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Preparation method and application of actinomycin d

A technology of actinomycin and actinomycetes, applied in the field of preparation of actinomycin D, to achieve the effect of broad-spectrum and high-efficiency antagonistic activity

Active Publication Date: 2020-08-04
TARIM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently registered antibiotics for the prevention and treatment of Verticillium wilt have not been reported

Method used

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  • Preparation method and application of actinomycin d
  • Preparation method and application of actinomycin d
  • Preparation method and application of actinomycin d

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The source and isolation culture of embodiment one Streptomyces mutans TRM45540

[0022] From the 0-30cm soil samples in the ancient city of Loulan, Xinjiang (1400m above sea level, 89 ° 22' 22 " in east diameter, 40 ° 29' 55 " in north latitude), adopt the coating plate method, with 5%, 10%, 15%, 20 % and 25% sodium chloride in 8 kinds of media, 28 ℃ constant temperature culture, a total of 35 strains of actinomycetes of 7 genera were isolated, and finally separated and screened out The biocontrol bacteria TRM45540 with the best antibacterial effect.

Embodiment 2

[0023] The identification of embodiment two Streptomyces mutans TRM45540

[0024] In order to further confirm the taxonomic status of the biocontrol bacteria TRM45540, the present invention uses traditional classification methods and molecular classification methods to classify and identify the strain.

[0025] 1 Culture medium for morphology observation

[0026] Inorganic salt starch agar (ISP4) medium: starch 10g L -1 ,(NH 4 ) 2 SO 4 4g·L -1 ,MgSO 4 2g·L -1 ,K 2 HPO 4 2g·L -1 ,CaCO 3 1g·L -1 ,NaCl 40g·L -1 , agar 18g·L -1 , pH 7.2-7.4.

[0027] 2 primers

[0028] Synthesized by Aoke Biotechnology (Beijing) Co., Ltd. using actinomycetes universal primers. The sequences are: 27F (5'-AGAGTTTGATCCTGGCTC-3') and 1492R (5'-CGGCTACCTTGTTACGACTT-3')

[0029] 3 test method

[0030] 3.1 Morphological observation

[0031] Ordinary microscope observation: using the insert method, cultivate the biocontrol bacteria TRM45540 at 28°C. Use an optical microscope to observe...

Embodiment 3

[0052] Example 3 Utilize Streptomyces mutans TRM45540 to ferment and prepare Actinomycin D

[0053] Inoculate a small amount of mycelium in the ISP4 medium, cultivate at 28°C for 4 days, and obtain the seed liquid of Streptomyces vulbilis TRM45540, take 2mL (8% inoculum size) of the seed liquid and inoculate it in the corn flour fermentation medium for fermentation and cultivation. The fermentation conditions It is: rotation speed 180rpm, initial pH 7.2-7.4, liquid volume 250mL, fermentation temperature 28°C, fermentation 7d. Collect the fermentation broth, filter the fermentation broth with four layers of gauze, and load the sample onto the macroporous resin chromatographic column by wet method (fill 2.0kg of D101 or AB-8 macroporous resin per 100L of fermentation filtrate); then use 2 times the column volume of water, 10% ethanol solution, 30% ethanol solution, 50% ethanol solution, 70% ethanol solution, 95% ethanol solution elution; combined 70-95% ethanol eluate, concentra...

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Abstract

The invention discloses a new actinomycetes strain producing actinomyces D, Streptomyces mutabilis TRM45540, which was preserved in the U.S. Department of Agriculture Agricultural Research Service (ARS) Culture Collection on April 14, 2014, with a preservation number It is NRRL B-59117; also announced the seed culture medium and fermentation medium that utilize this bacterial strain to efficiently produce actinomycin D and its preparation process for extracting and isolating actinomycin D from fermentation liquid; Antibacterial activity test shows that the Bacteria and the actinomycin D produced by its fermentation have broad-spectrum and high-efficiency antagonistic activity against pathogenic bacteria such as Verticillium wilt of cotton, Pythium walnut, Staphylococcus aureus, Staphylococcus epidermidis, Candida albicans, etc. The safe production of crops and the prevention and control of diseases such as cow mastitis and women's vaginitis have important application value and good application and development prospects.

Description

technical field [0001] The invention relates to the fields of agriculture and medicine, in particular to a new application of actinomycin D, a new bacterial strain capable of producing actinomycin D and a preparation method for producing actinomycin D by using the bacterial strain. Background technique [0002] The two strains, Smelanochromogenes No.1779 and Streptomyces parvullus, can produce actinomycetes D. Their main mechanism of action is to hinder the function of RNA polymerase and inhibit the synthesis of RNA, especially mRNA, thereby hindering protein synthesis and inhibiting tumor growth. Combined use of radiation therapy can improve the sensitivity of tumors to radiation therapy. However, the effect of actinomycin D on plant pathogens of cotton Fusarium wilt, walnut pythium and the pathogenic bacteria of women's vaginitis, Candida albicans, has not been reported yet. [0003] Cotton fusarium wilt and cotton verticillium wilt are the most important diseases that har...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P21/02A01P3/00A61P31/10A61P31/04C12R1/465
CPCC12P21/02C12N1/205C12R2001/465
Inventor 张利莉万传星罗晓霞吕玲玲仼晓镤夏占峰
Owner TARIM UNIV
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