Cultivation method for inducing scutellaria baicalensis hairy root based on agrobacterium rhizogenes infection
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A technology of Agrobacterium rhizogenes and a cultivation method, applied in the field of artificial cultivation of Scutellaria baicalensis, can solve problems such as shortage of Scutellaria baicalensis resources, and achieve the effects of strong secondary metabolite ability, low energy consumption and low cost
Inactive Publication Date: 2011-11-02
SHAANXI UNIV OF SCI & TECH
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[0004] The problem to be solved by the present invention is to provide a cultivation method for inducing hairy roots of Scutellaria baicalensis based on Agrobacterium rhizogenes infection, which can induce hairy roots quickly and with a high induction rate, and solve the problem of shortage of Scutellaria baicalensis resources
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Embodiment 1
[0043] A method for cultivating hairy roots of Scutellaria baicalensis based on infection by Agrobacterium rhizogenes, comprising the following steps:
[0044] 1) Acquisition and pre-culture of Scutellaria baicalensis explants
[0045] The germinated Scutellaria baicalensis seeds are inoculated into a pre-medium and cultured at 25-28° C. to grow into seedlings, and the stem segments of the seedlings are cut out as explants.
[0046] Alternatively, directly take Scutellaria baicalensis seedlings and sterilize them, and then cut the stem segments of the seedlings as explants.
[0047] The obtained sterile Scutellaria baicalensis explants were inoculated on the pre-culture medium, and pre-cultured for 2-3 days at 25-28°C under dark conditions;
[0048] The pre-medium is MS solid medium containing 0.15-1.0 mg / kg 6-BA and 0.1-0.35 mg / kg NAA (specifically, 1 LMS medium supplemented with 30 g sucrose and 5 g agar).
[0049] 2) Activation of A. rhizogenes and transfection of explant...
Embodiment 2
[0070] The optimized culture of hairy roots is the same as the screening method in Example 1, the difference is that the supplementary components of the optimized medium are different.
[0071] Described optimized medium is added 40g / L of glucose, 0.5g / L of hydrolyzed milk protein, 50mg / L of Aspergillus niger elicitor, 1.0mmol / L of Ca on the hairy root liquid basic medium. 2+ and 0.25mg / L NAA, pH 7.0.
Embodiment 3
[0073] The optimized culture of hairy roots is the same as the screening method in Example 1, the difference is that the supplementary components of the optimized medium are different.
[0074] Described optimized medium is to add 20g / L sucrose, 0.45g / L ammonium nitrate, 40mg / L Aspergillus oryzae elicitor, 1.0mmol / L Mg on hairy root liquid basic medium 2+ And 0.25mg / L 6-BA, pH value is 4.0.
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Abstract
The invention discloses a cultivation method for inducing scutellaria baicalensis hairy root based on agrobacterium rhizogenes infection, which uses an agrobacterium rhizogenes bacteria liquid to infect root of a scutellaria baicalensis explant and induces the scutellaria baicalensis explant to form a hairy root, the scutellaria baicalensis hairy root presents yellow hairy on a MS solid medium, so that scutelloside can be stably synthesized, an ITS sequencing is performed and the hairy root can be identified as the scutellaria baicalensis hairy root of a scutellaria. Then seed selection is performed for further in a seed selection medium and a cultivation condition can be optimized to select lines which raise the content of active ingredient scutelloside used for hairy root traditional Chinese medicines. Hairy root can be obtained through scutellaria of a scutellaria introduced by agrobacterium rhizogenes, wherein the inductivity can reach 10 - 40%, the obtained scutellaria baicalensis hairy root presents yellow hairy which is capable of stably synthesizing scutelloside. According to the invention, the scutellaria baicalensis hairy root of a scutellaria is cultivated on the optimized medium, the growth is rapid and the biomass can be increased for 23 -35 times in 30 - 35 days, and the cultivation is performed under the dark conditions. The invention has the characteristics of low energy consumption and low cost, which is suitable for industrialization production.
Description
technical field [0001] The invention belongs to the technical field of artificial cultivation of scutellariae, and relates to a cultivation method for inducing hairy roots of scutellaria baicalensis based on Agrobacterium rhizogenes infection. Background technique [0002] Scutellaria baicalensis Georgi. (Scutellaria baicalensis Georgi.) is a plant of the Labiatae family and is a commonly used traditional Chinese medicine. It is cold in nature and bitter in taste. It belongs to the lung, heart, liver, gallbladder, and large intestine meridians. Scutellaria baicalensis is a perennial herb, which likes mild climate and is resistant to cold and drought. It is mainly distributed in Hebei, Liaoning, Shaanxi, Shandong, Inner Mongolia and other places. The main components are baicalin, baicalein, wogonin, and wogonin. With the in-depth international research on Scutellariae, the main component of Scutellariae baicalin has good effects in scavenging oxygen free radicals, reducing t...
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