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A kind of Pseudomonas fluorescens and its application

A technology of Pseudomonas fluorescens and Pseudomonas acid, applied in the field of microorganisms, can solve problems such as low fermentation level, and achieve the effects of single composition of fermentation products and cost reduction

Active Publication Date: 2015-11-18
ZHEJIANG RUIBANG LAB
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem to be solved by the present invention is to provide a kind of fermentation unit with very high pseudomonic acid A in view of the defect that the fermentation level of pseudomonic acid A produced by the production bacteria of pseudomonic acid A is relatively low. Production bacteria of pseudomonic acid A and its application

Method used

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  • A kind of Pseudomonas fluorescens and its application
  • A kind of Pseudomonas fluorescens and its application
  • A kind of Pseudomonas fluorescens and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The acquisition of embodiment 1 bacterial classification

[0032] 1. Starting strain

[0033] Pseudomonas fluorescens ATCC 49323 (purchased from ATCC).

[0034] 2. The cultivation process of bacteria

[0035] The starting strain Pseudomonas fluorescens was inoculated on the slant medium, cultured at 30°C for 3 days, washed with sterile water, inoculated on the seed medium at 1% (v / v) inoculum, and cultured at 28°C for 20 Hours, inoculate the fermentation medium at 2% (v / v) inoculum, and culture at 28°C for 4-5 days. Different stages of seed culture can be set according to different fermentation culture scales.

[0036] 3. HPLC analysis conditions of monocytic acid A:

[0037] The fermentation broth was taken, centrifuged at 12000rpm for 10min, and the supernatant was taken for HPLC analysis.

[0038] Mobile phase: 50mmol / L sodium dihydrogen phosphate (pH6.3): acetonitrile=75:25.

[0039] Detection wavelength: 229nm

[0040] HPLC column: C18, 250*4.6mm

[0041] F...

Embodiment 2

[0053] Example 2 Morphological and cultural characteristics of Pseudomonas fluorescens CGMCC No.7147

[0054] Pseudomonas fluorescens CGMCCNo.7147 is a gram-negative bacterium that does not produce spores. After being cultured in LB medium for 3 days, the diameter of the colony is between 1-3mm, and it is a round colony with a protruding center. The color of the colony is milky white . According to "Bergey's Handbook of Systematic Bacteriology", the physiological response of the strain is shown in Table 1, indicating that the strain is Pseudomonasfluorescens.

[0055] Table 1. Physiological responses of Pseudomonas fluorescens CGMCCNo.7147

[0056]

[0057]

Embodiment 3

[0058] Example 3 Identification of 16SrDNA of Pseudomonas fluorescens CGMCC No.7147 strain

[0059] Genomic DNA was extracted from Pseudomonas fluorescens CGMCCNo.7147 for 16SrDNA analysis.

[0060] Designed primers, 7F5'–CAGAGTTTGATCCTGGCT-3', and

[0061] 1540R5'–AGGAGGTGATCCAGCCGCA-3';

[0062] Carry out PCR amplification, the composition of PCR reaction system is as follows:

[0063] Reagent

Volume (μl)

Template (genomic DNA20-50ng / μl)

0.5

5×Buffer (including Mg 2+ )

2.5

dNTPs (2.5mM each)

1

7F (10 μM)

0.5

1540R (10μM)

0.5

Taq enzyme

0.5

Add double steamed H 2 O to

25

[0064] The PCR conditions are shown in the table below:

[0065]

[0066] The results of agarose gel electrophoresis of the PCR products are shown in figure 2 . DNA sequencing was performed on the PCR product, and the sequence was compared with the 16SrDNA in the Genbank database. The results are sh...

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Abstract

The invention discloses a pseudomonic acid A producing strain and application thereof. The pseudomonic acid A producing strain is pseudomonas fluorescens, and the preservation number is CGMCC No.7147. The pseudomonas fluorescens has very high pseudomonic acid A fermentability, and the capability of the pseudomonas fluorescens for producing the pseudomonic acid A is many times that of the conventional producing strain, and more than 5 times that of the starting strain; the fermentation unit of the pseudomonas fluorescens can reach more than 2500 mg / L in a 5000 L fermentation tank; moreover, the fermentation product compositions of the pseudomonas CGMCC No.7147 is simple, and has no analogues of the pseudomonic acid A, so that the cost of separating and purifying the pseudomonic acid A from fermentation liquor is reduced greatly.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a Pseudomonas fluorescens and its application. Background technique [0002] Pseudomonic acid A, also known as mupirocin, the chemical name is 9-{4-[5-2,3-epoxy-5-hydroxy-4-methylhexyl]-3,4-di Hydroxytetrahydropyran-2-yl}-3-methylbut-2-enoyloxy}nonanoic acid, the structural formula is shown in Formula I. It is a compound with antibacterial activity isolated from the fermentation culture of Pseudomonas in 1971. [0003] [0004] Pseudomonic acid A acts on the binding point between isoleucine-tRNA synthetase and isoleucine in bacteria, thereby hindering the synthesis of amino acids, consuming tRNA in cells, and making RNA synthesis and protein synthesis The process is stopped, thereby inhibiting the growth of bacteria. Pseudomonic acid A has strong antibacterial activity against Gram-positive bacteria, such as Staphylococcus aureus and Staphylococcus epidermidis. It i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P17/16C12R1/39
Inventor 卜光明
Owner ZHEJIANG RUIBANG LAB
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