Induction and rapid propagation method of salvia chinensis hairy roots

A hairy root and purple ginseng technology, which is applied in the directions of horticultural methods, botanical equipment and methods, biochemical equipment and methods, etc., can solve problems such as research gaps in the induction and propagation of purple ginseng hairy roots, so as to improve the induction rate, The effect of protecting the ecological environment and rapid induction

Active Publication Date: 2019-08-27
CHINA PHARM UNIV
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  • Description
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  • Application Information

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Problems solved by technology

However, the research on the induction

Method used

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  • Induction and rapid propagation method of salvia chinensis hairy roots
  • Induction and rapid propagation method of salvia chinensis hairy roots
  • Induction and rapid propagation method of salvia chinensis hairy roots

Examples

Experimental program
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Effect test

Embodiment 1

[0041] Example 1 Screening and optimization of the optimal induction system for hairy roots of purple ginseng

[0042] 1. Experimental method

[0043] 1.1 Effects of different A. rhizogenes strains and explant types on the induction of hairy roots of purple ginseng

[0044] (1) Obtaining sterile seedlings of purple ginseng tissue culture: Select the nodes, rhizomes and leaves of the young buds of the purple ginseng plant in the current year as explants, shake them in 0.5% sodium hypochlorite for 8 minutes, and then use sterile water to rinse twice, each time 5min times. Then disinfect with 0.1% mercuric chloride for 5 to 8 minutes, and then rinse with sterile water for 4 times, 5 minutes each time. After that, the sterilized explants were evenly spread on the sterilized filter paper, and the residual water on the surface was dried. cm in size, were placed on MS solid medium containing 0.5 mg / L, 1 mg / L and 2 mg / L TDZ, respectively, and placed in a light incubator. About 20d...

experiment example 2

[0067] Experimental Example 2 Molecular detection of hairy roots of purple ginseng induced and propagated by the present invention

[0068] 1. Experimental method

[0069] When A. rhizogenes infects plants to form hairy roots, the rhizogen rol gene in the T-DNA region of the Ri plasmid of A. rhizogenes A4 is transferred into the plant genome, and finally hairy roots are induced. In order to confirm whether the induced roots are hairy roots, in addition to morphological observation and distinction, it is also necessary to perform molecular level detection on the hairy roots to determine whether the rol gene is inserted into the induced hairy roots.

[0070] After the hairy roots induced in Experimental Example 1 grew to a length of 1 to 3 cm, the genomic DNA of 3 randomly selected hairy roots of purple ginseng was extracted using the plant genomic DNA extraction kit of Beijing Qingke Xinye Biotechnology Co., Ltd., The rol genes (rolB and rolC genes) contained in hairy roots we...

experiment example 3

[0086] Experimental Example 3 LC-MS detection of RAs in the hairy roots of purple ginseng induced and propagated by the present invention

[0087] 1. Experimental method

[0088] After the hairy roots of purple ginseng were cultured in MS liquid medium for 60 days, LC-MS was used to detect whether the hairy roots contained RAs.

[0089] Weigh 1 g of the hairy roots dried to constant weight obtained in Experimental Example 1 and place them in a 25 mL conical flask with a stopper, pipette 10 mL of 83% methanol to soak overnight, ultrasonically treat at room temperature for 86 min, and extract 3 times. The solution was transferred and combined into an EP tube, centrifuged at 13,500 × g for 5 min, and the supernatant was transferred to a new EP tube for use. 1 mL of the supernatant was filtered through a 0.22 μm nylon filter for UPLC-MS detection.

[0090] Based on the structural characteristics and mass spectrometry fragmentation rules of RAs, Waters ACQUITY UPLC H Classsystem ...

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Abstract

The invention discloses an induction and rapid propagation method of salvia chinensis hairy roots, belonging to the technology of biological culture. The method comprises the following steps: (1) infecting salvia chinensis explants with an agrobacterium rhizogenes infection solution; (2) co-culturing the infected explants; (3) transferring the co-cultured explants to an induction medium for induction culture; (4) performing degerming subculture and multiplication culture on salvia chinensis hairy roots generated by induction. Agrobacterium rhizogenes type, explant type and key parameters suchas pre-culture time, infection time and co-cultivation time of the explants, which significantly affect the induction efficiency of the salvia chinensis hairy roots, are optimized to significantly improve the induction effect of the salvia chinensis hairy roots, achieve rapid and efficient induction of the salvia chinensis hairy roots, and expand sources for obtaining cyclic peptide; the method has the advantages of convenience in material extraction, simpleness in operation, fast induction, genetic stability and the like.

Description

technical field [0001] The invention belongs to biological culture technology, and particularly relates to a method for inducing and rapidly multiplying hairy roots of purple ginseng. Background technique [0002] Purple ginseng Rubia yunnanensis (Franch.) Diels, also known as Yunnan madder, is a medicinal plant of the Rubiaceae (Rubiaceae) genus (Rubia). 1700-2500 meters of thickets, grass slopes or roadside. Its roots and rhizomes are fleshy and strip-shaped. It is a special medicinal material of Yunnan, red ginseng, and is commonly used as an anti-tumor drug in the folk. Traditional Chinese medicine believes that small red ginseng is sweet, warm in nature, invigorating blood and relaxing tendons, removing blood stasis and regenerating, and nourishing blood. Myocardial ischemia, antioxidant and other activities, have important medicinal value and economic value. [0003] The chemical constituents of red ginseng mainly include anthraquinones, arborane-type triterpenes, a...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H4/00
CPCC12N15/8205A01H4/001
Inventor 谭宁华缪媛媛
Owner CHINA PHARM UNIV
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