Production method of high-titer phages

A production method and phage technology, applied in the fields of biotechnology and microbial products, can solve the problems of complex phage production mode conditions, restricted phage development, and poor production effect, and achieve the effects of convenient application, high production efficiency, and reasonable components.

Inactive Publication Date: 2016-11-09
SUZHOU IRIVET BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, in the above-mentioned process, it is necessary to expand the production of phages. The production methods of phages in the prior art have complex conditions, and are often inefficient, and the production effect is not good, which seriously restricts the development of phage applications.

Method used

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Examples

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Embodiment 1

[0029] A method for producing high-titer lactobacillus phage, the steps are completed under 4% humidity conditions, comprising the following steps:

[0030] 1) The frozen Lactobacillus phage mother liquid is added to the Lactobacillus bacterial liquid according to the volume ratio of 1:1, and mixed evenly; the titer of the phage in the phage mother liquid is 10 6 PFU / mL;

[0031] 2) Shake the above mixture at 32°C for 10 minutes at a speed of 80 r / min;

[0032] 3) Spread the mixture in step 2) evenly on the solid medium, and incubate it upside down at 32°C for 8 hours; the solid medium includes the following components by weight: 0.8 parts of tryptone, yeast extract 0.8 parts of powder, 1 part of agarose, 0.3 parts of lactose, 0.3 parts of sodium chloride, 0.1 parts of potassium chloride, 0.5 parts of magnesium sulfate, 0.1 parts of disodium hydrogen phosphate and 100 parts of deionized water;

[0033] 4) Dissolve a single colony on the solid medium in 1 mL of phage buffer, ...

Embodiment 2

[0040] A method for producing high-titer Vibrio alginolyticus bacteriophage, the steps are completed under 6% humidity conditions, comprising the following steps:

[0041] 1) The frozen Vibrio alginolyticus phage mother solution was added to the Vibrio alginolyticus bacterial solution according to the volume ratio of 1:2, and mixed evenly; the titer of phage in the phage mother solution was 10 12 PFU / mL;

[0042] 2) Shake the above mixture at a temperature of 40°C for 20 minutes at a speed of 120 r / min;

[0043] 3) Spread the mixture of step 2) evenly on the solid medium, and incubate it upside down at 40°C for 16 hours; the solid medium includes the following components by weight: 1.2 parts of tryptone, yeast extract 1.5 parts of powder, 2 parts of agarose, 0.6 parts of lactose, 0.8 parts of sodium chloride, 0.5 parts of potassium chloride, 2 parts of magnesium sulfate, 0.3 parts of disodium hydrogen phosphate and 100 parts of purified water;

[0044]4) Dissolve a single co...

Embodiment 3

[0051] A method for producing high-titer mycobacteriophage, the steps are completed under 5% humidity conditions, comprising the following steps:

[0052] 1) The frozen Mycobacterium phage mother solution is added to the Mycobacterium bacterial solution according to the volume ratio of 1:1.5, and mixed evenly; the titer of the phage in the phage mother solution is 10 9 PFU / mL;

[0053] 2) Shake the above mixture at 37°C for 15 minutes at a speed of 100 r / min;

[0054] 3) Spread the mixture in step 2) evenly on the solid medium, and incubate it upside down at 37°C for 10 hours; the solid medium includes the following components by weight: 0.9 parts of tryptone, yeast extract 1 part of powder, 1.5 parts of agarose, 0.5 parts of lactose, 0.6 parts of sodium chloride, 0.3 parts of potassium chloride, 0.9 parts of magnesium sulfate, 0.2 parts of disodium hydrogen phosphate and 100 parts of deionized water;

[0055] 4) Dissolve a single colony on the solid medium in 0.8 mL of phag...

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Abstract

The invention provides a production method of high-titer phages. In a production process, an OD value of a phage culture solution is detected timely; a host bacterium solution of the phage, when cultivated to a logarithmic phase that proliferative activity is maximized, is kept and is subjected to enlarge cultivation; and centrifugal cracking and filtering are conducted to remove bacteria, so that high-titer phage liquid is obtained in a large scale. The production method of the high-titer phage disclosed by the invention is reasonable, convenient for application and high in production efficiency, and the produced high-titer phages are high in titer and good in thermal stability.

Description

technical field [0001] The invention belongs to the field of biotechnology and microbial products, and in particular relates to a production method of high titer phage. Background technique [0002] Phage is a general term for a class of viruses that can infect microorganisms such as bacteria, actinomycetes, and spirochetes. No toxicity to animals and plants. With the development of molecular biology technology, phage has been widely used in many fields such as disease diagnosis and water treatment. Phages have also become important molecular and genetic research tools due to their ability to insert genes into host DNA. Using phages, many ingenious experiments can be designed. [0003] However, in the above process, it is necessary to expand the production of phages. The production methods of phages in the prior art have complex conditions, and often have low efficiency and poor production effect, which seriously restricts the development of phage applications. Contents...

Claims

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Application Information

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IPC IPC(8): C12N7/00
CPCC12N7/00C12N2795/00011
Inventor 许维素宋增福陈彪
Owner SUZHOU IRIVET BIOTECH
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