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Chemical inducer for radix astragali adventitious root tissue culture and application of chemical inducer in radix astragali adventitious root culture

A chemical inducer, a technology for root tissue in applications, botanical equipment and methods, horticulture, etc.

Inactive Publication Date: 2022-07-08
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Cis-3-hexenal feedback inhibits the key enzyme of its synthesis pathway - lipoxygenase, and its application in the synthesis and accumulation of non-volatile active ingredients of medicinal plants has not been reported at home and abroad

Method used

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  • Chemical inducer for radix astragali adventitious root tissue culture and application of chemical inducer in radix astragali adventitious root culture
  • Chemical inducer for radix astragali adventitious root tissue culture and application of chemical inducer in radix astragali adventitious root culture
  • Chemical inducer for radix astragali adventitious root tissue culture and application of chemical inducer in radix astragali adventitious root culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment １

[0027] Example 1: Induction and subculture of adventitious roots of Astragalus mongolica

[0028] The tissue culture seedlings of Astragalus mongolica, which have been propagated for a long time in the laboratory (> 5 years) and are in the vigorous growth stage, are used as starting materials. figure 1 As shown in A, select the terminal buds with good growth, 1-2 cm in length, cut them aseptically, and transfer them to MS solid medium containing 0.7% agar, 30 g / L sucrose, and 1 mg / L naphthalene acetic acid. Cultured at 25°C, 16 hours (light) / 8 hours (dark) for 8 weeks, the early morphological characteristics of the induced adventitious roots are shown in the appendix figure 1 In B, the morphological characteristics of adventitious roots used in liquid culture are shown in the appendix figure 1 in C.

[0029] Subsequently, after aseptic shearing and washing of adventitious roots with sterile water, the culture was transferred to 30 mL 1 / 2 MS liquid medium (100 mL conical f...

Embodiment 2

[0030] Example 2: Lipoxygenase extraction, activity assay and optimization of phenidone incubation dose in vitro

[0031] Weigh 1.00g of Astragalus adventitious root with good growth at random, grind it into powder with liquid nitrogen, add pH9.0, 0.05 mol / L boric acid buffer solution according to the material ratio of 1:10 (w:v), mix well, extract in ice-water bath Centrifuge at 8000 rpm for 2 h at 4 °C for 10 min, transfer the supernatant to a new centrifuge tube, the obtained supernatant is the crude lipoxygenase extract, and store at 4 °C for future use. Under low temperature conditions, measure 30 μL of crude lipoxygenase extract into a new centrifuge tube, add 1000 μL of pH 9.0, 0.435 mmol / L linoleic acid solution preheated to 25°C, and mix well After incubation at 25°C for 10 min, 970 μL of absolute ethanol was added to terminate the reaction; at the same time, the reaction system with the addition of anhydrous ethanol and then the substrate solution was used as a blank...

Embodiment 3

[0034] Example 3: In vitro screening of green leaf volatiles with lipoxygenase activity inhibition

[0035] Select the same batch of crude lipoxygenase extract, pipette 120 μL into a new centrifuge tube, and add 50 mM of the following green leaf volatiles solution at a volume ratio of 30:1: n-hexanal, n-hexanol, hexyl acetate, cis -3-hexenal, cis-3-hexenol, trans-2-hexenal, trans-2-hexenol, cis-3-hexenyl acetate, and set normal control and solvent control at the same time (adding the same volume ratio of methanol) and positive control (obtained by screening in Example 2), mix well, incubate at 25°C for 10 min, and measure OD according to the method in Example 2 235 , the inhibition rate was calculated; the number of replicates per treatment: 3.

[0036] The results are attached image 3 , it can be seen from the figure that among the detected green leaf volatiles, cis-3-hexenal has the highest inhibition rate, which is at the same level as that of the positive control phenid...

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Abstract

The invention belongs to the technical field of chemical induction of traditional Chinese medicine radix astragali adventitious roots, and provides a chemical inducer for radix astragali adventitious root tissue culture and application of the chemical inducer in radix astragali adventitious root culture in order to solve the problems of green and sustainable development of radix astragali resources at present. Cis-3-hexenal is dissolved in water to prepare a mother solution with the concentration of 200 mmol / L, and the mother solution is stored at the temperature of 4 DEG C for standby application after being filtered and sterilized by a sterile filter membrane. The growth of the radix astragali adventitious roots and the synthesis and accumulation of triterpenes and isoflavone active ingredients are promoted in a dose-dependent manner, so that the browning of radix astragali adventitious root culture is inhibited; and synthesis and accumulation of astragaloside I, isoastragaloside I, astragaloside II, astragaloside III, cycloastragaloside II, liquiritin, formononin, calycosin, glucoside of calycosin and the like in the roots are promoted. The additive is expected to be developed into a green additive which is applied to an astragalus mongholicus adventitious root tissue culture system to improve the yield of root culture and the content of active substances.

Description

technical field [0001] The invention belongs to the technical field of chemical induction of Chinese medicine Astragalus adventitious root, and in particular relates to a chemical inducer for Astragalus adventitious root tissue culture and its application in Astragalus adventitious root culture; A chemical elicitor of the synthesis and accumulation of active ingredients. The addition of low concentration of cis-3-hexenal to the culture medium promoted the increase of the biomass of adventitious root tissue culture of astragalus, as well as the increase of the contents of glucoside, formononetin and astragaloside IV in the culture. Background technique [0002] Green leaf volatiles are linoleic acid, linolenic acid or conjugated galactolipids released by phospholipase hydrolysis of cell membrane lipids when plants are subjected to environmental stress such as mechanical damage, drought, fungal infection or under normal conditions. , A class of small molecular compounds forme...

Claims

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Application Information

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IPC IPC(8): A01H4/00
CPCA01H4/002A01H4/005
Inventor 孙海峰张晴晴张靖妮
Owner SHANXI UNIV
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