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Basophilic micrococcus and alkaline xylanase produced from basophilic micrococcus and application

An alkalophilic micrococcus and xylanase technology, applied in alkaline xylanase and application fields, can solve the problems of incapability of large-scale industrial production of heat-resistant alkaline xylanase, short service life, poor stability and the like, Achieve the effect of being conducive to large-scale fermentation production, inhibiting bacterial contamination, and stabilizing enzyme activity

Inactive Publication Date: 2015-04-29
SHENZHEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there are still some problems in the direct use of xylanase in industry: poor stability, low substrate specificity, short service life and high cost.
The development of industrial application of xylanase has been carried out internationally, but only a small number of companies can produce xylanase for the paper industry, and there is still room for improvement in its alkali resistance and heat resistance; Systematic research on heat-resistant alkaline xylanase, but it is also impossible to produce heat-resistant alkaline xylanase on a large scale

Method used

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  • Basophilic micrococcus and alkaline xylanase produced from basophilic micrococcus and application
  • Basophilic micrococcus and alkaline xylanase produced from basophilic micrococcus and application
  • Basophilic micrococcus and alkaline xylanase produced from basophilic micrococcus and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1 Isolation and cultivation of strains

[0051] 1. Enrichment culture

[0052] The alkaline xylanase-producing strains were screened from Shenzhen mangrove soil. Weigh 1g of soil sample, add it to 100mL of sterile water, shake well to make a bacterial suspension, respectively take 1mL of bacterial suspension and insert it into 5mL enrichment medium that has been sterilized beforehand, at 37℃, 200rpm for enrichment Set culture for 48h.

[0053] Enrichment medium: xylan 8.0g / L, peptone 10g / L, NaCl 15g / L, KH 2 PO 4 1.5g / L, Na 2 HPO 4 ·12H 2 O9.0g / L, MgSO 4 ·7H 2 O 2.0g / L, pH 9.0.

[0054] 2. Flat screen

[0055] Dilute the enriched and cultured bacterial solution by 10 times, and take the dilution factor as 10 -3 , 10 -5 , 10 -7 Spread 0.5 mL of the bacterial suspension on an alkaline selective medium plate containing xylan, and incubate at 37°C for 48 hours. Pick a single colony with a larger transparent circle into the alkaline slant medium. Repeat this many times until...

Embodiment 2

[0064] Example 2 Separation and purification of alkaline xylanase produced by Micrococcus sp.S1

[0065] (1) Materials

[0066] 1. Bacteria

[0067] Alkaliophilic Micrococcus is an alkaline xylanase-producing strain Micrococcus sp.S1 (CCTCC NO:M 2014295) screened from Shenzhen mangrove soil.

[0068] 2. Medium

[0069] Selection medium: Xylan 8.0g / L, KNO 3 1.0g / L, MgSO 4 ·7H 2 O 0.5g / L, NaCl 15g / L, KH 2 PO 4 1.5g / L, solid medium plus 15-20g / L agar, pH 9.0; culture temperature is 37°C.

[0070] 3. Main reagents

[0071] The ion exchange chromatography media DEAE Sepharose Fast Flow and SP-sepharose Fast Flow cation exchange chromatography media were all purchased from GE. Acrylamide and N,N-methylene bisacrylamide are imported from Serva. DNA polymerase and DNA molecular weight markers are products of Bao Biology; protein molecular weight markers are products of Fermentas (MBI); ethylene diamine tetraacetic acid (EDTA), ethylene glycol diethyl ether diamine tetraacetic acid (EGTA), guan...

Embodiment 3

[0094] Example 3 Study on the enzymatic properties of alkaline xylanase

[0095] (1) Test method

[0096] 1. Optimal reaction pH and pH stability

[0097] Take an appropriate amount of the pure enzyme solution of alkaline xylanase obtained in Example 2 and add 1% xylan solutions with different pH values ​​to determine the enzyme activity according to a conventional method. At the same time, the pure enzyme solution was stored under different predetermined pH conditions for 30 minutes, and then the enzyme activity was measured at pH 9.0 according to the conventional method.

[0098] 2. Research on the optimum reaction temperature and temperature stability

[0099] A proper amount of the pure enzyme solution of alkaline xylanase obtained in Example 2 was placed in different temperature conditions and reacted for 20 minutes to determine its enzyme activity. At the same time, the pure enzyme solution was placed under different temperature conditions (30°C to 80°C) for 10 minutes, and then...

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Abstract

The invention discloses a basophilic micrococcus and alkaline xylanase produced by the basophilic micrococcus and an application. The basophilic micrococcus is a bacterial strain for producing alkaline xylanase, which is obtained by screening from mangrove soil in Shenzhen. The invention further discloses the alkaline xylanase produced from the basophilic micrococcus. The enzyme has excellent enzymatic characteristics of wide pH application range, heat resistance and alkaline resistance; metal ions, a surfactant and the like are suitable for industries such as papermaking. The basophilic micrococcus disclosed by the invention belongs to alkali-resistant extremophiles, can be cultivated under a high-alkali condition, and has the characteristics that living contaminants can be effectively inhibited, and large-scale fermenting production is facilitated.

Description

Technical field [0001] The invention belongs to the field of biological technology, and particularly relates to a kind of alkaliphilic micrococcus and the alkaline xylanase produced therefrom and application thereof. Background technique [0002] Xylanase is widely distributed in bacteria, fungi, yeast, actinomycetes, ruminant rumen, snails, crustaceans, land plant tissues and various invertebrates, among which neutral or alkaline xylan derived from bacteria The enzyme has high alkali resistance and thermal stability. Xylanase has a wide range of applications. In the paper industry, it can be used in pulp bleaching and waste paper deinking; in the feed industry, xylanase can reduce the chyme viscosity caused by water-soluble xylan in wheat-based diets. Increased sex, thereby improving the efficiency of digestive enzymes on substrates. At the same time, xylanase can act on insoluble non-starch polysaccharides, break plant cell walls, and release nutrients; it is also widely used ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/42D21C9/10C12R1/265
CPCC12N9/248C12N1/205C12R2001/265D21C9/10
Inventor 邢苗刘森林陈伟钊
Owner SHENZHEN UNIV
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