Snow lotus flavone active ingredient production method by cultivating acaleph snow lotus trichome

A technology of snow lotus flavonoids and hairy roots, which is applied in the field of plant biotechnology engineering, can solve problems that have not been seen yet, and achieve the effects of reducing cost and price, protecting the ecological environment, and saving electricity

Inactive Publication Date: 2004-08-18
INST OF BOTANY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, it is an effective way to solve the problem of snow lotus resources through hairy root induction and culture technology. However, it is quite difficult to estab

Method used

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  • Snow lotus flavone active ingredient production method by cultivating acaleph snow lotus trichome
  • Snow lotus flavone active ingredient production method by cultivating acaleph snow lotus trichome
  • Snow lotus flavone active ingredient production method by cultivating acaleph snow lotus trichome

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] 1. Establish a high-frequency regeneration system for jellyfish and snow lotus

[0042] 1.1 Seed disinfection treatment

[0043] Source of seeds: Dried jellyfish snow lotus (S.medusa Maxim) picked in Qilian Mountains in 2001, peeled off the seeds from the inflorescence

[0044] Cleaning: Choose healthy, full-bodied, mildew-free seeds, rinse them with tap water 4 times, soak them overnight at room temperature, and rinse them with running water for 1 hour.

[0045] Disinfection: Soak in 70% ethanol for 30s, act on 0.01% mercuric chloride for 10-14min, wash with sterile water 4-6 times.

[0046] Sterile seed culture: use hormone-free MS medium, 0.6% agar. 25°C, light intensity 2000lux, continuous light, the seeds germinated successively after 3 days.

[0047] 1.2 Inducing cotyledons to produce clustered seedlings

[0048] Explant treatment: select the green seedlings with cotyledons unfolded after 3 days of germination, cut at the cotyledon petiole, and leave a cotyled...

Embodiment 2

[0057] Embodiment 2: establish jellyfish snow lotus transformation system

[0058] 2.1 Transformation preparation experiment 1-determining the strain of Agrobacterium rhizogenes:

[0059] Determination of Agrobacterium strains: The infection effect of Agrobacterium rhizogenes LBA9402, R1601, and R1000 was tested with tobacco, and it was found that R1601 was the most toxic, and the time it took for the infected tobacco leaves to root was the shortest, and the induced hair roots had typical Morphological characteristics of hairy roots (Kanamycin 50mg / l, slender, many hairs, weak gravitropism, growth along the bottle wall, oblique secondary roots), and the frequency of 90% of the same family plant Annua annua R1601 Hairy roots were induced, so we determined to use Agrobacterium rhizogenes R1601 to infect Saussurea jellyfish.

[0060] 2..3 Transformation preparation experiment 2-determination of Km screening pressure

[0061] The clustered seedlings were cultured in the maintena...

Embodiment 3

[0074] The screening of embodiment 3 hairy roots

[0075] Delayed screening: Insert the leaves with roots into the delayed screening medium 1 / 2MS (20mg / )+Cef (500mg / l)++IAA (0.5-1.0mg / l)+NAA (0.5mg / l)+GA 3 (0.5mg / l), keep low light culture for 5-8 days to promote the induction and growth of hair roots.

[0076] Km screening: cut off the 1.0---2.5cm long root tips of all roots and insert them into the screening medium 1 / 2MS(20mg / l)+Cef(300mg / l)+Km(50mg / l), low light culture, After 10 days, 1.0-2.5 cm root tips of the Km-resistant root system were cut out for liquid culture.

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Abstract

The present invention is the process of culturing Saussurea involucrata hairy root with Saussurea involucrata hairy root cell line and liquid and producing Saussurea involucrata flavone compound. The process includes the following steps: culturing green cotyledon of three days age in caespitose seedling culture medium to inducing caespitose seedling; culturing single plant with the caespitose seedling in seedling growing and strengthening culture medium; culturing in root inducing culture medium to obtain seedling with satus root; cutting the foliage into explant, dark culturing the explant and transforming with mediating rooting agrobacterium to inducing rooting; re-culturing the foliage with hairy root in screening culture medium; cutting root tip of selected anti-Km root line with positive mannopine test for expanding culture; drying, crushing, separating and extracting the cultured material to prepare Saussurea involucrata flavone compound.

Description

technical field [0001] The invention belongs to the field of plant biotechnology engineering, and specifically relates to a method for cultivating hairy roots of snow lotus with high yield of flavonoids, and also includes separation, extraction and preparation process methods of flavonoids. technical background [0002] Snow lotus, also known as snow lotus, is one of the three rare plants in my country. As early as in "Northwest Territories" and "Xiangyuan Xiaozhi", there are records about snow lotus, and the two are combined into one, collectively called snow lotus. As snow lotus, the name of the native plant varies with the place of origin. For example, S.tridactyla Sch.Bip. is used in Tibet and other places; S.lanicps Hand.-Mazz. is used in Sichuan, Yunnan and other places; Qinghai and other places use jellyfish snow lotus S.medusa Maxim; Qinghai also uses snow rabbit S.gossypiphora D.Don.; It belongs to the subgenus Subgen Eriocorye of Saussurea, and is generally distri...

Claims

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Application Information

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IPC IPC(8): A01H4/00A01H5/06C07D311/30C07H1/08C07H17/065
Inventor 赵德修陈亚琼付春祥
Owner INST OF BOTANY CHINESE ACAD OF SCI
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