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Bacterial strain capable of producing alkali protease and industrialized liquid fermentation method of bacterial strain

A liquid fermentation and protease technology, applied in the field of bioengineering, can solve the problem of high production cost, achieve the effects of simple operation, mild culture conditions and lower production cost

Active Publication Date: 2013-11-27
SHANDONG LONGKETE ENZYME PREPARATION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the enzyme activity of alkaline protease that can be industrially produced in the market is generally low, making the production cost corresponding to the unit enzyme activity relatively high, which is far from meeting the needs of the current detergent and other industries

Method used

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  • Bacterial strain capable of producing alkali protease and industrialized liquid fermentation method of bacterial strain
  • Bacterial strain capable of producing alkali protease and industrialized liquid fermentation method of bacterial strain
  • Bacterial strain capable of producing alkali protease and industrialized liquid fermentation method of bacterial strain

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Experimental program
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Effect test

Embodiment 1

[0037] Embodiment 1: Screening of starting bacterial strain

[0038] (1) Take 2g soil sample from the saline-alkali land rich in organic matter, 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, continuously Shake the Erlenmeyer flask to mix the soil sample, 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.

[0039] (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.

[0040]...

Embodiment 2

[0046] The mutagenesis screening of embodiment 2 bacterial strains

[0047] (1) Ultraviolet mutagenesis

[0048] Aspirate the 10-12h bacterial solution of the originating strain and pour it into a flat dish, and the amount of bacterial solution in each flat dish is 10mL. Place the plate containing the bacterial solution on the magnetic stirrer under the ultraviolet lamp in the mutagenesis box, and the vertical distance between the plate and the ultraviolet lamp is 30 cm. Turn on the UV lamp and preheat for 30 minutes to stabilize the light wave. Adjust the rotation speed of the stirring bar. After the rotation speed is stable, open the lid of the plate for irradiation, and start timing. The irradiation time of the plate is 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 minutes respectively. Take out 0.1ml of bacterial suspension from each plate and make appropriate dilutions to obtain bacterial suspensions with different dilutions. Draw 0.3ml of the diluted bacterial suspension to coat ...

Embodiment 3

[0063] A kind of liquid submerged aerated fermentation method using Ap180 alkalophilic bacillus strain as production strain, comprises the steps:

[0064] (1) First-level seed culture: Inoculate the slant strain of Bacillus alkalophilus Ap180 into a 500-ml shaker flask, with a medium volume of 100 ml, a rotary shaker at 180 rpm, a culture temperature of 34°C, and a culture time of 11 hours;

[0065] (2) Secondary seed cultivation: Put the first-grade seeds into 500 ml second-grade seed shake flasks according to the inoculation amount of 10%, and the cultivation conditions are the same as the first-grade seeds;

[0066] (3) Tertiary seed cultivation: Inoculate the secondary seeds with 10% inoculum amount into a 5000 ml tertiary seed shaker flask, with a medium capacity of 1000 ml, a rotary shaker at 100 rpm, and a culture temperature of 34°C. time 11h;

[0067] (4) First-level seed tank cultivation: Put the third-level seeds into the first-level seed tank with a total volume o...

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Abstract

The invention discloses a bacterial strain capable of producing alkali protease and an industrialized liquid fermentation method of the bacterial strain, and belongs to the field of bioengineering technology. The bacterial strain is mutant strain Ap180 of Bacillus alcalophilus, and the preservation number is CGMCC No.7545. The bacterial strain is obtained by repeat UV-induced mutation, nitrosoguanidine-induced mutation and selection of a wild bacterial strain obtained from saline lands. Characteristics of the bacterial strain are that: efficiency of alkali protease production is high, and stability is excellent. The Bacillus alcalophilus strain, which is capable of producing alkali protease and is suitable for industrialized production , is provided in the invention, and related fermentation mechanism is optimized. According to the fermentation mechanism, operation processes are simple, cultivating conditions are mild, fermenting enzyme activity is more than 80000u / ml, production cost of enzyme activity per unit is reduced, and the requirements of market are met. Applications of the alkali protease in fields such as washing agent, forage, and leather industry are more extensive.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, in particular to an alkaline protease-producing bacterial strain and an industrialized liquid fermentation method thereof. Background technique [0002] Alkaline Protease is a protease with an optimum pH of 9-11, with a molecular weight of 26,000-34,000 Daltons, and is generally stable at a pH of 5-10. Because its active center contains serine (Ser), it belongs to serine protease. It not only hydrolyzes peptide bonds, but also has the functions of hydrolyzing ester bonds, amide bonds, transesterification and transpeptide. Widely used in detergent, medical, silk, leather and other industries. [0003] In recent years, with the improvement of people's living standards and the needs of environmental protection, phosphorus-containing detergents have been basically eliminated, replaced by a large number of applications of various enzyme-added detergents. Because the aqueous solution of deter...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/54C12R1/07
Inventor 王兴吉刘顺启孙硕张杰
Owner SHANDONG LONGKETE ENZYME PREPARATION
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