Efficient inducing method of rhizoma coptidis hairy roots

A technology of hairy root and Coptis chinensis is applied in the biological field to achieve the effects of reducing browning rate and mortality, enhancing antioxidant capacity and improving stress resistance

Inactive Publication Date: 2019-03-26
赵静
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no report on the use of Agrobacterium rhizogenes to induce hairy roots from Coptidis Rhizoma

Method used

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  • Efficient inducing method of rhizoma coptidis hairy roots
  • Efficient inducing method of rhizoma coptidis hairy roots
  • Efficient inducing method of rhizoma coptidis hairy roots

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] A high-efficiency induction method of Coptis chinensis hairy roots, specifically comprising the steps of:

[0040] S1. Acquisition of sterile tissue culture seedlings

[0041] Select robust Coptis chinensis plants, wash them with flowing tap water for 30 minutes, cut the stems with buds on the plants in a purification workbench, wash them with sterile water for 2 to 3 times, soak them in 75% ethanol for 30 to 50 seconds, and wash them off with sterile water. Finally, soak in 0.2% mercuric chloride for 8-10 minutes, wash away the residue with sterile water; place the sterilized stems on sterile filter paper to absorb the moisture on the surface, and then inoculate them with 0.5mg / LNAA, 0.1mg / L6- On the MS solid medium of BA, culture under the conditions of temperature 22°C, light intensity 1800lx, and light time 13h / d until they grow into sterile tissue culture seedlings with true leaves.

[0042] S2. Pre-cultivation of explants

[0043] Under aseptic conditions, cut t...

Embodiment 2

[0050] Example 2 Effect of Explant Types on the Induction Rate of Coptis Rhizoma Hairy Roots

[0051] Under sterile conditions, the cotyledons, true leaves, petioles and stems of Coptidis aseptic tissue cultured seedlings were cut into 0.5 cm 2 The small piece of is used as explant, and other operation steps are all the same as embodiment 1, compares the influence of explant type on Coptis chinensis hairy root induction rate, the result is as follows figure 1 shown.

[0052] test results( figure 1 ) showed that the type of explant had a significant effect on the induction rate of Coptidis hairy roots. In the experiment, the order of induction ability of four kinds of explants to hairy root was: true leaf>stem segment>petiole>cotyledon. Since the transformation of Agrobacterium rhizogenes only occurs in the S phase of cell division (that is, the DNA synthesis phase), vigorously growing tissues and organs—true leaves of sterile tissue culture seedlings—are more likely to in...

Embodiment 3

[0053] Example 3 Effect of preculture on induction rate of Coptis chinensis hairy roots

[0054] 1) Effect of pre-culture time on induction rate of Coptis chinensis hairy roots

[0055] The pre-cultivation time of the explants in Example 1 is set to 0, 1, 2, 3, 4, 5d, and other operation steps are all the same as in Example 1, and the influence of the pre-culture time on the induction rate of Coptis chinensis hairy roots is compared, the result Such as figure 2 shown.

[0056] test results( figure 2 ) showed that the pre-culture time had a significant effect on the induction rate of Coptis chinensis hairy roots. Compared with no pre-culture (pre-culture time is 0d), the induction rate of Coptis chinensis hairy roots has been improved after pre-culture. Within 1-3 days of pre-culture, the induction rate of Coptis chinensis hairy roots increased with the prolongation of pre-culture time, and the induction rate was the highest at 3 days, and then gradually decreased after 3...

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Abstract

The invention discloses an efficient inducing method of rhizoma coptidis hairy roots. The efficient inducing method specifically comprises the following steps: S1, sterile tissue culture seedlings areobtained; S2, explants are pre-cultured; S3, agrobacterium rhizogenes is activated; S4, the hairy roots are induced and sterilized; S5, liquid culture of the hairy roots is conducted; and S6, PCR detection of the hairy roots is conducted. By systematacially researching multiple factors influencing generation of the rhizoma coptidis hairy roots, certain methods for increasing the inducing rate ofthe hairy roots are explored, an efficient inducing system of the rhizoma coptidis hairy roots is built for the first time, and the inducing rate of the hairy roots reaches up to 88.7%; and the hairyroots obtained through the efficient inducing method are easy to breed and culture on a large scale, the potency of generating secondary metabolites is achieved, and the foundation for future development and utilization of the rhizoma coptidis hairy roots is laid.

Description

technical field [0001] The invention relates to a method for inducing hairy roots of a medicinal plant Coptis chinensis by using Agrobacterium rhizogenes. Belongs to the field of biotechnology. Background technique [0002] Coptis ( Coptis chinensis Franch. ), also known as Weilian, Chuanlian, Jizhulian, Jizhuhuanglian, Guanglian, is a perennial herb of the Ranunculaceae Coptis genus. It is distributed in Sichuan, Guizhou, Hunan, Hubei, Shaanxi, Zhejiang, Guangdong and other regions in China. The dried rhizomes of Coptis chinensis, Coptis triangularis, and Coptis yunnanensis in Coptis genus are the traditional Chinese medicine "Coptis chinensis", which was first recorded in "Shen Nong's Materia Medica", and has a history of more than two thousand years. Traditional Chinese medicine believes that Coptis chinensis is bitter in taste and cold in nature, and enters the heart, liver, stomach, and large intestine meridians. It has the effects of clearing away heat and dampness, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 赵静
Owner 赵静
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