A kind of neutral endoglucanase produced by alkaliphilic streptomyces and its application

A technology of endoglucanase and streptomyces, which is applied in the field of neutral glucanase and application, can solve the problems of restricting the use and development of bacteria and molds, and achieve the benefits of large-scale fermentation production and stable enzyme activity , the effect of inhibiting bacterial contamination

Inactive Publication Date: 2017-10-20
SHENZHEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For reasons such as these, the use and development of bacteria and molds are limited

Method used

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  • A kind of neutral endoglucanase produced by alkaliphilic streptomyces and its application
  • A kind of neutral endoglucanase produced by alkaliphilic streptomyces and its application
  • A kind of neutral endoglucanase produced by alkaliphilic streptomyces and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] The isolation and cultivation of embodiment 1 bacterial strain

[0049] 1. Enrichment culture

[0050] A strain producing neutral endoglucanase was screened from mangrove soil in Shenzhen. Weigh 1g of soil sample, add it to 100mL sterile water, shake well to make a bacterial suspension, take 1mL of the bacterial suspension and put it into 5mL enrichment medium that has been sterilized beforehand, and enrich the medium at 37°C and 200rpm. Set cultured for 48h.

[0051] Enrichment medium: CMC-Na 8.0g / L, peptone 10g / L, NaCl 15g / L, KH 2 PO 4 1.5g / L, Na 2 HPO 4 12H 2 O 9.0g / L, MgSO 4 ·7H 2 O 2.0g / L, pH 9.0.

[0052] 2. Flat plate primary screening

[0053] Dilute the enriched bacterial solution 10 times step by step, and take the dilution factor as 10 -3 、10 -5 、10 -7 0.5mL of the bacterial suspension was spread on the alkaline selective medium plate containing CMC-Na, cultured at 37°C for 48h, and the single colony with a large transparent circle was picked in...

Embodiment 2

[0062] The enzymatic activity of the neutral endoglucanase produced by Example 2 Streptomyces sp.H31

[0063] (1) Materials

[0064] 1. Strains

[0065] Streptomyces alkalophilus is a neutral endoglucanase-producing strain Streptomyces sp.H31 (CCTCC NO:M 2015003) screened from Shenzhen mangrove soil.

[0066] 2. Medium

[0067] Selection medium: 10g / L CMC-Na (sodium carboxymethylcellulose), 5g / L yeast powder, 5g / L NaCl, 2g / LKH 2 PO 4 , 5g / L NH 4 NO 3 , 0.3g / L MgSO 4 , 1g / L (NH 4 ) 2 SO 4 , solid medium plus agar 15-20g / L, pH 9.0; culture temperature is 37°C.

[0068] (2) Experimental method

[0069] 1. Extraction of crude enzyme solution

[0070] Put the Streptomyces sp.H31 strain into the liquid selection medium, cultivate it at 37°C and 200rpm for 48h, freeze and centrifuge the 48h culture solution at 14000rpm for 20min, and take the supernatant; The supernatant was concentrated by ultrafiltration to obtain a crude enzyme solution.

[0071] 2. Enzyme activity a...

Embodiment 3

[0095] (1) Materials

[0096] 1. Strains. Streptomyces alkalophilus is a neutral endoglucanase-producing strain Streptomyces sp.H31 (CCTCC NO:M 2015003) screened from Shenzhen mangrove soil. E.coli TOP10F' was purchased from Invitrogen;

[0097] 2. Carrier. The Escherichia coli cloning vector pMD-19T (simple) was purchased from Dalian Bao Biological Company, and the Escherichia coli expression vector pET-28a(+) (Novagen, KanR) was purchased from Novagen Company.

[0098] 3. Medium

[0099] (1) Selection medium: 10g / L CMC-Na (sodium carboxymethylcellulose), 5g / L yeast powder, 5g / L NaCl, 2g / LKH 2 PO 4 , 5g / L NH 4 NO 3 , 0.3g / L MgSO 4 , 1g / L (NH 4 ) 2 SO 4 , solid medium plus agar 15-20g / L, pH 9.0; culture temperature is 37°C.

[0100] (2) LB medium: tryptone 10g / L, yeast extract 5g / L, NaCl 10g / L, solid medium plus agar 15-20g / L, pH 9.0, autoclaved at 121°C for 20min.

[0101] 4. Main reagents. DNA polymerase and DNA molecular weight markers are products of Bao Biol...

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Abstract

The invention discloses an alkaliphilic streptomyces, a neutral endoglucanase produced by it and an application thereof. The alkaliphilic streptomyces is a neutral endoglucanase-producing strain screened from Shenzhen mangrove soil. The invention also discloses the neutral endoglucanase produced by the above-mentioned alkaliphilic streptomyces. The enzyme has a wide range of pH adaptability, and is suitable for washing and papermaking industries such as heat resistance, alkali, metal ions and surfactants. Enzymatic properties, can be used in industries such as washing and papermaking. The alkaliphilic streptomyces of the present invention belongs to alkali-resistant extremophile microorganisms, can be cultivated under high alkaline conditions, has the characteristics of effectively inhibiting contamination by miscellaneous bacteria, and is beneficial to large-scale fermentation production.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to an alkaliphilic streptomyces and the neutral glucanase produced by it and its application. Background technique [0002] Cellulose is the main component of plant fiber, accounting for about 30-50% of its dry weight. It is the most widely distributed natural carbohydrate and the most abundant and cheapest renewable resource on the earth. With the rapid increase of the world population, the rapid development of the global economy, the shortage of food and the reduction of non-renewable resources such as oil, coal and natural gas at an alarming rate, the development and utilization of renewable cellulose resources has attracted widespread attention around the world and has become A major topic of research in countries all over the world. The study of cellulase has opened up a broad way for this, especially in the past 10 years, with the rapid development of modern b...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N9/42C12R1/465
CPCC12N9/2437C12Y302/01004C12N1/205C12R2001/465
Inventor 刘森林区晓阳邢苗陈伟钊
Owner SHENZHEN UNIV
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