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A high-efficiency chitosanase-producing bacterium and its fermentation method

A chitosan enzyme and bacteria-producing technology, applied in the biological field, can solve the problems of low yield of oligosaccharides, difficult product separation, environmental pollution, etc.

Active Publication Date: 2019-12-27
领先生物农业股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the methods for preparing chitosan oligosaccharides mainly include chemical degradation method and enzymatic hydrolysis method. The yield of degraded oligosaccharides by chemical method is low, the separation of products is difficult, and it causes serious environmental pollution.

Method used

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  • A high-efficiency chitosanase-producing bacterium and its fermentation method
  • A high-efficiency chitosanase-producing bacterium and its fermentation method
  • A high-efficiency chitosanase-producing bacterium and its fermentation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] ——Fermentation culture of GS115-LX strain (100L tank)

[0069] Medium composition:

[0070] 1) Solid medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0071] 2) Shake flask seed medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0072] 3) Basal salt medium: 10ml of 85% phosphoric acid, 3g of calcium sulfate, 3g of potassium sulfate, 5g of magnesium sulfate heptahydrate, 20g of glycerol, add water to a total volume of 1L, and sterilize at 121°C for 30min;

[0073] 4) Feed medium: methanol.

[0074] In the following experiments, the formula of the medium was not changed, and this medium was used.

[0075] First, culture on a slant, inoculate the glycerol tube strain preserved by the GS115-LX strain on the slant of the test tube slant solid medium, and then place it in a constant temperature incubator and culture it on a slan...

Embodiment 2

[0080] ——Fermentation culture of GS115-LX strain (100L tank)

[0081] Medium composition:

[0082] 1) Solid medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0083] 2) Shake flask seed medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0084] 3) Basal salt medium: 10ml of 85% phosphoric acid, 3g of calcium sulfate, 3g of potassium sulfate, 5g of magnesium sulfate heptahydrate, 20g of glycerol, add water to a total volume of 1L, and sterilize at 121°C for 30min;

[0085] 4) Feed medium: methanol.

[0086] In the following experiments, the formula of the medium was not changed, and this medium was used.

[0087] First, culture on a slant, inoculate the glycerol tube strain preserved by the GS115-LX strain on the slant of the test tube slant solid medium, and then place it in a constant temperature incubator and culture it on a slan...

Embodiment 3

[0092] ——Fermentation culture of GS115-LX strain (500L tank)

[0093] Medium composition:

[0094] 1) Solid medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0095] 2) Shake flask seed medium: yeast extract powder 1%, peptone 1%, glucose 2%, agar 1.8%, add water to 100%, sterilize at 115°C for 20 minutes;

[0096] 3) Basal salt medium: 10ml of 85% phosphoric acid, 3g of calcium sulfate, 3g of potassium sulfate, 5g of magnesium sulfate heptahydrate, 20g of glycerol, add water to a total volume of 1L, and sterilize at 121°C for 30min;

[0097] 4) Feed medium: methanol.

[0098] First, culture on a slant, inoculate the glycerol tube strain preserved by the GS115-LX strain on the slant of the test tube slant solid medium, and then place it in a constant temperature incubator and culture it on a slant at a temperature of 28-30°C for 2-3 days to obtain Inclined bacteria;

[0099] Then, for primary seed cul...

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Abstract

The invention relates to a high-efficiency chitosanase-producing bacterium and a fermentation method thereof. The present invention has obtained a genetically engineered bacterium Pichia pastoris (Pichia pastoris) GS115-LX bacterial strain producing chitosanase by genetic engineering cloning-construction-recombination, fermented by the bacterial strain according to the fermentation culture method of the present invention The chitosan enzyme activity of the fermented broth obtained by cultivating can reach 500-1000U / ml, the enzyme activity is very high, no need of purification, and can be directly used for the degradation of chitosan. It has broad application prospects in the field of industrial production of chitosan oligosaccharides.

Description

technical field [0001] The invention relates to a high-efficiency chitosanase-producing bacterium and a fermentation method thereof. Belongs to the field of biotechnology. Background technique [0002] Chitosan is a linear polysaccharide composed of β-(1,4)-D-glucosamine. In nature, this polymer is partially acetylated. In fact, chitosan is used broadly to describe polymers composed of D-glucosamine and N-acetyl-D-glucosamine residues. [0003] The molecular weight of chitosan macromolecules is usually around hundreds of thousands. Because of the interaction of internal and external hydrogen bonds in the molecule, it can only be dissolved in a small number of dilute acid solutions, but not in water. Oligochitosan is an oligosaccharide formed by using chitosan as a raw material and degrading and preparing 2 to 10 glucosamines connected by β-(1,4) glycosidic bonds through bioengineering technology. Due to its small molecular weight, good water solubility, easy absorption, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/19C12N9/42C12R1/84
Inventor 苏海榆孙杉杉徐志文
Owner 领先生物农业股份有限公司