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Bacillus amyloliquefaciens for high production of neutral protease

A technology for dissolving starch spores and protease, which is applied in the field of microbial screening, can solve the problems of pollution, unstable fermentation, low fermentation unit of neutral protease, etc., and achieves the effect of increasing yield and having broad prospects.

Active Publication Date: 2015-11-25
WEIFANG KANGDIEN BIOTECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The research and development of neutral protease has been started since the 1970s, and industrialization was realized in the early 1980s at home and abroad, but the fermentation unit of neutral protease has always been low, the fermentation is unstable, and it is susceptible to bacteriophage and yeast. pollution

Method used

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  • Bacillus amyloliquefaciens for high production of neutral protease
  • Bacillus amyloliquefaciens for high production of neutral protease
  • Bacillus amyloliquefaciens for high production of neutral protease

Examples

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Effect test

Embodiment 1

[0017] Example 1 The ultraviolet mutagenesis and screening of the bacterium Bacillus amyloliquefaciens FN

[0018] Starting bacteria: Bacillus amyloliquefaciens FN ( Bacillus amylolique faciens FN), the strain was isolated by the inventor Fu Juan in May 2014 from the soil of the forest farm in Laoshan District, Qingdao City, Shandong Province, and can ferment and produce neutral protease.

[0019] Bacterial suspension preparation

[0020] Streak inoculation of the starting bacterium Bacillus amyloliquefaciens FN on LB slant (yeast powder 0.5%, peptone 1%, NaCl 1%, agar 2%), culture at 37°C for 24 hours; add 5 mL of 0.85% sterile saline, Wash all the bacteria on the inclined surface, transfer them into a sterile test tube containing glass beads, vortex and shake for 10 minutes, and completely break into single-cell bacteria; transfer all the bacteria suspension into a 15mL centrifuge tube, and centrifuge at 6000rpm for 3 minutes to collect bacteria cells, take the supernata...

Embodiment 2

[0040] Example 2 Enzymatic property analysis of protease produced by Bacillus amyloliquefaciens FN61

[0041] 2.1 Optimum pH analysis

[0042] Using disodium hydrogen phosphate-citric acid buffer solution with pH values ​​of 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, and 9.0, and borax-boric acid buffer solution with pH values ​​of 8.5 and 9.0, the above mutant strain FN61 Dilute with the fermentation supernatant of the starting strain FN, the substrates are also prepared with buffers corresponding to the pH value, and the protease enzyme activity is measured at 40°C, with the highest enzyme activity as 100%, the relative enzyme activity is calculated, and the pH is calculated. - Relative enzyme activity curve. The result is as figure 1 As shown, the optimum pH value of the protease produced by the mutant strain FN61 obtained in the present invention is 7.5, which is a neutral protease, which is consistent with the starting strain. However, the relative enzyme activity of the pr...

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Abstract

The invention relates to the technical field of functional microorganism screening, and in particular relates to a mutant strain, namely bacillus amyloliquefaciens FN61, which is obtained through ultraviolet mutagenesis, wherein the preservation number of the mutant strain is CCTCC NO: M2015527. The mutant strain can be used for significantly improving the yield of neutral protease, and fermenting enzyme activity in a 20L tank can reach up to14135U / ml, which is 42% higher than that of an initial strain. The optimum action pH of the neutral protease produced by the mutant strain is 7.5, which is consistent with the initial strain, but relative enzyme activity of the protease produced by the mutant strain is generally higher than that of the initial strain with a pH range of 6.5-8.5, showing that the protease produced by the mutant strain more sui table than the initial strain to play a role in a neutral condition; and the optimum action temperature of the protease produced by the mutant strain is at 40 DEG C, which is equal to that of the initial strain, so that an unexpected technical effect is achieved. The neutral protease produced by the mutant strain FN61 disclosed by the invention can be widely applied to the fields of feed and food processing; and the neutral protease is broad in prospect.

Description

technical field [0001] The invention belongs to the technical field of microbial screening, and in particular relates to a strain of bacillus amyloliquefaciens with high production of neutral protease and its application. Background technique [0002] Protease is a class of enzymes that catalyze the hydrolysis of peptide bonds, and can hydrolyze macromolecular proteins into amino acids, small peptides and other products. Neutral protease refers to a class of protease with the most suitable pH of 6.0-7.5. As a biocatalyst, it has the properties and advantages of fast catalytic reaction speed, no industrial pollution, and wide adaptability of catalytic reaction conditions. Microorganisms are favored by people because of their fast growth rate, small growth space required, wide biochemical diversity and genetic operability. According to statistics, microbial protease occupies about 40% of the world's entire enzyme sales market, and has become the main source of protease produc...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/54C12R1/07
Inventor 付娟齐建纪海涛刘文瑶郭小飞徐玉婷石增秀
Owner WEIFANG KANGDIEN BIOTECH
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