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Method for producing biocontrol bacillus amyloliquefaciens agent by liquid-solid biphasic fermentation

A technology of Bacillus amyloliquefaciens and Bacillus amyloliquefaciens, which is applied in the field of liquid-solid two-phase fermentation to produce bio-control Bacillus amyloliquefaciens inoculum, which can solve the problem of bio-control Bacillus amyloliquefaciens inoculum, which is difficult to obtain high-density live bacteria and Problems such as spores and metabolites cannot be fully utilized, so as to achieve the effects of easily expanding production, improving the level of comprehensive utilization, and accelerating growth and reproduction

Inactive Publication Date: 2018-11-13
湖南慧谷农业生态研究院有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, liquid submerged fermentation has high production costs, difficulty in obtaining high-density viable bacteria and spores, inability to fully utilize metabolites, complicated follow-up treatment (such as vacuum concentration of fermentation broth, adsorption of solid powder carrier, drying, etc.), and waste liquid discharge However, pure solid fermentation is not suitable for promotion and use due to the multi-stage solid expansion that increases the risk of bacterial contamination and the process is complex and the fermentation cycle is long.
How to increase the spore rate as much as possible while ensuring the effective number of viable bacteria while taking into account the production cost has become a bottleneck problem that plagues the actual industrial production of biocontrol Bacillus amyloliquefaciens inoculum

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1 liquid-solid two-phase fermentation produces the method for biocontrol Bacillus amyloliquefaciens bacterial agent

[0035] Including the following steps:

[0036] (1) Strain activation: pick a small amount of bacterial lawn from the Bacillus amyloliquefaciens strains preserved on the NA slope, use an inoculation loop, separate them by marking on the NA plate, and culture them in a constant temperature incubator at 30°C for 24 hours;

[0037] (2) Shake flask seed culture: pick a ring of activated bacterial lawn with an inoculation loop, inoculate it in YPD seed medium (1.2L in total) for cultivation, shaker culture conditions: temperature 36°C, shaker speed 160r· min-1, culture time 12h, get shake flask seed solution;

[0038] (3) fermentor submerged liquid fermentation culture: the shake flask seed liquid gained in the step (2) is inserted in the 100L fermentor by volume ratio 2% inoculum, the fermentation medium composition in the tank: bran 10g / L, Soybe...

Embodiment 2

[0043] Embodiment 2 liquid-solid two-phase fermentation produces the method for biocontrol Bacillus amyloliquefaciens bacterial agent

[0044] Including the following steps:

[0045] (1) Strain activation: pick a small amount of bacterial lawn from the Bacillus amyloliquefaciens strains preserved on the NA slope, use an inoculation loop, separate them by marking on the NA plate, and culture them in a constant temperature incubator at 30°C for 24 hours;

[0046] (2) Shake flask seed culture: Pick a ring of activated bacterial lawn with an inoculation loop, inoculate it in YPD seed medium (0.5L in total) for cultivation, shaker culture conditions: temperature 36°C, shaker speed 160r. min -1 1. Cultivation time 12h, shake flask seed solution;

[0047] (3) Fermentation tank submerged liquid fermentation culture: the shake flask seed liquid gained in step (2) is inserted in the 100L fermentation tank by volume ratio 1% inoculum, and the fermentation medium composition in the tank...

Embodiment 3

[0053] Embodiment 3 liquid-solid two-phase fermentation produces the method for biocontrol bacillus amyloliquefaciens

[0054] Including the following steps:

[0055] (1) Strain activation: pick a small amount of bacterial lawn from the Bacillus amyloliquefaciens strains preserved on the NA slope, use an inoculation loop, separate them by marking on the NA plate, and culture them in a constant temperature incubator at 30°C for 24 hours;

[0056] (2) Shake flask seed culture: pick a loop of activated bacterial lawn with an inoculation loop, inoculate it in YPD seed medium (1.8L in total) for cultivation, shaker culture conditions: temperature 36°C, shaker speed 160r· min -1 12 hours of incubation time, the shake flask seed solution was obtained;

[0057] (3) Fermentation tank submerged liquid fermentation culture: the shake flask seed liquid gained in step (2) is inserted in the 100L fermentation tank by volume ratio 3% inoculum, and the fermentation medium composition in the...

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Abstract

The invention provides a method for producing a biocontrol bacillus amyloliquefaciens agent by liquid-solid biphasic fermentation, wherein the method comprises the following steps: bacterial strain activation, shaking flask seed culture, deep liquid fermentation culture with a fermentation tank, solid fermentation culture with a porous shallow dish and fermentation post-treatment. The liquid fermentation culture medium comprises the components: bran, soybean powder, NaCl, MgSO4.7H2O and CaCO3, and the solid fermentation culture medium mainly contains bran, corncob powder, soybean meal, sucroseor molasses, and rice hulls. The number of effective living bacteria in the obtained bacterial agent product is more than or equal to 5.0*10<10> cfu / g, and the spore rate is 95% or more. The method has the advantages of simple process, low production cost, easy control and management, high yield of living bacteria and spores, and simple post-treatment, and is beneficial to application and promotion of the biocontrol microbial agent.

Description

technical field [0001] The invention relates to the technical field of microbial fermentation, in particular to a method for producing a bacterial agent against Bacillus amyloliquefaciens by liquid-solid two-phase fermentation. Background technique [0002] Bacillus amyloliquefaciens is a common biocontrol bacillus closely related to Bacillus subtilis. Studies have shown that most of Bacillus amyloliquefaciens can produce broad-spectrum antibacterial active substances, compete with pathogenic bacteria for nutrients, induce plant systemic resistance, and promote plant growth. They also have fast growth, simple nutrition, and can produce heat-resistant Stress-resistant spores, strong colonization and reproduction capabilities, harmless to humans and animals, and no environmental pollution. At present, the biocontrol of Bacillus amyloliquefaciens has a remarkable effect on the prevention and control of soil-borne diseases of pepper, potato, cucumber, rice, rape, lettuce and ot...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A01N63/00A01P3/00C12R1/07
CPCA01N63/00C12N1/20
Inventor 郑双凤谭石勇谭武贵罗志威郭帅聂勇丰来
Owner 湖南慧谷农业生态研究院有限公司
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