Tripterygium wilfordii regenerated plant and its preparation process

A technique for regeneration of plants and preparation process, applied in the biological field, can solve problems such as instability of active ingredients in culture, difficulty in large-scale culture, low yield of callus or suspension cell culture, etc.

Inactive Publication Date: 2002-09-04
上海延农生物工程有限公司 +2
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  • Abstract
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AI Technical Summary

Problems solved by technology

[0005] 1. Callus or suspension cells are undifferentiated cultures, while wild Tripterygium wilfordii uses plants as medicinal materials, which are differentiated products. There are essential differences between the two
[0006] 2. The culture yield of c

Method used

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  • Tripterygium wilfordii regenerated plant and its preparation process

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042]Example 1 Clonal Variation and Mutagenesis of Tripterygium wilfordii Cells or Callus

[0043] Take the young tissues of wild tripterygium wilfordii or seedlings as explants, sterilize with 75% ethanol for 30 seconds, sterilize with fenfenzolin or bleach for 15 minutes, wash with sterile water, cut into thin slices for induction culture, induce culture The base is 2 / 3MS, 2,4-dichlorophenoxyacetic acid (2,4-D) 1.0mg / L, naphthaleneacetic acid (NAA) 0.1mg / L, kinetin (KT) 0.1mg / L, 5% sucrose , 0.6% agar, cultured for 35 days to induce callus formation. Then carry out subculture of 2 to 3 generations, the subculture medium is: 2 / 3N6, hydrolyzed casein (CH) 500mg / L, 2,4-dichlorophenoxyacetic acid (2,4-D) 0.5mg / L L, indole butyric acid (IBA) 0.5 mg / L, kinetin (KT) 0.1 mg / L, 4% sucrose, 0.6% agar, each generation was cultured for 30 days to obtain a large number of callus. Put a large number of calli into liquid medium (2 / 3N 6 , 500mg / L hydrolyzed casein (CH), 2,4-dichlorophen...

Embodiment 2

[0046] Example 2 Plant Regeneration of Tripterygium wilfordii Suspension Culture Cells or Cell Clusters

[0047] The bright yellow suspension culture cells or cell clusters that have been subcultured or repaired are subjected to differentiation culture. The differentiation medium is 1 / 2MS, indole acetic acid (IAA) 0.1mg / L, kinetin (KT) 0.5mg / L, 6-benzylaminopurine (6-BA) 1.5mg / L, 3% sucrose, 0.6% agar, culture temperature is 22~25 ℃, and daily light is 12 hours, after 30 days, select thick normal seedling from regenerated seedling.

Embodiment 3

[0048] Example 3 Screening of Tripterygium wilfordii strains with high reproduction coefficient and active ingredient content

[0049] Carry out proliferation culture to the regenerated plants, the proliferation medium is 2 / 3MS, hydrolyzed casein (CH) 500mg / L, kinetin (KT) 0.5mg / L, 6-benzylaminopurine (6-BA) 0.5mg / L , naphthalene acetic acid (NAA) 0.1mg / L, 3% sucrose, 0.6% agar, select stout, fast-growing strains, get its part plants and carry out the mensuration of total diterpene lactone content (Lin Qishou etc., " Chinese herbal medicine component chemistry " 1977, page 390, Beijing, Science Publishing House), with the Tripterygium wilfordii extract sheet (batch number 20000505) of Leishi Pharmaceutical Shanghai No.

[0050] Select the high regenerated plant of total diterpene lactone content to carry out the mensuration of triptolide content again (Ye Xiaochuan, "Journal of Pharmaceutical Analysis", 1997, the 5th phase of 17 volumes: 319 pages), the standard sample of trip...

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Abstract

The preparation method of regenerative plant of thunder god vine mainly utilizes the techniques of clonal variation, mutation, chromosome doubling, regeneration and hormoneless propagation, etc. Saidinvention not only can make mass propagation, and the propagation coefficient of every generation can be up to 5-6 times, and the triptolide alcohol content is 0.009-0.015%, and its total diterpene lactone content also is higher than that of commercial thunder god vine extract, and they can be retained stably.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to a regenerated plant of Tripterygium wilfordii and its preparation process. Background technique [0002] Tripterygium wilfordii Hook f. is a plant of the genus Tripterygium wilfordii Hook in the family Euonymus. Pharmacological experiments show that Tripterygium wilfordii has obvious anti-tumor, anti-inflammatory, immunosuppressive and anti-fertility effects. The current research results show that tripterygium wilfordii diterpene lactones are its main active ingredients. Tripterygium wilfordii has complex chemical components and extensive pharmacological effects, and has important clinical application value. [0003] However, because Tripterygium wilfordii is a perennial woody vine with slow growth, it has been widely used clinically, resulting in serious shortage of wild resources. Therefore, the artificial cultivation of Tripterygium wilfordii plays an ine...

Claims

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Application Information

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IPC IPC(8): A01H4/00
Inventor 黄剑华陆瑞菊孙月芳王亦菲周润梅俞庆全
Owner 上海延农生物工程有限公司
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