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Microorganism for producing beta-1,3-glucanase and application thereof

A glucanase, microorganism technology, applied in the direction of hydrolase, bacteria, etc., can solve the problem of insufficient bacteria source

Inactive Publication Date: 2009-10-28
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the defect that the existing β-1,3-glucanase produces insufficient bacterial sources, the present invention provides a microorganism and its application for producing β-1,3-glucanase

Method used

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  • Microorganism for producing beta-1,3-glucanase and application thereof
  • Microorganism for producing beta-1,3-glucanase and application thereof
  • Microorganism for producing beta-1,3-glucanase and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] The isolation and screening of chitosanase producing bacteria H12, the steps are as follows:

[0039] The soil samples were collected in Baotu Spring Brewery, planted on the screening medium, and cultivated at 30°C for 3 days, picked the edge of the colony with a large transparent circle of the degraded yeast cell wall, and transferred it to the screening medium again, at 30°C Cultivate for 3 days, pick out the thalli at the edge of the transparent circle of the degraded yeast cell wall, and separate them by streaking to obtain pure colony strains. The strain was identified as chitosanase-producing strain H12 (Mitsuaria chitosanitabida H12), and its 16SrRNA gene sequence is shown in Seq. Nol.

[0040] The screening medium used is: Angel yeast 5.0g, CaCl 2 2H 2 O 1.36g, agar 10g, add water to 1L, sterilize at 121°C for 30min.

Embodiment 2

[0041] Embodiment 2: Fermentation culture and β-1,3-glucanase activity determination of chitosanase producing bacteria H12

[0042] The fermentation medium is: 1-2% (w / v) Angel yeast, pH 6-7, autoclaved at 121° C. for 30 minutes. Inoculate the chitosanase-producing bacteria H12 obtained by the separation and screening of the present invention at an inoculation amount of 3-5% (v / v), culture at 30° C. at 160 rpm for 15h-18h on a shaking table to obtain crude enzyme liquid.

[0043] The obtained crude enzyme liquid takes laminarin as a substrate, and uses 3,5-dinitrosalicylic acid (DNS) method to measure the activity of β-1,3-glucanase, and the β-1,3-glucanase activity of the crude enzyme liquid is - Glucanase activity is 3-5U / ml. The optimum reaction temperature of β-1,3-glucanase is 50°C, and the optimum reaction pH is pH 6. The optimal reaction temperature of β-1,3-glucanase is as follows figure 1 shown. The optimal reaction pH of β-1,3-glucanase is as follows figure 2 s...

Embodiment 3

[0046] Example 3: Preparation of crude enzyme preparation of β-1,3-glucanase

[0047] With the fermented and cultivated crude enzyme liquid of chitosanase producing bacteria H12 obtained in Example 2, centrifuge 30min at 4000-5000rpm, get the supernatant, slowly add solid ammonium sulfate until the concentration of ammonium sulfate is 70-75% (w / v) Saturation, centrifuge at 8000-10000rpm for 30min, put the obtained precipitate into a dialysis bag with a semi-permeable membrane, tie the bag tightly and place it in a 1000mL beaker of 0.01mol / L acetate buffer solution with pH 6.0, dialyze overnight in the refrigerator, and replace About 4 times with acetate buffer. Remove the enzyme solution in the dialysis bag, freeze overnight at -20°C, and freeze-dry in a freeze dryer for 36 hours to obtain a crude enzyme preparation of β-1,3-glucanase in the form of dry powder.

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Abstract

The invention relates to a microorganism for producing beta-1,3-glucanase and application thereof. The microorganism for producing the beta-1,3-glucanase has a category name of Mitsuaria chitosanitabida H12 and a preservation number of CGMCC 2949, and is applied to preparing the beta-1,3-glucanase. By preparing bacterial fermentation cultivating crude enzyme solution of the Mitsuaria chitosanitabida H12, a crude enzyme solution dry powder preparation of the beta-1,3-glucanase is further prepared. The beta-1,3-glucanase and the crude enzyme solution dry powder preparation of the beta-1,3-glucanase can be widely applied to the fields of brewing industry, feed industry, biological control, medicaments and the like.

Description

technical field [0001] The invention relates to a microorganism producing beta-1,3-glucanase and a preparation method and application of a crude enzyme preparation prepared by fermentation and culture of bacteria, belonging to the field of biotechnology. technical background [0002] β-glucan is widely distributed in nature and is one of the structural macromolecules that make up the cell walls of higher plants, yeasts and filamentous fungi. β-glucan widely exists in wheat crops, and its main structure is β-1,3-1,4-glucan. The polysaccharide content in yeast cell wall is 85-90%, and its main component is β-1, 3-glucan of β-1,6-branch. Yeast cell walls are an important source of β-1,3-glucan. The main component of the cell wall of many important plant pathogenic fungi is also β-1,3-glucan, which together with chitin constitutes the network skeleton of the fungal cell wall. [0003] β-1,3-glucanase (β-1,3-glucanase, EC3.2.1.39) is a kind of hydrolase that can specifically a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/24
Inventor 卢雪梅樊志华刘巍峰段峰张创
Owner SHANDONG UNIV
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