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Bacillus sp. A-5 and application thereof

A technology of Bacillus subtilis and strains, applied in the field of bioengineering, can solve the problems of low production efficiency of γ-polyglutamic acid

Active Publication Date: 2019-10-25
SHANGHAI ACAD OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In view of the above-mentioned shortcoming of the prior art, the object of the present invention is to provide a kind of Bacillus subtilis sp.A-5 and its application, for solving the problem of low production efficiency of gamma-polyglutamic acid in the prior art

Method used

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  • Bacillus sp. A-5 and application thereof
  • Bacillus sp. A-5 and application thereof
  • Bacillus sp. A-5 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] A strain of synthetic γ-polyglutamic acid is screened from home-made natto and named Bacillus subtilis sp.A-5. The strain has been registered and preserved in the China Center for Type Culture Collection (CCTCC) on March 15, 2019, and the preservation number is CCTCC NO: M2019157.

[0042] The Bacillus strain of the present invention was sent to the Extremophile Laboratory of Zhejiang University for morphological identification in December 2018, and the identification results were:

[0043] The morphological characteristics are: cultured in an incubator at 37°C overnight, the colony is irregularly round, milky white, with irregular edges and opaque; Gram staining is positive; under the microscope, it is rod-shaped and can form spores; the size is (1.20~3.00 )μm×(0.45~1.40)μm, with blunt round ends. Such as figure 1 shown.

[0044] The main physiological characteristics of the A-5 strain are: alkaline phosphatase, esterase (C4), esterase lipase (C8), acid phosphatase,...

Embodiment 2

[0051] (1) The γ-polyglutamic acid solution can react with the sodium hydroxide solution of the cationic surfactant cetyltrimethylammonium bromide (CTAB) at a suitable concentration to form a suspension.

[0052] The principle is that in solution, the nitrogen atom of CTAB pairs with the oxygen atom of the carboxyl group in γ-polyglutamic acid to form a water-insoluble γ-polyglutamic acid-CTAB complex. In this experiment, the absorbance of the suspension formed by γ-polyglutamic acid and CTAB at 250nm is directly proportional to the concentration of γ-polyglutamic acid, and has a good linear relationship. First prepare a γ-polyglutamic acid standard solution with a certain concentration gradient, mix it with an equal volume of CTAB sodium hydroxide solution, let it stand at room temperature for 3-4min, measure its absorbance at 250nm, and draw a standard curve as y =0.0166x-0.134(R 2 = 0.9996).

[0053] Table 1 Absorbance of different concentrations of γ-polyglutamic acid at...

Embodiment 3

[0059] The method for efficiently preparing gamma-polyglutamic acid by using the gamma-polyglutamic acid synthetic strain in Example 1, the steps are as follows:

[0060](1) Slant surface activation: Bacillus sp.A-5 was streak-inoculated on the slant medium with an inoculation loop, and cultured overnight at 37°C. The components of the slant medium are: beef extract 3.0g / L, peptone 10g / L, sodium chloride 5.0g / L, pH 7.2.

[0061] (2) Seed culture: Pick a single colony from the slant culture medium and inoculate it into the seed culture medium containing an appropriate amount of glutamic acid, cultivate overnight at 200 rpm at 37° C. until the mid-late logarithmic period. The components of the seed medium are: glucose 10g / L, yeast extract 2g / L, sodium glutamate 10g / L, pH value 7.2, sterilized at 115°C for 15min.

[0062] (3) Fermentation culture: insert the seed liquid obtained in step (2) into the fermentation medium, the inoculum size is 5%, the liquid volume of the shaker fl...

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Abstract

The invention provides bacillus sp. A-5 and application thereof. The preservation number of the bacillus sp. A-5 is CCTCC NO: M2019157. The conversion rate of gamma-polyglutamic acid prepared from bacillus sp. A-5 by fermenting exceeds 75%, the molecular weight exceeds 4000 KDa, and the conversion rate is far higher than the conversion rate in the prior art.

Description

technical field [0001] The invention relates to a bacillus subtilis sp.A-5 and an application thereof, belonging to the field of bioengineering. Background technique [0002] γ-polyglutamic acid is a kind of anionic polypeptide polymer formed by connecting several glutamic acid monomers (D-type or L-type) through γ-amide bonds. It can be synthesized by chemical methods or microbial fermentation. Mostly between 100-1000kDa, its average molecular weight is about 1.23×10 4 KDa. γ-polyglutamic acid does not have a typical peptide chain structure, and the basic skeleton is a straight chain fiber. γ-polyglutamic acid, as a new type of environmentally friendly green polymer bioproduct, has unique physical and chemical properties such as degradability, film formation, fiber formation, plasticity, and adhesion, and can be used in pharmaceutical manufacturing , food processing and preservation, cosmetics, manufacturing, agriculture and many other fields. [0003] The advantages of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P13/02C12R1/125
CPCC12P13/02C12R2001/125C12N1/205
Inventor 白娜玲吕卫光张翰林郑宪清李双喜张娟琴张海韵
Owner SHANGHAI ACAD OF AGRI SCI
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