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Geobacillus thermodenitrificans obtained by artificial mutagenesis and application thereof in cellulose degradation

A thermoglucosidase, Geobacillus technology, applied in the field of microorganisms, can solve the problem that P.thermoglucosidasius has not been reported in cellulose degradation, and achieve the effect of high enzyme concentration

Active Publication Date: 2020-12-22
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The ability of P.thermoglucosidasius to degrade cellulose has not been reported so far

Method used

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  • Geobacillus thermodenitrificans obtained by artificial mutagenesis and application thereof in cellulose degradation
  • Geobacillus thermodenitrificans obtained by artificial mutagenesis and application thereof in cellulose degradation
  • Geobacillus thermodenitrificans obtained by artificial mutagenesis and application thereof in cellulose degradation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Isolation, Purification and Identification of High Temperature Cellulose Degrading Bacteria

[0029]Pig carcasses and straw compost samples were separated and purified in beef extract peptone medium, enriched with carboxymethyl cellulose sodium basal medium, selected strains were spotted on cellulose Congo red medium, cultured at 60°C for 72 hours, and after preliminary screening, A total of 27 single strains were obtained from the sodium carboxymethylcellulose medium plate, and then re-screened to measure the cellulase activity of each strain, and finally a strain with the highest enzyme activity was selected, which was identified as Geobacillus by sequencing thermodenitrificans CMC-4, after high-temperature domestication and ultraviolet mutagenesis, a strain with high cellulase production was obtained. This strain was sent to the China Center for Type Culture Collection on July 16, 2020 for preservation, preservation number: CCTCC NO: M2020324, taxonomic name: Parageo...

Embodiment 2

[0033] Thermoglucosidase Geobacillus Parageobacillus thermoglucosidasius BGSC95A1 cellulase enzyme activity assay:

[0034] The strain Parageobacillus thermoglucosidasius BGSC95A1 and its original strain Geo bacillus thermodenitrificans CMC-4 were inoculated in the liquid fermentation medium with 2% inoculum amount, cultured at 60°C and 150r / min for 72h, and the fermentation broth of the strain was centrifuged at 8000r / min for 30min to remove bacteria. Body and culture medium, the supernatant is the crude enzyme solution.

[0035] The formula of the liquid fermentation medium: beef extract 10g, peptone 10g, yeast extract 10g, sodium chloride 5g, potassium dihydrogen phosphate 1g, magnesium sulfate 0.2g, distilled water supplemented to 1000mL, natural pH. Sterilize at 121°C for 20 minutes.

[0036] Extract and obtain crude enzyme liquid, survey its carboxymethyl cellulose enzymatic activity, the detection method of enzymatic activity (being enzyme concentration) is as follows:...

Embodiment 3

[0049] Effects of different carbon sources on the enzyme production of Geobacillus thermoglucosidase BGSC95A1:

[0050] 1) with 1% carboxymethylcellulose sodium (CMC-Na), 1% microcrystalline cellulose (micro-cellulose), 1% soluble starch (starch), 1% glucose (glucose), 1% straw (straw) ) is the carbon source, and the adjusted pH value is 7.2. Inoculate the activated Geobacillus thermoglucosidase BGSC95A1 bacteria liquid (ie CMC-4-1) with 2% inoculum into a 250 mL Erlenmeyer flask containing 50 mL of liquid fermentation medium. After cultivating on a shaker at 55°C and 200r / min for 72h, centrifuge the strain fermentation broth at 8000r / min for 30min to remove the bacteria and the medium, and the supernatant is the crude enzyme solution; each group has 3 replicates without adding any carbon source 1. Peptone is CK group as nitrogen source; Measure cellulase activity (the method for embodiment 2), the enzyme activity of each group of detection, result is shown in the table below...

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Abstract

The invention belongs to the field of microorganisms, and discloses Geobacillus thermodenitrificans obtained by artificial mutagenesis and application thereof in cellulose degradation. The applicant separates a strain of Geobacillus thermoslucanase with the highest cellulase activity from a sample of pig carcass and straw compost in a high-temperature period, and obtains a strain of Geobacillus thermodenitrificans with high cellulase yield after high-temperature domestication and ultraviolet mutagenesis. The preservation number of the strain is CCTCC NO: M2020324. The Geobacillus thermodenitrificans BGSC95A1 with cellulase activity is reported for the first time. A plurality of carbon sources can be utilized, the Geobacillus thermodenitrificans has a degradation effect on a plurality of celluloses, and the cellulase concentration in fermentation liquor produced by fermentation is high.

Description

technical field [0001] The invention belongs to the field of microorganisms, and in particular relates to a thermoglucosidase geobacillus obtained by artificial induction and its application in cellulose degradation. Background technique [0002] As the most abundant and least utilized resource in the biosphere, cellulose can be hydrolyzed by microorganisms with the aid of a multi-enzyme system (Hahn- al 2006). Cellulase is a compound enzyme with synergistic effect, mainly including endoglucanase (EC3.2.1.4 endo-1 4-β-D-glucanase), exoglucanase (EC3.2.1.91, exo-1 4-β-D-glucanase) and β-glucosidase (EC.3.2.1.21, β-glucosidase). The three enzymes are coordinated to participate in the degradation of natural cellulose (Li Na 2006). Endoglucanase mainly works in the non-crystalline region of cellulose molecules, hydrolyzes β-1,4 glycosidic bonds, and makes long-chain cellulose molecules The chains become small molecular celluloses, creating more cleavage sites for exoglucanas...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/42C12P19/02C12P19/14C12P19/12C12R1/01
CPCC12N9/2437C12P19/14C12P19/12C12P19/02C12R2001/01C12N1/205Y02W30/40
Inventor 郑嵘韩珍珍杨旭晨刘建华李宇飞蒋思文柴进
Owner HUAZHONG AGRI UNIV
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