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Preparing and regenerating method of tetracycline producing strain protoplast

A protoplast and tetracycline technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, bacteria, etc., can solve the problems of low mutation rate of strains and poor breeding effect, and achieve the effect of increasing the mutation rate of strains

Active Publication Date: 2014-08-27
NORTH CHINA PHARMA COMPANY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a preparation and regeneration method of protoplasts of tetracycline-producing bacteria, so as to solve the problems of low strain mutagenesis rate and poor breeding effect in existing traditional breeding techniques

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] a. Mycelium cultivation:

[0035] Make a spore suspension from the slant spores of the tetracycline-producing bacteria (Streptomyces aureofaciens), inoculate it in the mycelium medium, and the inoculation amount is not less than 10 6 cells / ml, cultured with shaking at 30°C for 26 hours to obtain a mycelium culture solution, the mycelium was inspected under a microscope and showed a net shape, and the supernatant was discarded by centrifugation (3500 rpm, 10 minutes) to obtain mycelium.

[0036] Among them, the strains of tetracycline slant spores are from the Chinese General Catalog of Microbial Strains, and the strain preservation number is CPCC220020.

[0037] The mycelium medium is prepared from distilled water, and based on the total volume of 100ml mycelium medium, it contains the following components: glucose 1g, peptone 0.4g, yeast extract 0.4g, dipotassium hydrogen phosphate 0.4g, dihydrogen phosphate Potassium 0.2g, magnesium sulfate 0.05g, glycine 1g. Steril...

Embodiment 2

[0044] a. Mycelium cultivation:

[0045] Make a spore suspension from the slant spores of the tetracycline-producing bacteria (Streptomyces aureofaciens), inoculate it in the mycelium medium, and the inoculation amount is not less than 10 6 cells / ml, cultured with shaking at 30°C for 26 hours to obtain a mycelium culture solution, the mycelium was inspected under a microscope and showed a net shape, and the supernatant was discarded by centrifugation (3500 rpm, 10 minutes) to obtain mycelium.

[0046] Among them, the strains of tetracycline slant spores are from the Chinese General Catalog of Microbial Strains, and the strain preservation number is CPCC220020.

[0047] The mycelium medium is prepared from distilled water, and based on the total volume of 100ml mycelium medium, it contains the following components: glucose 1g, peptone 0.4g, yeast extract 0.4g, dipotassium hydrogen phosphate 0.4g, dihydrogen phosphate Potassium 0.2g, magnesium sulfate 0.05g, glycine 1g. Steril...

Embodiment 3

[0054] a. Mycelium cultivation:

[0055] Make a spore suspension from the slant spores of the tetracycline-producing bacteria (Streptomyces aureofaciens), inoculate it in the mycelium medium, and the inoculation amount is not less than 10 6 cells / ml, cultured with shaking at 30°C for 26 hours to obtain a mycelium culture solution, the mycelium was inspected under a microscope and showed a net shape, and the supernatant was discarded by centrifugation (3500 rpm, 10 minutes) to obtain mycelium.

[0056] Among them, the strains of tetracycline slant spores are from the Chinese General Catalog of Microbial Strains, and the strain preservation number is CPCC220020.

[0057] The mycelium medium is prepared from distilled water, and based on the total volume of 100ml mycelium medium, it contains the following components: glucose 1g, peptone 0.4g, yeast extract 0.4g, dipotassium hydrogen phosphate 0.4g, dihydrogen phosphate Potassium 0.2g, magnesium sulfate 0.05g, glycine 1g. Steril...

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Abstract

The invention aims to provide a preparing and regenerating method of a tetracycline producing strain protoplast. The method comprises the following steps: (a) preparing a tetracycline slant culture-medium; (b) preparing a spore suspension; (c) preparing a mycelium culture-medium; (d) preparing a regeneration medium; (e) culturing mycelium; (f) enzymatically hydrolyzing a cell wall; (g) separating a protoplast; and (h) regenerating the protoplast. A novel method for breeding tetracycline strains, provided by the invention, has the advantage as follows: problems that the tetracycline protoplast bacterial colony cannot be regenerated, the spore generating capacity of the tetracycline protoplast bacterial colony is low and the passage of the tetracycline protoplast bacterial colony cannot be made are solved.

Description

technical field [0001] The invention relates to a method for preparing and regenerating microbial protoplasts, in particular to a method for preparing and regenerating protoplasts of tetracycline-producing bacteria. Background technique [0002] Tetracycline (Tetracycline, TC) is a broad-spectrum antibiotic that inhibits bacterial protein synthesis. It is named for its molecular structure containing a tetracene basic skeleton. It is mainly used for infections of various Gram-positive cocci and Gram-positive bacilli. It is widely used in medicine, animal husbandry and agriculture. my country has started industrial production of tetracycline since the 1960s, and it has a history of more than 50 years. The tetracycline-producing bacteria are Streptomyces in actinomycetes. At present, the breeding of its excellent strains mainly adopts traditional breeding methods. After long-term artificial mutation breeding and natural separation, the strains have formed a high production ca...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12R1/485
Inventor 张彩霞张曦
Owner NORTH CHINA PHARMA COMPANY
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