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Method for promoting accumulation of triterpene in betula platyphylla suk. suspension cell by utilizing endophytic fungi elicitor

A fungal elicitor and endophytic fungus technology, applied in the field of bioengineering, to achieve the effects of considerable yield, short cycle and simple reproduction and cultivation

Inactive Publication Date: 2010-09-08
NORTHEAST FORESTRY UNIVERSITY +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report on the use of fungal elicitors to increase the content of triterpenes in birch cells

Method used

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  • Method for promoting accumulation of triterpene in betula platyphylla suk. suspension cell by utilizing endophytic fungi elicitor
  • Method for promoting accumulation of triterpene in betula platyphylla suk. suspension cell by utilizing endophytic fungi elicitor
  • Method for promoting accumulation of triterpene in betula platyphylla suk. suspension cell by utilizing endophytic fungi elicitor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1. screening significantly improves the endophytic fungus of birch triterpenes

[0031] The hyphae of 59 kinds of endophytic fungi isolated from white birch bark were fermented and cultured in PDA liquid medium for 7 days, and then the harvested bacterial liquid and bacterial cells were homogenized and then sterilized at 121°C for 20 minutes. The final concentration of sugar was 60 μg / ml was prepared as elicitor. Add fungal elicitors on the 8th day of birch suspension cell growth, treat 3 flasks of suspension cells with elicitor bacteria prepared by each strain, harvest the cells after 3 days of induction, and measure the dry weight of the cells and the content of triterpenes in the cells.

[0032] Birch suspension cells grown for 10 weeks were inoculated in B5 medium with an inoculum volume of 4% (W / V). The inoculated birch suspension cells were cultured at 24-26°C under 1500-2000 Lux light. On the 8th day of cell culture, elicitors prepared from 59 kinds o...

Embodiment 2

[0033] Example 2. Optimizing the induction effect of the polysaccharide elicitor prepared by screening strains

[0034] Birch suspension cells grown for 10 weeks were inoculated in B5 medium with an inoculum volume of 4% (W / V). The birch suspension cells inoculated were cultured at 24-26°C under 1500-2000 Lux light, and three concentrations ( 40 μg / ml, 100 μg / ml, 400 μg / ml) BE43 fungal polysaccharide elicitor, and the cells were harvested after induction and culture for 1d, 2d, and 3d, and the harvested cells were dried at 60°C and weighed. The content of total triterpenes in the cells was determined colorimetrically with betulin as a standard and vanillin-glacial acetic acid solution as a dye. The results showed that after adding different concentrations of fungal polysaccharide elicitors, the dry weight of the cells decreased to varying degrees, while the content of triterpenes increased to varying degrees. Among them, the concentration of 40 μg / ml elicitor was added on th...

Embodiment 3

[0035] Example 3. Optimizing the induction effect of the protein elicitor prepared by screening strains

[0036]Birch suspension cells grown for 10 weeks were inoculated in B5 medium with an inoculum volume of 4% (W / V). The inoculated birch suspension cells were cultured at 24-26°C under 1500-2000 Lux light, and on the 8th day of cell culture, fungal protein elicitors with a concentration of 40 μg / ml, 80 μg / ml, and 120 μg / ml were added, respectively, on the 1d of induction culture. , 2d, and 4d were harvested, and the harvested cells were dried at 60°C and weighed dry weight. The content of total triterpenes in the cells was determined colorimetrically with betulin as a standard and vanillin-glacial acetic acid solution as a dye. The results showed that after adding different concentrations of mycoprotein elicitors, the dry weight of cells and the content of triterpenes were increased to varying degrees. Among them, the concentration of 80 μg / ml fungal protein elicitor was a...

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Abstract

The invention provides a method for promoting accumulation of triterpene in betula platyphylla suk. Suspension cell by utilizing endophytic fungi elicitor. The method utilizes endophytic fungi separated from betula latyphylla suk. bark, the endophytic fungi is prepared into fungi elicitor, appropriate concentration of fungi elicitor is added in appropriate stage of betula platyphylla suk. suspension cell growth, after induction is carried out for a period of time, cells are collected, and endophytic fungi capable of effectively improving accumulation of triterpene is screened out as phomopsis sp. On the basis, induction effect of polysaccharide elicitor and protein elicitor prepared by the fungi is further optimized; 40Mug / ml of polysaccharide elicitor is added on the eighth day of betula platyphylla suk. cell culture, triterpene content in cell after induction for one day reaches up to 29.47mg / g, being 1.78 times that of the control group; 80Mug / ml of fungus protein elicitor is added on the eighth day of betula platyphylla suk. cell culture, induction culture is carried out for four days, and triterpene content in cell reaches up to 34.49mg / g, being 2.47 times that of the control group. In the invention, the type and concentration of the added elicitor, growth stage of betula platyphylla suk. suspension cell added with appropriate elicitor and induction time of elicitor are determined, induction technology of effectively improving accumulation of triterpene in betula platyphylla suk. suspension cell is formed, and important theory and technical basis are laid for betula platyphylla suk. triterpene production by utilizing cell engineering technology, thus the invention has wide application potential and promotion prospect.

Description

Technical field: [0001] The invention relates to a practical technique for increasing triterpene content in white birch suspension cells, which belongs to the technical field of bioengineering. Background technique: [0002] White birch (Betulaplatyphylla Suk.) is a deciduous tree belonging to the genus Betula Linn of the family Betulaceae. It is mainly distributed in Japan, Korea, Russia, China and other Eurasian continents in temperate Asia. As early as recorded in "Compendium of Materia Medica", birch bark can be used for the treatment of jaundice, mastitis, scabies and other diseases. Studies in recent years have shown that one of the main active components of birch bark is triterpenoids, accounting for about 25% to 30% of the dry weight of the bark. Betula triterpenoids have antibacterial, antiviral, antitumor, lipid-lowering, choleretic and hepatoprotective effects (Kessler et al., 2007), especially triterpenoids such as betulin and betulinic acid are used as natural ...

Claims

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Application Information

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IPC IPC(8): C12P33/00C12P1/00C12N1/14C12R1/645
Inventor 詹亚光范桂枝翟俏丽李晓灿齐凤慧曾凡锁由香玲
Owner NORTHEAST FORESTRY UNIVERSITY
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