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Plant endophytic fungus Eupenicillium brefeldianum F4a and its application

A plant endophytic fungus and Penicillium technology, applied in the field of microorganisms, can solve problems such as low BFA yield, and achieve the effects of reducing treatment and pollution, simple and efficient extraction and separation, and good industrialization prospects.

Active Publication Date: 2015-09-02
SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the yield of BFA produced by microbial fermentation reported so far is relatively low.

Method used

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  • Plant endophytic fungus Eupenicillium brefeldianum F4a and its application
  • Plant endophytic fungus Eupenicillium brefeldianum F4a and its application
  • Plant endophytic fungus Eupenicillium brefeldianum F4a and its application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] The identification of embodiment 1 plant endophyte Penicillium brescens F4a

[0030] Plant endophytic fungus F4a was preserved in the China Center for Type Culture Collection on December 18, 2012. The address of the preservation unit is China. Wuhan. Wuhan University. The preservation number is CCTCC M2012531. The taxonomic name is Penicillium brezheng F4a ( Eupenicillium brefeldianum F4a).

[0031] Strain F4a grows well on PDA and Martin's medium; the early colonies are pure white, and gradually the center of the colony turns yellow-green; the surface of the colony is fluffy, without exudate and soluble pigment; after maturity, the colony has radial folds, and the center Slightly raised, forming a round plaque with a distinct color, yellowish on the back. Under the microscope, there is no enlarged apical capsule at the top, and the conidia structure is rare, and the spores are born on the conidiophores in a broom shape, and the conidia on the top of the small stalks a...

Embodiment 2

[0035] Example 2 Fermentation of strain F4a to prepare BFA

[0036] 2.1 Preparation of seed solution: first inoculate the strain F4a on the prepared PSA medium plate by streaking method, and culture at a constant temperature of 28°C for 5 days; In a 250mL Erlenmeyer flask filled with 50mL PSB medium, culture on a constant temperature shaker at 180rpm at 28°C for 72h;

[0037] The PSB medium is: 20.0 g sucrose, 200 g potato extract, balanced to 1 L with distilled water, pH 5.0-8.0, sterilized at 115° C. for 30 min.

[0038] PSA is a solid medium with 18 g of agar added per liter of PSB.

[0039] 2.2 Fermentation in large bottles: use a 3L Erlenmeyer flask, fill each bottle with 0.70L of PSB medium, and sterilize at 115°C for 30 minutes before use. 50mL of inoculated seed solution per bottle, cultivated in a constant temperature shaker at 28°C and 180rpm for 7-13d, and fermented 10L in total;

[0040] 2.3 Obtaining crude extract: filter the obtained fermentation broth with 4-...

Embodiment 3B

[0043] The structural analysis of embodiment 3BFA

[0044] BFA is colorless crystals, ESI-MS gives ( image 3 ) m / z303.2[M+Na] + (Calcd. for C 16 h 24 o 4 Na, 303.1572) deduced that its molecular formula is C 16 h 24 o 4 . exist 1 H-NMR (600MHz, CD 3 OD) spectra (Table 1 and Figure 4 ), the low field region δ H 7.46 (1H, dd, J=15.5, 2.2Hz) and 5.82 (1H, d, J=15.5Hz) form a trans double bond, and one end is connected to the quaternary carbon; δ H 5.27 (H, dd, J=15.1, 9.7Hz) and 5.77 (1H, ddd, J=15.1, 9.7, 4.4Hz) form another trans double bond, and the two ends are respectively connected to CH and CH 2 group. Hydrogen δ on 3 oxygenated carbons H 4.04 (1H, brd, J=9.1Hz), 4.80 (1H, m) and 4.21 (1H, m) and the signal δ of a methyl proton attached to CH H 1.24 (3H, d, J=6.2Hz). 13 C-NMR (150MHz, CD 3 OD) gives 16 carbon signals (Table 1 and Figure 5 ), where one ester carbonyl carbon signals δ C 166.96; 4 unsaturated carbon signals δ C 153.69, 136.70, 129.99, 1...

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Abstract

The invention relates to the technical field of microbes, and concretely relates to a new plant endophytic fungus and its application in the preparation of brefeldin A (BFA). The plant endophytic fungus is Eupenicillium brefeldianum F4a, and is preserved in China Center for Type Culture Collection on Dec. 18, 2012 with the preservation number of CCTCC M2012531. The Eupenicillium brefeldianum F4a can generate high content BFA, and the highest fermentation output reaches 1.9g / L. The invention also provides a method for preparing the highly pure BFA through HP20 solid phase extraction and rapid silica gel column chromatography or a pure crystallization technology. The method has the advantages of low production cost, great reduction of treatment and pollution of organic matters, and good industrial prospect. The BFA prepared in the invention has antifungal and insecticidal activity, and is an ideal veterinary or agricultural candidate drug.

Description

technical field [0001] The invention relates to the technical field of microbes, in particular to a plant endophytic fungus Penicillium breenii F4a and its application. Background technique [0002] Plant endophytes refer to a class of microorganisms that survive part or all of their life cycle inside healthy plant tissues without causing the host plant to show obvious symptoms of infection (at least temporarily). In the long-term evolution process, it has formed a specific mutually beneficial symbiotic relationship with the host, its special habitat, and its growth and metabolism are restricted by many factors such as the host and external environmental factors. Nowadays, the important physiological and ecological functions played by endophytes and their huge application potential in the fields of agriculture, forestry and medicine have gradually become a research hotspot at home and abroad. Therefore, the abundant and diverse plant endophytes have become an important sour...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12P17/08C07D313/00A01N43/22A01P3/00A01P7/00A61K31/365A61P31/10C12R1/645
Inventor 潘华奇胡江春王书锦
Owner SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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