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Method for preparing 1,3-propylene glycol coupling bacteriophage produced by microbial fermentation

A technology of microbial fermentation and propylene glycol, applied in the field of bioengineering, can solve the problems of low phage titer, high cost of medium, and low culture density of host cells, so as to avoid the bacteria treatment steps, improve the infection effect, and shorten the infection the effect of time

Active Publication Date: 2020-12-22
DALIAN UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The object of the present invention is to provide a method for producing 1,3-propanediol-coupled phage by microbial fermentation to solve the problems of low host cell culture density, high culture medium cost and low phage titer in the prior art of phage preparation. method

Method used

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  • Method for preparing 1,3-propylene glycol coupling bacteriophage produced by microbial fermentation
  • Method for preparing 1,3-propylene glycol coupling bacteriophage produced by microbial fermentation
  • Method for preparing 1,3-propylene glycol coupling bacteriophage produced by microbial fermentation

Examples

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Effect test

Embodiment 1

[0049] Example 1 Fermentation of 1,3-propanediol

[0050] The fermentation experiment of this embodiment was carried out in a 5L fermenter, the initial fermentation medium volume of the fermenter was 2L, and the inoculum size was 10%. The initial glycerin concentration was 40g / L, and glycerol with a purity of 96% was added during the fermentation process to control the total residual glycerol concentration in the fermenter to be 15-20g / L. The temperature and rotational speed of the fermentation process were controlled at 37° C. and 250 rpm. The pH in the fermentation process was controlled at 7.0 by dropping 5mol / L NaOH solution. The changes of products and biomass during the fermentation process are shown in figure 1 shown. The host cell density reaches the maximum OD at 15 hours of fermentation 650 9.4, the concentration of 1,3-propanediol at the end of fermentation can reach up to 55g / L, and the production intensity is 1.57g / L / h.

Embodiment 2

[0051] The preparation of embodiment 2 phage

[0052] According to the method described in Example 1, using glycerol as a substrate, the host bacteria were obtained by adding glycerol in batches, and a certain amount of host bacteria at different fermentation time points were taken respectively, centrifuged at 10000rpm for 10min, and the supernatant was mainly 1,3 -Products such as propylene glycol, the obtained bacteria sludge is used to mix with the phage suspension in a certain ratio, and the infection index (MOI) is 0.1, and the infection experiment is carried out. The infection experiment was carried out in a 150mL shake flask (working volume: 50mL), the culture medium was a fermentation medium with a glycerol concentration of 40g / L, and the culture conditions were 37°C and 200rpm. After 2.5h, the infection was terminated, and the phage lysate was obtained. The concentration of the lysate, the pH value of the lysate and the titer of the phage in the phage lysate are shown...

Embodiment 3M

[0055] The influence of embodiment 3MOI on phage infection

[0056] The host cells in the final stage of fermentation (35h) in Example 1 were taken for MOI optimization test, and four values ​​of MOI=0.1, 0.05, 0.01, and 0.002 were respectively taken for infection test. OD of lysate obtained from infection 650 See Table 2 for values, pH and phage titers. When the MOI was 0.01, the phage titer was as high as 2.6×10 9 pfu / mL.

[0057] Table 2. Experimental results of infecting host cells at the end of fermentation at different MOIs

[0058]

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Abstract

The invention belongs to the technical field of biological engineering, and provides a method for preparing 1, 3-propylene glycol coupling bacteriophage produced by microbial fermentation. The methodcomprises the following steps of: inoculating klebsiella pneumonia into a fermentation culture medium, and taking glycerin as a substrate to ferment and produce 1, 3-propylene glycol; and taking fermentation broth or the bacterial sludge of the fermentation broth, and carrying out co-culture with a bacteriophage suspension so as prepare the bacteriophage. By use of the method disclosed by the invention, the problem in an existing bacteriophage preparation technology that pathogen host preparation wastes time and money is solved, and the problem of thallus processing and safety of 1, 3-propylene glycol production bacteria can be solved. The method gives consideration to 1, 3-propylene glycol fermentation and bacteriophage preparation, the preparation cost of the bacteriophage can be greatlysolved, worries about future of 1, 3-propylene glycol fermentation are eliminated, and the method disclosed by the invention is a method for preparing 1, 3-propylene glycol and bacteriophage, and hasa good industrial application prospect.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a method for preparing 1,3-propanediol-coupled phage by microbial fermentation. Background technique [0002] Since the 21st century, the problem of bacterial resistance caused by the abuse of antibiotics has become a global public health crisis, and the development of new antibiotics in trouble has led to an increasingly severe situation in clinical antibacterial treatment. According to a 2016 British government report, it is estimated that by 2050, the number of deaths caused by "super bacteria" will increase to 10 million people every year. The treatment of multi-drug-resistant bacteria and pan-drug-resistant bacterial infections is facing a severe situation where no drugs are available, which makes the development of a powerful, large population with indications, significant clinical effect, and unique bactericidal mechanism different from the existing ones. Antimicro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/18C12N1/20C12N7/00C12R1/22
CPCC12P7/18C12N1/20C12N7/00C12N2795/00051
Inventor 修志龙张志荣王晓丽孙亚琴
Owner DALIAN UNIV OF TECH
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