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Bacillus amyloliquefaciens and method for producing chymosin by fermenting using same

A technology of amylolytic spores and rennet, which is applied in the field of fermented production of rennet, Bacillus amyloliquefaciens strains, and Bacillus amyloliquefaciens, which can solve problems such as gaps and achieve control of growth conditions, high enzyme production levels, and easy growth conditional effect

Active Publication Date: 2012-09-05
四川生力源生物工程有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, compared with mold, the rennet activity produced by bacteria still has a certain gap. Currently, the level of enzyme production by bacterial fermentation does not exceed 3000 Soxhlet units per milliliter of fermentation broth

Method used

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  • Bacillus amyloliquefaciens and method for producing chymosin by fermenting using same
  • Bacillus amyloliquefaciens and method for producing chymosin by fermenting using same

Examples

Experimental program
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Effect test

Embodiment 1

[0020] Example 1 Screening, isolation and identification of Bacillus amyloliquefaciens CCTCC NO: M 2011045

[0021] 1. Bacillus amyloliquefaciens JNU002 ( Bacillus amyloliquefacines JNU002) screening separation

[0022] Bacillus amyloliquefaciens JNU002 was isolated and screened from bran purchased from Sanliqiao Grain and Oil Market in Wuxi City. Get 6 grams of bran into a 250 milliliter sterilized Erlenmeyer flask containing 10 small glass beads, add 100 milliliters of sterile saline, vibrate and cultivate on a shaker at 150 rpm for 20 minutes, and let stand for 2 hours; Transfer 90 ml of the above mixture into a conical flask filled with 90 ml of nutrient broth medium (peptone 5g / L, glucose 5g / L, sodium chloride 5g / L), at 33°C and 150 rpm Cultivate for 12 hours; take the above culture solution and dilute it to 10 -6 、10 -7 、10 -8 Take 100 microliters of the diluted solution of each concentration and spread it on the milk medium plate (skim milk powder 25g / L, agar 16g...

Embodiment 2

[0036] Example 2 Production of chymosin by liquid fermentation of Bacillus amyloliquefaciens CCTCC NO: M 2011045

[0037] (1) Slant culture: Bacillus amyloliquefaciens CCTCC NO: M 2011045 was inoculated on the slant medium and cultured at 32°C for 24 hours. The slant medium used was (unit: g / L): glucose 20, yeast extract 10, agar strip 20, prepared with tap water, pH natural, sterilized at 115°C for 20 minutes.

[0038](2) Seed culture: Use an inoculation loop to take a loop of the grown bacteria on the slant and insert it into the seed medium, cultivate at 35°C for 12 hours, and the shaker rotates at 150 rpm. The seed medium used is (gram / liter): Bran 30, sodium chloride 5, magnesium sulfate heptahydrate 5, potassium dihydrogen phosphate 2, calcium carbonate 3, prepared with tap water, pH natural; each 250 ml triangular bottle 30ml of seed medium, sterilized at 121°C for 30 minutes.

[0039] (3) Liquid fermentation culture: the fermentation medium used is (unit is gram per ...

Embodiment 3

[0040] Example 3 Production of rennet by liquid fermentation of Bacillus amyloliquefaciens CCTCC NO: M 2011045

[0041] The cultured seeds were placed in a 500 ml Erlenmeyer flask, in which the fermentation medium was 50 ml, the inoculum size was 100 microliters, and cultured at 32° C. for 60 hours with a shaker rotating at 150 rpm. The fermentation medium used is (g / L): glucose 20, bran 10, sodium chloride 6, magnesium sulfate heptahydrate 4.5, potassium dihydrogen phosphate 1, calcium carbonate 3, prepared with tap water, pH natural, 121°C Sterilize for 30 min. Other conditions were the same as in Example 2. After the cultivation was completed, the chymosin enzyme activity of the fermentation liquid was 3560 SU / mL.

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Abstract

The invention relates to Bacillus amyloliquefaciens and a method for producing chymosin by fermenting using the same and belongs to the technical field of microbial fermentation. The invention discloses a bacterial strain for producing the chymosin. The bacterial strain is named as Bacillus amyloliquefaciens JUN002 and is collected in China Center for Type Culture Collection, wherein the collection number is CCTCC No:M2011045. The method for producing the chymosin by using the bacterial strain through liquid fermentation comprises the following steps of: performing slant culture; (2) performing seed culture; and (3) performing liquid fermentation culture, namely inoculating cultured seeds into a fermentation culture medium according to the inoculation quantity of 0.1 to 0.2 percent to culture the seeds at the temperature of between 32 and 36 DEG C for 60 to 72 hours. The Bacillus amyloliquefaciens and the method for producing the chymosin by fermenting using the same have the advantages that: the chymosin is produced by using the Bacillus amyloliquefaciens CCTCC No:M2011045 through liquid fermentation, so the enzyme production level is high, the fermenting time is short, growth conditions are easy to control and the like; and the method is completely suitable for application to industrialized production.

Description

technical field [0001] A strain of Bacillus amyloliquefaciens producing chymosin, specifically a strain of Bacillus amyloliquefaciens, and a method for producing rennet by fermentation of the strain. The invention belongs to the technical field of microbial fermentation. Background technique [0002] α-casein and β-casein in casein micelles will not coagulate spontaneously under the action of κ-casein. Rennet is a key enzyme in cheese production. It makes raw milk coagulate into two stages: firstly, rennet specifically decomposes κ-casein into p-κ-casein and a macropeptide, making The micelles of casein lose their stability; then in Ca 2+ Under the action, chemical bonds are formed between p-κ-casein and α-casein and β-casein to form a precipitate. [0003] Traditionally, rennet is extracted from the fourth stomach (abomasum) of calves. With the continuous expansion of the cheese industry, the production of rennet by simply slaughtering young animals can no longer meet th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N9/54C12R1/07
Inventor 丁重阳王博达丁明亮王迎娜王望斐肖思维欧阳安然顾正华张梁石贵阳
Owner 四川生力源生物工程有限公司
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