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Plant cell engineering process of producing puerarin

A plant cell, puerarin technology, applied in plant cells, sugar derivatives, organic chemistry and other directions, can solve the problems of inability to expand cultivation, shortage of kudzu resources, inability to meet and other problems, and achieve small changes in effective components, good growth, The effect of meeting the requirements of suspension culture

Inactive Publication Date: 2003-03-19
SOUTH CHINA NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But there have also been many problems: (1) Puerarin and other kudzu drugs used clinically are all derived from wild kudzu plants. Due to the expansion of cities and the extensive exploitation of kudzu resources, kudzu resources are increasingly in short supply, which is far from enough (2) Pueraria thomosonii Benth., the cultivar of Pueraria thomosonii Benth., is widely planted among the people, and its starch content is high, but its medicinal component puerarin content is lower than that of Pueraria lobata, which cannot meet the corresponding requirements. need
Such as adopting the method of Ginkgo or Yunnan Taxus callus suspension culture (Yu Rongmin etc., Ginkgo cell suspension culture and its ginkgolide production, Biological Engineering Journal, 1999 No. 2; Gan Fanyuan, etc., Yunnan Taxus cells Suspension culture of Suspension Culture, Journal of Plant Physiology, 1997 1) to carry out kudzu suspension cell culture, its effect is very poor, as follows: in a very short period of time, the cells aggregate into clusters, browning occurs, easy to die, or melt to death , cannot be expanded at all

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] (1) Induce callus formation in isolated organs: take mature seeds and carry out surface disinfection, place them on wet filter paper for dark germination for 5 days to obtain aseptic seedlings, take the leaves of aseptic seedlings and cut them into 0.3 cm long, and put them in the following formula Cultivate in solid medium for 10 days: MS+0.5mg / L NAA+1.0mg / L6-BA, the culture conditions are temperature (25±2)°C, dark or weak light.

[0024] (2) Subculture of callus: the callus produced in step (1) is moved into the subculture medium of the following formula for subculture: B 5 +1.0mg / L NAA+1.0mg / L6-BA+100mg / L hydrolyzed casein, the culture conditions are the same as step (1), the culture time is 8 days, and the fresh medium is replaced 3 times during the period, the result is that the granules of the callus are constantly Dilated and loosened.

[0025] (3) Suspension culture of callus: prepare the new liquid culture medium of following formula: B 5 +1.0mg / L NAA+1.0mg / ...

Embodiment 2

[0027] Others are with embodiment 1, and the time of dark germination of seed in step (1) is 3 days, and the leaf of aseptic seedling is cut into 1.0 centimeter section, and medium formula is: B 5 +2.0mg / L IAA+0.5mg / LCPPU; medium in step (2) adopts MS+2.0mg / L , 4-D+0.1mg / L KT+200mg / L hydrolyzed casein, and the culture time is 5 days, Replace the fresh medium twice during the period; the newly prepared medium in step (3) adopts MS+2.0mg / L2, 4-D+0.1mg / L KT+1000mg / L hydrolyzed casein, take 30ml mixed medium , the callus inoculation density was 1.5 g per 45 ml, and the culture time was 5 days. The desired puerarin is obtained as a result.

Embodiment 3

[0029] Others are with embodiment 1, in the step (1) with the stem segment of healthy plant as explant, adopt 75% ethanol surface disinfection for 30 seconds, then use 0.1% mercuric chloride surface disinfection for 5 minutes (add surfactant few drops), Medium formula is B 5 +1.0mg / L2,4-D+1.0mg / L TDZ, the culture time is 15 days; the medium in step (2) uses MS+0.5mg / L2,4,5-T+0.5mg / L KT+400mg / L hydrolyzed casein, the culture time is 5 days, during which the fresh medium is replaced once; the newly prepared medium in step (3) is MS+0.5mg / L2,4,5-T+0.5mg / L KT+ 100mg / L chitosan, take 100ml of mixed culture medium, callus inoculation density is 3g per 45ml, culture time is 3 days. The desired puerarin is obtained as a result.

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Abstract

The plant cell engineering process of producing puerarin includes the following steps: inducing excised organic to form callus tissues; succesive transfer culture of the callus tissues; and the suspension culture of the callus tissues. In each of steps, specially prepared culture medium is used. The production process of puerarin of the present invention has short period, high productivity and high quality.

Description

(1) Technical field [0001] The invention relates to a method for producing puerarin, specifically, a method for producing puerarin by adopting plant cell suspension culture on the basis of in vitro culture and asexual reproduction of Pueraria plants. (2) Background technology [0002] Plant cell engineering is a modern high-tech that combines biology and engineering based on the suspension culture of plant cells, tissues or organs for the purpose of obtaining specific products. The production of cell secondary metabolites by plant cell engineering is the most researched and widely used field of plant cell engineering. [0003] Pueraria lobata (Willd.) OhWi, a perennial herbaceous vine belonging to the genus Pueraria of Papilionaceae, is a traditional medicinal and edible resource in my country and other East Asian countries. The active ingredients of its tuber root (commonly known as Puerariae radix, Puerariae radix) are puerarin-based isoflavone compounds, which have the f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H17/065C12N5/04
Inventor 李玲张春荣刘慧丽
Owner SOUTH CHINA NORMAL UNIVERSITY
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