Marine streptomyces griseorubens HN60 and application thereof

A technology of marine actinomycetes and actinomycetes, which is applied in the direction of preparation methods of bacteria and peptides, biochemical equipment and methods, etc., which can solve the biological activities of complex structures, increase the demand for new drugs, and increase the drug resistance of pathogenic microorganisms, etc. problems to achieve good antitumor activity

Active Publication Date: 2020-10-09
QUFU NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The increase in the drug resistance of pathogenic microorganisms has increased the demand for new drugs, and chemical synthesis cannot completely solve the demand for new drugs. Microbial secondary metabolites often have the characteristics of complex structure and strong biological activity. Therefore, microbial secondary metabolites Metabolites as source of drug lead compounds

Method used

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  • Marine streptomyces griseorubens HN60 and application thereof
  • Marine streptomyces griseorubens HN60 and application thereof
  • Marine streptomyces griseorubens HN60 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Weigh 1 g of sample, dissolve it in 9 mL of sterile water, and shake on a shaker at 28 °C for 30 min. Take the upper layer of soil suspension for gradient dilution, respectively diluted to 10 -1 ~10 -3 100 uL from each gradient was applied to Gao's No. 1 medium, YMG medium, HVA medium, and sodium propionate medium (all media were added with 25 ug / mL of naphthyridone before inverting the plate) acid to inhibit fast-growing bacteria, especially Gram-negative bacteria and 75 ug / mL of potassium dichromate to inhibit the growth of bacteria and fungi), pick a single colony for further purification, and isolate the culture medium for a single colony Gao's No. 1 medium. Streaking was performed 3 times in a row to purify a single colony to obtain strain HN60.

[0036] 16S rDNA Identification of Marine Actinomycetes HN60: 16S rDNA Sequence Amplification and Sequencing: Using Positive Bacterial Genome Kit to Extract Genomic DNA, Primer Sequence

[0037] Upstream: 5'-AGAGTTTGAT...

Embodiment 2

[0041] Fermentation of marine Streptomyces flatus HN60 and the acquisition of crude extract: the composition of the medium used is: malt extract powder 10 g, glucose 4 g, yeast extract powder 4 g, natural seawater 1L, pH 7.2~7.4. Streptomyces grivis was inoculated in YMG liquid medium according to the inoculum amount of 8%, and cultured at 30°C for 6 days. After the cultivation, the culture solution was centrifuged to remove the bacteria, and the supernatant was extracted with ethyl acetate until the ethyl acetate phase was in a colorless state, the ethyl acetate phase was collected, filtered with absorbent cotton, and rotary evaporated to obtain the crude extract I. The crude extract I was dissolved in 95% methanol and then extracted with an equal volume of petroleum ether until the petroleum ether phase was colorless and the extraction was completed. The methanol phase was removed by rotary evaporation to obtain the final crude extract. A total of 5.3 g was obtained.

Embodiment 3

[0043]Separation and purification of secondary metabolites: Weigh 60 g of petroleum ether-swelled silica gel and pack it into a column. Dissolve 5.3 g of crude extract in 5 mL of dichloromethane and load the sample. After loading, the sample is eluted. The specific eluent is: Petroleum ether-ethyl acetate (500 mL-500 mL), petroleum ether-dichloromethane-ethyl acetate (130 mL-130 mL-130 mL), petroleum ether-dichloromethane-ethyl acetate (300 mL-600 mL-600 mL), dichloromethane-ethyl acetate (100mL-200 mL), dichloromethane-methanol (100mL-200mL). Use Erlenmeyer flasks for collection, and collect one bottle per 100 mL. The collected solution was tested by silica gel thin layer chromatography, and the components with the same test results were combined. Finally, Fr.1 (0.3 g), Fr.2 (4.3 g), and Fr.3 (0.7 g) were obtained.

[0044] Dissolve Fr.2 (4.3 g) in 4 mL of acetonitrile, and then load the sample for reverse-phase silica gel column chromatography with gradient elution of acet...

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Abstract

The invention belongs to the field of marine actinomycete resource development and utilization, and particularly relates to marine streptomyces griseorubens HN60 and an application thereof. The marinestreptomyces griseorubens HN60 is used as a production strain of actinomycin V and actinomycin D, a pH value of the culture medium is 7.2-7.4, and a culture medium comprises 10g of malt extract powder, 4g of glucose, 4g of yeast extract powder and 1L of natural seawater. The extracted actinomycin V and actinomycin D have good anti-staphylococcus aureus activity.

Description

technical field [0001] The invention belongs to the field of actinomycete resource utilization, and relates to marine streptomyces griseus HN60 and its application, in particular to a marine streptomyces HN60 and its application in the preparation of anti-tumor active substances. Background technique [0002] Cytobacteria belong to the phylum Actinomycetes, are Gram-positive bacteria, and are rarely Gram-negative bacteria. They have a high (G+C)% DNA content and a complex life cycle. Due to the diversity of secondary metabolites and rich biological activities, actinomycetes are often considered as important microbial resources. Studies have shown that among the more than 30,000 microbial secondary metabolites found so far, more than 40% are produced by actinomycetes, and 70% of the known natural antibiotics are produced by actinomycetes, and some actinomycetes produce Secondary metabolites have been used in real life, such as vancomycin used in clinical medicine, Jinggangmy...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P21/02C07K7/06C07K1/20C07K1/16C07K1/14A61P35/00C12R1/465
CPCC07K7/06A61P35/00C12R2001/465C12N1/205C12P21/02
Inventor 杨革孙萍车程川刘金锋巩志金
Owner QUFU NORMAL UNIV
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