Inducing method of psammosilene tunicoides hairy roots

A root system and hairy technology, applied in the field of induction of hairy root systems of golden iron locks, can solve the problems of decreased growth rate, impact on harvest yield, death, etc., and achieve the effect of improving the induction transformation rate.

Active Publication Date: 2015-08-19
DALIAN PRACTICAL BIOTECH
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  • Abstract
  • Description
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AI Technical Summary

Problems solved by technology

The outstanding problems existing in the preparation and liquid culture of Jintiezhao hairy roots: (1) The induction rate of hairy roots is low, about 20%, which is determined by the inherent characteristics in the interaction process between pla

Method used

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  • Inducing method of psammosilene tunicoides hairy roots

Examples

Experimental program
Comparison scheme
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preparation example Construction

[0028] Preparation of reference solution: Accurately weigh about 20mg of oleanolic acid standard substance (dried to constant weight at 120°C under reduced pressure), dissolve it in absolute ethanol, and set the volume in a 50mL volumetric flask to obtain the oleanolic acid reference substance Sample liquid.

[0029] Preparation of the sample solution: Accurately weigh about 30 mg of the crude total saponins of Aurantia chinensis dried to constant weight, dissolve it in absolute ethanol, and set the volume in a 50 mL volumetric flask to obtain the sample solution.

[0030] Determination of the maximum absorption wavelength: Accurately measure 0.6mL of the standard solution and 5mL of the sample solution, place each in a 10mL volumetric flask, evaporate the solvent on a water bath at 75°C, and then add a newly prepared 5% vanillin-glacial acetic acid solution (50mg vanillin dissolved in 10mL glacial acetic acid) 0.4mL, perchloric acid 1.6mL, shake well, heat in a water bath at ...

Embodiment 1

[0035] The high-efficiency induction of embodiment 1 golden iron lock hairy root system

[0036] (1) Take the leaves of Jintiesuo aseptic seedlings, remove the tissue around the leaf edge, and use 10mmol / L CuCl 2 After soaking the explants in the aqueous solution for 10 minutes, dry the excess water on the surface of the explants with sterile filter paper;

[0037](2) Explants treated with Agrobacterium rhizogenes dipping step (1):

[0038] 2a. Agrobacterium rhizogenes ACCC10060 bacterial liquid was inoculated in YEB solid medium, and cultured in the dark at 25°C ± 1 for 2-3 days to obtain clones of Agrobacterium rhizogenes ACCC10060;

[0039] 2b. Pick the clone of Agrobacterium rhizogenes ACCC10060, inoculate it in YEB medium, and culture it with shaking at 25°C±1; take 1ml of the bacterial solution with an OD value of 0.6-0.8 and place it in 100ml YEB liquid medium and shake it to OD When the value is 0.6, the bacteria liquid is collected, and the bacteria are collected by...

Embodiment 2

[0048] Embodiment 2: the comparison induction of golden iron lock hairy root system

[0049] According to the method of embodiment 1, but omit the CuCl with 10mmol / L in the step (1) 2 The step of immersing the explant in the aqueous solution for 10 minutes is followed by the next step with the cleaned Jintiesu explant to induce the generation of the hairy root of Jintiezio.

[0050] In the culture process of this embodiment, 4-6 days after inoculation, hairy roots will occur at the petiole and leaf veins, and hairy roots will be produced in 5-8 days, and the rooting time is longer than that of Example 1; the induced transformation rate is lower than 30%..

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Abstract

The invention relates to an inducing method of psammosilene tunicoides hairy roots, and particularly relates to a method for inducing a psammosilene tunicoides explants to produce hairy roots by adopting rooting agrobacterium. The method comprises the step of treating the explants by utilizing 10 to 15 mmol/L of CuCl2 aqueous solution before the induction. According to the method, the psammosilene tunicoides leaves are treated by utilizing CuCl2, the leaves are immersed by virtue of the rooting agrobacterium, the hairy roots can be produced on leaf veins of a leaf stalk and leaf margins in 2 to 4 days, the rooting time is shortened, the rooting can be generated in 3 to 5 days, the inducing conversion rate is greatly increased, and the inducing conversion rate can reach 60 percent or more. The hairy roots grow rapidly in a hormone-free culture medium by virtue of screening.

Description

technical field [0001] The invention relates to a method for efficiently inducing and cultivating a hairy root system by using biotechnology, especially a method for efficiently inducing a gold iron lock hairy root system. Background technique [0002] Psammosilene tunicoides (Psammosilene tunicoides W.C.Wu et C.Y.Wu) is a perennial herbaceous plant of the genus Psammosilene tunicoides in the family Caryophyllaceae, also known as Dudingzi, Xiaobawang, Kunming Aphrodisiac, Jinsi Aiduoduo, Tunicoides, etc. It is a unique single-genus species in southwest my country Medicinal plants. The roots of Jintiesuo are mainly used as medicine, and the main medicinal ingredients are triterpenes, triterpene saponins, cyclic peptides and lactams. Pharmacological studies have found that the extracts of Jintiesuo are mainly analgesic and anti-inflammatory, blood stasis and bleeding, anti-tumor, immune regulation, antibacterial It is used for the treatment of bruises, rheumatism, gas pain, tr...

Claims

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Application Information

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IPC IPC(8): A01H4/00
Inventor 张宗申王超金朝霞雒怀宇杨思雨窦旖君陈阳王发德
Owner DALIAN PRACTICAL BIOTECH
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