Phage lyase composite powder and preparation method and application thereof

A phage lyase, compound powder technology, applied in the directions of lyase, application, additional food elements, etc., can solve the problems of loss of activity, easy oxidation of residues, reduced activity, etc., to improve the bursa index and promote the daily increase The effect of regenerating and maintaining micro-ecological balance

Pending Publication Date: 2019-12-20
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Phage lyase belongs to the protease class, and it is easily interfered by gastric acid and bile salt during direct feeding; in addition, a large amount of pepsin and trypsin in the gast

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0022] Embodiment 1: The composition and mass percentage of this phage lyase compound powder are phage lyase TSPpgh 0.15%, phage lyase MMPpgh 0.2%, trehalose 0.5%, citric acid 0.1%, mannitol 0.2%, vitamin C 0.2 %, diatomite 98.65%.

[0023] The preparation method of the above-mentioned phage lyase compound powder is as follows: the phage lyase TSPpgh gene and the phage lyase MMPpgh gene are respectively cloned into the pET-28a(+) plasmid, and transformed into Escherichia coli BL21 after successful verification to obtain a phage lyase containing phage lyase. The genetically engineered bacteria with the TSPpgh gene and the genetically engineered bacteria with the phage lyase MMPpgh gene were inoculated into 300 mL of seed medium respectively, and cultured at 37°C to OD 600 = 2.0, then inoculate the fermentation medium (using a 20L automatic fermenter (Shanghai Bailun Biotechnology Co., Ltd.)) with an inoculum of 2%, and cultivate it at 37°C and 200rmp / min for 10h to the biomass ...

Example Embodiment

[0025] Example 2: The composition and mass percentage of the phage lyase compound powder in this example are phage lyase TSPpgh 0.2%, phage lyase MMPpgh 0.15%, trehalose 0.3%, citric acid 0.2%, mannitol 0.3%, vitamin C 0.3%, diatomite 98.55%.

[0026] The preparation method of the above-mentioned phage lyase compound powder is as follows: the phage lyase TSPpgh gene and the phage lyase MMPpgh gene are respectively cloned into the pET-28a(+) plasmid, and transformed into Escherichia coli BL21 after successful verification to obtain a phage lyase containing phage lyase. The genetically engineered bacteria with the TSPpgh gene and the genetically engineered bacteria with the phage lyase MMPpgh gene were inoculated into 300 mL of seed medium respectively, and cultured at 37°C to OD 600 = 1.5, then inoculate the fermentation medium (using a 20L automatic fermenter (Shanghai Bailun Biotechnology Co., Ltd.)) with an inoculum of 1.5%, and cultivate it at 37°C and 150rmp / min for 12h to...

Example Embodiment

[0028] Example 3: In vitro antibacterial activity detection of phage lyase composite powder

[0029] 1. Take 1 g of the phage lyase compound powder prepared in the above example in a 4 mL EP tube, and take 1 g of sterilized diatomaceous earth in a 4 mL EP tube as a control;

[0030] 2. The experimental group and the control group were respectively added with 3 mL of PBS;

[0031] 3. Add dilution to 10 in the experimental group and the control group respectively -5 200μL of the bacterial solution to be tested;

[0032] 4. After sufficient shaking, incubate in a 37°C water bath for 30 minutes, shaking every 5 minutes;

[0033] 5. After incubation, take 300 μL of double-layer medium for detection, and invert at 37°C for 12 hours to count colonies;

[0034] The components of the lower solid medium are: NaCl 10.0g / L, tryptone 10.0g / L, yeast powder 5.0 g / L, agar powder 15.0 g / L;

[0035] The components of the upper semi-solid medium are: NaCl 10.0g / L, tryptone 10.0g / L, yeast pow...

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Abstract

The invention relates to phage lyase composite powder. The composite powder composition comprises the following components in percentage by mass: 0.15%-0.2% of phage lyase TSPpgh, 0.15%-0.2% of phagelyase MMPpgh, 0.3%-0.5% of trehalose, 0.1%-0.2% of citric acid, 0.2%-0.3% of mannitol, 0.2%-0.3% of vitamin C and 98.3%-98.9% of diatomite. The phage lyase composite powder is subjected to in-vitro antibacterial activity detection by adopting a two layer plating method. After the composite powder is stored for 12 months at normal temperature, the enzyme activity is kept at 95% or above (the enzymeactivity loss is within 5%). The composite lyase preparation can be used for replacing antibiotics to inhibit proliferation of harmful pathogenic bacteria such as escherichia coli, salmonella and staphylococcus aureus in animal intestines, and can maintain intestinal microecological balance.

Description

technical field [0001] The invention belongs to the technical field of enzyme preparations, and in particular relates to a composite powder of phage lyase and its preparation method and application. Background technique [0002] Phage lyase is a type of cell wall hydrolytic enzyme that can hydrolyze peptidoglycan. After phage infects the host, the double-stranded DNA phage destroys the bacterial cell wall structure through the holin-lyase cleavage system, thereby causing the host to rupture. Lyases generally have two to three domains involved in the catalysis and binding of substrates. As a new type of bactericidal agent, the high affinity of lyase is related to the species-specific cell wall sugar, and the latter is a necessary component for the survival of bacteria, so it is difficult for bacteria to develop resistance to lyase. In nature, almost all All bacteria have corresponding phages, so the development and research of lyase is of great value. There are two ways for...

Claims

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Application Information

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IPC IPC(8): C12N9/88A23K20/189A23K50/75
CPCA23K20/189A23K50/75C12N9/88
Inventor 林连兵张关令熊燕嵇歆彧蔡赛波邓先余王峰张棋麟郭军
Owner KUNMING UNIV OF SCI & TECH
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