±0.1, liquid volume of fermentation tank 2500L, 0-24h culture conditions: temperature 30°C, stirring rate 160r / min, ventilation volume 100m 3 / h, tank pressure 0.06Mpa, 25-36h culture conditions: temperature 30°C, stirring rate 120r / min, ventilation volume ...

Embodiment 3

[0059] Embodiment 3 Utilizes quick-acting carbon and nitrogen source high-density fermentation to produce biocontrol Bacillus amyloliquefaciens bacterial agent

[0060] (1) Strain activation: same as in Example 1.

[0061] (2) Preparation of primary seed solution in shake flask: same as in Example 1.

[0062] (3) Preparation of the secondary seed liquid in the seed tank: same as in Example 1.

[0063] (4) Fermentation tank high-density fermentation culture: the seed liquid gained in the step (3) is inserted in the 5000L fermentation tank by the inoculum size of 2% after microscopic examination without assorted bacteria pollution, and the fermentation medium composition in the tank: glucose 50000g , yeast extract 25000g, NaCl 2500g, MgSO 4 5000g, CaCO 3 2500g, initial pH 6.5±0.1, liquid volume of fermentation tank 2500L, 0-24h culture conditions: temperature 30°C, stirring rate 160r / min, ventilation volume 100m 3 / h, tank pressure 0.06Mpa, 25-36h culture conditions: tempera...

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Abstract

The invention provides a high-density fermentation method of Bacillus amyloliquefaciens and a method for preparing bacterial agent thereof. High-density fermentation medium containing bran or glucose, soybean meal or yeast extract, NaCl, MgSO 4 , CaCO 3 , initial pH 6.0-7.0, bran and soybean meal can be replaced by glucose and yeast extract respectively. Under the condition of 1000L fermentation tank scale, the effective number of viable bacteria reaches 8.3×10 9 More than cfu / mL, the number of spores reaches 7.6×10 9 More than cfu / mL, sporulation rate is more than 91.6%. The preparation method of the bacterial agent comprises centrifugally collecting the bacterial slime and spray-drying to obtain a dry powder preparation, and the number of effective viable bacteria in the dry powder preparation is ≥9.0×10 10 cfu / g, shelf life at room temperature up to 18 months. The fermentation process of the invention is simple, the raw materials are cheap and easy to obtain, and the cost is low, which is suitable for large-scale production in factories.

Description

technical field [0001] The invention relates to the technical field of microbial fermentation, in particular to a method for high-density fermentation of Bacillus amyloliquefaciens with biocontrol function and a method for preparing dry powder bacterial agent. Background technique [0002] Plant diseases not only reduce the yield and quality of crops and cause direct economic losses, but also produce toxic substances, which pose a threat to human and animal health and seriously restrict the sustainable development of agriculture. At present, plant disease control is mainly based on disease-resistant varieties and chemical control. However, the selection cycle of disease-resistant varieties is long and difficult, and chemical control is easy to cause environmental and food pollution. With the increasing requirements of human beings on the environment and food safety and quality The higher the value, the less popular chemical control becomes. Green, pollution-free and harmles...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12R1/07
CPCC12N1/20
Inventor 郑双凤谭石勇谭武贵丰来罗志威徐滔明张冬雪
Owner 湖南慧谷农业生态研究院有限公司