A method for producing beta-galactosidase by liquid fermentation of Bacillus coagulans

A galactosidase, fermentation and culture technology, applied in microorganism-based methods, glycosylases, biochemical equipment and methods, etc., can solve the problems of scarcity, low β-galactosidase enzyme activity, etc. The effect of production cost, simple process operation and mild culture conditions

Active Publication Date: 2019-01-11
SHANDONG LONGKETE ENZYME PREPARATION
View PDF3 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the enzyme activity of β-galactosidase produced by natural enzyme-producing strains is generally low, and the resources of excellent natural strains are scarce. Therefore, it is of great significance to breed new excellent strains with high enzyme production.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A method for producing beta-galactosidase by liquid fermentation of Bacillus coagulans
  • A method for producing beta-galactosidase by liquid fermentation of Bacillus coagulans
  • A method for producing beta-galactosidase by liquid fermentation of Bacillus coagulans

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Embodiment 1 starts the screening of bacterial strain

[0056] (1) Take 2g of soil sample from the breeding base of the dairy farm, add it to a small triangular flask containing 50mL of cooled sterile water, shake the table at 200r / min, shake for 20min, then place it in a water bath at 80°C for 10min, and shake it constantly Mix the soil samples in the triangular flask, let it stand for 5 minutes, absorb 100 μL of the supernatant, and gradually dilute to 10 -1 ~10 -9 Concentration, select 10 -3 , 10 -4 , 10 -5 , 10 -6 Concentration coated beef extract peptone plate, cultivated at 37°C for 24h, and continued to cultivate at 30°C for 24h.

[0057] (2) According to the characteristics of the shape, size, surface structure, edge structure, texture, gloss, transparency, color and soluble pigments of the microbial colony, use the inoculation loop to pick out the isolated single colony and move it to the screening culture Base plate, and numbered, 34 ℃ for 48h.

[0058]...

Embodiment 2

[0064] The mutagenesis selection of embodiment 2 bacterial strains

[0065] (1) After absorbing and activating the starting strain, collect the bacteria by centrifugation, resuspend the bacteria with 5% glycerol, count with a hemocytometer until the bacterial concentration is 10 7 -10 8 cells / mL, as the starting bacterial suspension.

[0066] (2) Turn on the normal temperature and pressure plasma system, wipe the inside and outside of the operating room with alcohol cotton, and turn on the ultraviolet light for 30 minutes to sterilize. After the sterilization, take 10 μL of the bacterial suspension and spot it on the rough surface of the slide, and transfer the slide to the operating room table with tweezers under sterile conditions. Open the helium gas valve, set the gas flow and mutagenesis time for mutagenesis. The mutagenesis time was set as 90s, 120s, 150s, 180s, 210s respectively. After each mutagenesis, the slides were placed in an EP tube containing 990 μL sterile ...

Embodiment 3

[0078] Example 3 Liquid Fermentation Produces Endo-β-Galactosidase and Its Extraction and Purification

[0079] (1) First-level seed cultivation: put the Bacillus coagulans mutant strain BR-171 into a 500-ml shake flask, with a medium volume of 100 ml, a rotary shaker at 180 rpm, and a culture temperature of 34-35°C , culture time 11h;

[0080] (2) secondary seed cultivation: the primary seed is inserted into a 500 milliliter secondary seed shake flask according to the inoculum size of 10%, and the cultivation condition is the same as that of the primary seed;

[0081] (3) Tertiary seed cultivation: insert the secondary seed into a 3000 ml third-stage seed shake flask with 10% inoculum size, 800 ml of culture medium, 100 rpm on a rotary shaker, and a culture temperature of 34-35°C , culture time 11h;

[0082] (4) One-level seed tank cultivation: the three-level seeds are inserted into the first-level seed tank with a total volume of 50L with 3.5% inoculum size, the fermentat...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention belongs to the technical field of fermentation engineering, in particular to a method for producing beta-galactosidase by liquid fermentation of Bacillus coagulans and the produced beta-galactosidase. Bacillus coagulans is specifically Bacillus coagulans (Bacillus coagulans) mutant BR-171, with the deposit number of CGMCC No. 14413. The strain is capable of producing beta-galactosidase stably and efficiently. The yield is more than 4000u/ml after submerged fermentation in a 50L fermentor and fed-batch fermentation for 84h. The beta-galactosidase produced by the strain has an optimum pH of 5.0, and still has an activity of more than 80% in the range of beta-galactosidase pH 2.5-8.5. The pH tolerance of galactosidase is suitable for most dairy products. The optimum temperatureof beta-galactosidase is 60 DEG C, and the activity of beta-galactosidase still reaches 88.4% after incubation at 65 DEG C for 2 h, which indicated that beta-galactosidase had significant heat resistance. It has high industrial value in dairy industry.

Description

Technical field: [0001] The invention belongs to the technical field of fermentation engineering, in particular to a method for producing beta-galactosidase by liquid fermentation of bacillus coagulans and the produced beta-galactosidase. Background technique: [0002] Bacillus coagulans, a Gram-positive bacterium, belongs to the phylum Firmicutes (or Firmicutes). Bacillus coagulans belongs to the genus Bacillus in taxonomy, the cells are rod-shaped, terminal spores, without flagella, decompose sugars to generate L-lactic acid, and are homolactic acid fermenters. The optimum growth temperature is 45-50°C, and the optimum pH is 6.6-7.0. Bacillus coagulans is a facultative anaerobic bacterium that can grow in both aerobic and anaerobic environments, can adapt to a low-oxygen intestinal environment, has a high tolerance to acid and bile, can carry out lactic acid fermentation, and produces The L-lactic acid can lower the pH value of the intestinal tract, inhibit harmful bacte...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/38C12N1/20C12R1/07
CPCC12N1/20C12N9/2471C12Y302/01023
Inventor 郭庆文刘文龙王兴吉郭庆亮王克芬张杰
Owner SHANDONG LONGKETE ENZYME PREPARATION
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap