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Methyl nutritional type bacillus spore-production fermentation medium and culture method thereof

A methylotrophic and fermentation medium technology is applied in the field of methylotrophic bacillus spore-producing fermentation medium and its culture field, which can solve the problems of long shelf life, difficulty in producing spores, influence and the like, and achieves a low production cost. Effect

Inactive Publication Date: 2015-11-04
SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are few studies on the fermentation process of methylotrophic bacillus with spores as the final product, and it is difficult to produce spores under conventional culture conditions, which affects its becoming a spore preparation with high efficiency, low toxicity, low cost and long shelf life. Therefore There is an urgent need to optimize the fermentation formula of methylotrophic Bacillus 9912, find out the best medium formula and fermentation conditions, in order to increase the content of spores in the fermentation broth, reduce production costs, and lay the foundation for large-scale industrial production of wettable powder

Method used

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  • Methyl nutritional type bacillus spore-production fermentation medium and culture method thereof
  • Methyl nutritional type bacillus spore-production fermentation medium and culture method thereof
  • Methyl nutritional type bacillus spore-production fermentation medium and culture method thereof

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Effect test

Embodiment 1

[0029] Activation of the strain

[0030] Methylotrophic Bacillus 9912-D was streaked and inoculated on the modified starch agar medium, cultured at 30°C for 48 hours, picked and observed under a microscope, the typical single colony was streaked and purified again, and then inoculated on the slant of the modified starch test tube, cultured at 30°C 48h standby. The typical colony morphology on the modified starch agar medium is irregular at the edge, the surface is dry and slightly wrinkled, the colony is thicker and creamy in color; the bacterial morphology under the microscope: the cell is rod-shaped or columnar, and the size is 0.6-0.8×1.5- 3.0 μm, spores columnar or elliptic, not enlarged, with one flagella, terminal or lateral.

[0031] The composition of the modified starch medium is: 2% soluble starch, 0.5% beef extract, 0.5% yeast extract, 0.5% NaCl, and 1.8% agar. Natural pH, sterilized by high pressure steam at 115°C for 30 minutes (or sterilized at 121°C for 20 min...

Embodiment 2

[0033] Experimental Design and Statistical Analysis

[0034] 2.1 Single factor experimental design to screen suitable carbon and nitrogen sources

[0035] Carbon sources can not only provide energy for the growth and reproduction of microorganisms, but also are necessary components for the synthesis of bacteria. The enzyme systems of microorganisms are different, and the metabolism of different carbon sources has different effects on the quality of fermentation products. Nitrogen source is the source of cellular protein and nucleic acid substances, and has an important relationship with the normal metabolism of bacteria. It has been reported that some agricultural and sideline products, such as corn steep liquor, cornmeal, soybean cake powder, etc., can provide coenzymes and amino acids necessary for spore formation in the later stage of fermentation. Select 6 different carbon sources in the fermentation medium, namely cornmeal, bran, glucose, corn starch, sucrose, dextrin, a...

Embodiment 3

[0065] Single-factor experimental design to investigate the effects of pH, inoculum size, seed age, and liquid volume on spore production

[0066] The optimal cheap fermentation medium optimized above was used as the basic medium, and the influence of various factors on the spore yield and transformation rate was investigated by changing the inoculum size, liquid loading volume, initial pH, and seed age. The results showed that: the inoculum size 5 ~10%, liquid volume 500ml, shake bottle 50ml fermentation medium, initial pH 7.0~7.5, seed age 15~18h, fermentation 20~36h, aseptic sampling and determination of active spores up to 2×10 9 cfu / ml, the conversion rate is above 95%, and the predicted value of the model is 1.86×10 9 The cfu / ml should be higher.

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Abstract

The invention relates to optimization of a methyl nutritional type bacillus spore-production fermentation medium. The fermentation medium consists of the following components: 0.5-0.6% of corn meal, 0.08-0.2% of corn starch, 0.9-1.2% of bran, 0.18-0.54% of glucose, 0.05-0.15% of soybean cake powder, 0.16-0.2% of soybean peptone, 0.005-0.006% of KH2PO4, 0.006-0.008% of (NH4)2SO4, 0.004-0.005% of MnSO4, 0.0005-0.002% of CaCO3, 0.005-0.006% of MgSO4.7H2O, and the balance of water. The culture medium provided by the invention not only can meet the requirement of a shake flask fermentation experiment but also can successfully amplify the result of shake flask fermentation to a fermentation tank. Compared with the culture medium before optimization, the spore conversion rate and the spore yield of the formula of the culture medium are improved by 1-10 times, and the formula of the culture medium adopts low-price agriculture byproducts, so that the production cost is greatly reduced, and a foundation is laid for large-scale industrial production of wettable powder.

Description

technical field [0001] The present invention relates to a technique for optimizing the high-density fermentation production process of spores of methylotrophic bacillus by applying response surface method and single factor experiment design, and obtains a fermentation medium suitable for producing spores of methylotrophic bacillus and its cultivation method. Background technique [0002] Chemical pesticides have made great contributions to the prevention and control of plant pathogenic fungal diseases. However, problems such as the environment caused by chemical pesticides are becoming more and more serious. Therefore, biological control has been put on the agenda. In order to solve this problem, it is necessary to tap new microbial resources and develop new biological pesticides. [0003] Bacillus is widely distributed in nature and can produce a variety of antibacterial active substances. At present, it has been widely used in various industries such as medicine, pestic...

Claims

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Application Information

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IPC IPC(8): C12N3/00C12R1/07
Inventor 冯媛媛胡江春李学红潘华奇李春田王楠
Owner SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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