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Bacillus subtilis strain and applications thereof

A technology of Bacillus subtilis and strains, applied in the direction of bacteria, biochemical equipment and methods, microorganisms, etc., can solve the problem of not being able to use glycerin, and achieve the effect of cheap raw materials, high output, and short fermentation cycle

Active Publication Date: 2014-06-18
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] In view of the defect that existing Bacillus subtilis strains cannot use glycerol, especially crude glycerol, a by-product of biodiesel, to produce lipopeptide biosurfactin surfactin, therefore, the present invention separates and purifies a strain that metabolizes lipopeptide from the polluted soil of Shengli Oilfield The Bacillus subtilis-like biosurfactant was screened by atmospheric room temperature plasma (ARTP) mutagenesis to obtain a Bacillus subtilis strain that can use glycerol as the only carbon source to metabolize the lipopeptide biosurfactant surfactin

Method used

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  • Bacillus subtilis strain and applications thereof
  • Bacillus subtilis strain and applications thereof
  • Bacillus subtilis strain and applications thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0035] The present embodiment illustrates obtaining Bacillus subtilis G-34 ( Bacillus subtilis G-34) approach.

[0036] Include the following steps:

[0037] 1) Obtain biosurfactant strains in oil-contaminated soil by enriching the medium.

[0038] The oil-contaminated soil samples from Shengli Oilfield were taken, and the treated soil samples were inoculated into the enrichment medium to obtain the enrichment medium. Utilizing the hemolytic properties of biological surface active agents, the enriched culture solution was inoculated on the blood plate by streaking and coating separation methods, and cultured at 37°C for 24-48 hours to obtain a single colony with a large and transparent hemolytic zone.

[0039] The enrichment medium formula is: glucose 20 g / L, ammonium sulfate 1 g / L, sodium nitrate 2 g / L, magnesium sulfate 0.3 g / L, potassium dihydrogen phosphate 1 g / L, disodium hydrogen phosphate 4 g / L L, yeast powder 2 g / L.

[0040] The blood plate medium formula is: beef...

Embodiment 2

[0051] This example illustrates the effect on Bacillus subtilis strains Bacillus subtilis The results of preliminary identification of the type of biosurfactant produced by G-34.

[0052] by Bacillus subtilis The G-34 strain was used as a strain, and glycerol was used as a carbon source for shake flask fermentation. Bacillus subtilis The biosurfactant produced by G-34 was purified and analyzed by FT-IR.

[0053] The results of the analysis are attached figure 1 . In the IR spectrum, 3303cm-1 is the NH stretching vibration caused by hydrogen bonds between molecular chains, and 1656cm-1 and 1550cm-1 are phthalamide bands I and II. The hydrophilic group of the absorbing surface active agent molecule of the above characteristics is - the peptide chain. The two absorptions in the spectrogram 2872-2960cm-1 and 1243-1402cm-1 are the C-H stretching vibration table of the fatty acid family. The hydrophobic part of the surfactant molecule is a fatty half molecule, indicating th...

Embodiment 3

[0055] This example demonstrates that Bacillus subtilis Bacillus subtilis G-34 utilizes pure glycerol to ferment and produce the step of lipopeptide biosurfactant.

[0056] (1) Preparation of seed medium: yeast extract powder 5 g / L, peptone 10 g / L, NaCl 10 g / L. Sterilize at 121°C for 20 min and cool down for use.

[0057] (2) Fermentation medium preparation: glycerol 20 g / L, NH 4 NO 3 2 g / L, KH 2 PO 4 3 g / L, Na 2 HPO 4 10 g / L, MgSO 4 ·7H 2 O 0.2 g / L, FeSO 4 ·7H 2 O 0.02 g / L.

[0058] The medium was adjusted to pH 7.0 with phosphoric acid (1mol / L), sterilized at 121°C for 20min, and cooled for later use.

[0059] (3) pick a ring Bacillus subtilis The G-34 strain was placed in the culture medium obtained in step (1); placed at 37°C and 200 rpm for 12 hours to obtain seed liquid.

[0060] (4) Inoculate 1ml of the Bacillus subtilis G-34 seed solution obtained in step (3) into 50ml of the fermentation medium obtained in step (2). Placed at 37°C, cultured at 200r...

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Abstract

The invention discloses bacillus subtilis. The bacillus subtilis which is classified and named as G-34(Bacillus subtilis G-34) is collected in China Center for Type Culture Collection on January 6, 2014, and the collection register number is CCTCC NO:M2014003. A bacillus subtilis strain is obtained via mutation breeding on Bacillus subtilis strain separated from polluted soil of Shengli Oil Field; a culture medium with glycerin as the only carbon source can be rapidly utilized for preparing a lipopeptide biosurfactant surfactin at a high yield. With crude glycerin as a substrate, the pure surfactin can be obtained through the steps of bacillus subtilis fermenting-transforming, centrifugal degerming, acid precipitating, freeze-drying, and methanol extracting.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to Bacillus subtilis G-34 ( Bacillus subtilis G-34), especially relates to the method of using the bacterial strain to transform glycerol and ferment to produce lipopeptide biosurfactant. Background technique [0002] Lipopeptides are a class of compounds composed of hydrophilic peptide chains and lipophilic aliphatic chains. It contains a peptide chain composed of 7-10 amino acids and a β-hydroxy fatty acid chain or a β-amino fatty acid chain, wherein the hydroxyl or amine group on the fatty acid chain combines with the carboxyl group on the amino acid of the peptide chain to form a lactone bond or amide bond, The peptide chains are closed to form a cyclic lipopeptide. Because lipopeptide has a special chemical composition and amphiphilic molecular structure, it has broad application prospects in the fields of medicine, food, cosmetics and microbial oil recovery, and has become a rese...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P21/00C12R1/125
Inventor 李霜刘强黄和江凌
Owner NANJING UNIV OF TECH
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