Engineering bacterium for generating carbamoyl tobramycin and application thereof

A carbamoyl tobramycin and engineering bacteria technology, applied in bacteria, microorganisms, biochemical equipment and methods, etc., can solve problems such as pollution, environmental pollution, difficulties, etc., to reduce environmental pollution, reduce production costs, and simplify The effect of the production process

Inactive Publication Date: 2012-03-14
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because the structures and physicochemical properties of abramycin and kanamycin B are similar to carbamyl tobramycin, it is very necessary to separate the three components one by one from the fermentation broth in industrial production. difficulty
Not only the chromatographic

Method used

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  • Engineering bacterium for generating carbamoyl tobramycin and application thereof
  • Engineering bacterium for generating carbamoyl tobramycin and application thereof
  • Engineering bacterium for generating carbamoyl tobramycin and application thereof

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Experimental program
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Effect test

Embodiment 1

[0029] Source of reagents: pMD19-T and restriction endonucleases were purchased from TaKaRa Company, plasmid pAGe (containing resistance gene ermE) was donated by Dr. Shao Lei of Shanghai Institute of Pharmaceutical Industry, and plasmid pBR322 (containing tetracycline resistance gene) was provided by our laboratory save.

[0030] The present invention mainly comprises following major steps:

[0031] 1. Construct aprH-M gene replacement plasmid

[0032] Using the apramycin (also known as abramycin) biosynthetic gene cluster sequence (GenBank Accession Number AJ629123) published by Piepersberg et al. Breeding of carbamoyl tobramycin high-yielding strains and their fermentation characteristics [J]. Chinese Journal of Antibiotics, 2008, 33 (7): 442-445) Existence of abramycin biosynthetic gene clusters on chromosomal DNA. and then respectively in aprH-M Two pairs of primers SD1 / SD2 and SD3 / SD4 were designed for the upstream and downstream of the gene to extract the tot...

Embodiment 2

[0044] Embodiment 2: Preparation of tobramycin, a metabolite of Streptomyces melanogaster HM106

[0045] The engineering bacterium HM106 provided by the invention can be directly used to produce tobramycin, and the engineering bacterium is fermented to extract metabolites to prepare tobramycin as an antibacterial drug.

[0046] 1. Fermentation and cultivation of Streptomyces melanogaster HM106 engineering bacteria

[0047] Seed medium: glucose 5g, cornstarch 10g, soybean meal 15g, potassium dihydrogen phosphate 0.5g, potassium chloride 1g, calcium chloride 0.25g, magnesium sulfate 5g, add tap water to 1L.

[0048] Fermentation medium: glucose 15g, corn flour 20g, soybean cake powder 35g, magnesium sulfate 11g, calcium carbonate 7g, fish meal 6g, zinc sulfate 0.1g, ammonium sulfate 7g, cornstarch 20g, amylase 0.5g, add tap water to 1L, The pH was adjusted to 7.0-7.2 with sodium hydroxide solution.

[0049] The genetically engineered bacterium Streptomyces melanogaster HM106 o...

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Abstract

The invention relates to an aprH-aprM-inactivation-function engineering bacterium for mainly generating carbamoyl tobramycin and an application thereof, which belong to the technical field of medicine. The engineering bacterium is streptomyces tenebrarius HM106 (delta aprHM) and is registered and preserved by China General Microbiological Center Culture Collection Center on September 23, 2011, the preservation number is CGMCCNo.5246, and the engineering bacterium is used for preparing antibacterial medicine. The engineering bacterium mainly generating the carbamoyl tobramycin is obtained, theproduction process is simplified, the production cost is reduced, and simultaneously, the quality control of products is also convenient.

Description

Technical field [0001] The present invention is an antibiotic pharmaceutical field. It involves an engineering bacteria and its application. Specifically, the invention involves an engineering bacteria that is mainly produced by amino -metharomycin, which is used in antibiotic manufacturing. Background technique [0002] Dark chain mold ( Streptomycess tenebrarius ) It was a 1967 researcher from the American Celery Company being separated from the soil. The strain could produce a set of antibiotics, called Nebramycins, component figure 1 ; Aberla), 4 ( figure 2 ; Ammonia metharomycin B), 5 '( figure 2 ; Ammonia metharomycin) is the main component. When conducting biological research on each factor, these three components have good antibacterial effects, and they are important raw medicines.Over the years, the research of scientific research staff on dark chain molds is mainly concentrated in traditional conventional mutant breeding, genetic engineering breeding, and optimization ...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12P19/26C12R1/465
Inventor 洪文荣严绍德
Owner FUZHOU UNIV
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