Bacillus cereus producing keratinase and application thereof

A technology of Bacillus cereus and keratinase, applied in the field of microorganisms, can solve the problems that the original enzyme production cannot meet the needs of efficient transformation of livestock and poultry waste, and the enzyme production efficiency of strains is not very high

Pending Publication Date: 2018-04-03
BIOTECH RES CENT SHANDONG ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although a large number of keratin-degrading bacteria have been screened, the enzyme production efficiency of the strains is relatively not very high, and the production of ori

Method used

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  • Bacillus cereus producing keratinase and application thereof
  • Bacillus cereus producing keratinase and application thereof
  • Bacillus cereus producing keratinase and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Screening, purification, identification and preservation of strains

[0025] 1.1 Strain screening and purification

[0026] The mud samples were collected from the mud that had been submerged in feather residue for a long time near the sewage outlet of a food factory in Ningyang, Shandong. Weigh 1 g of sludge sample, put it into a conical flask containing 10 mL of sterilized physiological saline and glass beads, and shake it to fully mix the sludge sample with water. Take 1mL of the suspension and add it into a test tube filled with 9mL of sterilized physiological saline and mix well, this is 10 -1 Diluent. and so on to make 10 -2 ~10 -6 Solutions in several dilutions. Take 100 μL of the above-mentioned solution samples of different dilutions and apply them to a milk screening plate (10% skimmed milk powder sterilized at 116°C for 30 minutes and mix with beef extract peptone medium (beef extract 0.5%, peptone 1%, NaCl 0.5%, pH 7.5) Mix according to the ratio of 1:...

Embodiment 2

[0034] Crude Enzyme Solution Extraction and Feather Degradation

[0035] The seed liquid of B. cereus Y-15 was inserted into the optimized enzyme-producing medium according to the inoculum amount of 2%. After culturing at 30°C and 180r / min for 48 hours, the bacteria were removed by centrifugation to obtain the crude enzyme liquid. 1L of medium can obtain a total enzyme activity of 11000U of crude enzyme solution. The crude enzyme solution treats feathers at 45°C and 150r / min. After 24 hours, visual observation shows that the overall volume of the enzymatically hydrolyzed feathers becomes smaller, the amount of feathers decreases, and the solution changes from clear to cloudy. The feather structure was destroyed, the rachis curled up, most of the barbs of the feathers were degraded and disappeared, and only part of the rachis remained (attached Figure 5 ). Under the experimental conditions, the degradation rate of feather waste is 50%, and the degradation effect is remarkabl...

Embodiment 3

[0037] Optimization of Enzyme Production Conditions in B. cereus Y-15

[0038] Referring to the method of Tiwary and Gupta (Bioresource Technology, 2010, 101 (15): 6103-6110.), Feather peptone medium (Feather peptone medium, FM1) (0.5% of feather meal, 0.5% of peptone, 1% of glucose, K 2 HPO 4 0.3%, KH 2 PO 4 0.1%, pH 7.5, 1×10 5 Pa sterilization 20min). B. cereusY-15 seed solution was inserted into FM1 medium at 2% inoculum size, and cultured at 30°C and 180 r / min for 48 hours, and the enzyme activity of keratinase in the fermentation broth was determined to be 0.52 U / mL.

[0039] The enzyme production conditions of B. cereus Y-15 were optimized by comprehensive use of single factor experiment, PB design and central combination design.

[0040] 3.1 Single factor experiment

[0041] Select culture temperature (30°C, 37°C), medium pH (6.0-10.0, set a pH gradient for every 1.0), inoculum size (volume ratio) (2%, 4%, 6%, 8%), 9 Common carbon sources for cultivating bacte...

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Abstract

The invention belongs to the field of microorganism, particularly relates to ((i) bacillus cereus (/i)) Y-15 for efficiently degrading feather keratin, and further relates to application of the ((i) bacillus cereus (/i)) Y-15. The ((i) bacillus cereus (/i)) Y-15 is preserved in China General Microbiological Culture Collection Center in June 6, 2017, with the culture preservation number being CGMCCNO. 14221; the colonial morphology on a milk screening plate is as follows: bacterial colony is white and circular, the waxiness on the surface is non-transparent, and the edge is wavelike; gram staining shows a positive result, and the thallus is bacilliform. The ((i) bacillus cereus (/i)) Y-15 screened and cultured has excellent degradation effect on keratin.

Description

technical field [0001] The invention belongs to the field of microbes, in particular to Bacillus cereus Y-15 strain producing keratinase, and also relates to the application of the above strain. Background technique [0002] As a by-product of poultry breeding and slaughtering, long-term stacking of feathers will cause soil erosion, accumulation of heavy metals, proliferation of pathogenic microorganisms, and release of harmful gases, which will cause serious environmental problems. At the same time, the dense and complex structure of feathers is difficult to degrade, which brings difficulties to waste disposal. Traditional landfill, incineration, alkali treatment and other methods to treat and transform feather waste are not only expensive and cause secondary pollution, but also cause a variety of amino acids to be destroyed during the transformation process. In recent years, the use of microorganisms and keratinases to harmlessly treat keratin waste under mild conditions ...

Claims

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Application Information

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IPC IPC(8): C12N1/20A23K20/147A23K10/18C12R1/085
CPCA23K10/18A23K20/147C12N1/20C12N1/205C12R2001/085
Inventor 边婓岳寿松朱耀霞马德源彭振英毕玉平
Owner BIOTECH RES CENT SHANDONG ACADEMY OF AGRI SCI
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