CN111226793A - Method for producing astragalus membranaceus hairy roots in large scale - Google Patents
Method for producing astragalus membranaceus hairy roots in large scale Download PDFInfo
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- CN111226793A CN111226793A CN202010135220.0A CN202010135220A CN111226793A CN 111226793 A CN111226793 A CN 111226793A CN 202010135220 A CN202010135220 A CN 202010135220A CN 111226793 A CN111226793 A CN 111226793A
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- mother liquor
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- astragalus membranaceus
- hairy roots
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- 229910000366 copper(II) sulfate Inorganic materials 0.000 claims description 3
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- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 claims description 3
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- RADKZDMFGJYCBB-UHFFFAOYSA-N pyridoxal hydrochloride Natural products CC1=NC=C(CO)C(C=O)=C1O RADKZDMFGJYCBB-UHFFFAOYSA-N 0.000 claims description 3
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- DPJRMOMPQZCRJU-UHFFFAOYSA-M thiamine hydrochloride Chemical compound Cl.[Cl-].CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N DPJRMOMPQZCRJU-UHFFFAOYSA-M 0.000 claims description 3
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/005—Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8201—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation
- C12N15/8202—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation by biological means, e.g. cell mediated or natural vector
- C12N15/8205—Agrobacterium mediated transformation
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- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
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Abstract
The invention provides a method for producing astragalus root hairy roots in large scale, which comprises the steps of preparation of astragalus aseptic seedlings, activation and culture of agrobacterium tumefaciens, pre-culture and co-culture of the hairy roots and enlarged culture of the hairy roots.
Description
Technical Field
The invention relates to a method for producing astragalus hairy roots in a large scale, belonging to the technical field of astragalus biotechnology cultivation.
Background
Astragalus root belongs to perennial herb of leguminosae, and has been used as medicine by root, so far, more than 2000 years of history. The astragalus root has the main pharmacological actions of tonifying qi, strengthening exterior, promoting urination, expelling toxin, discharging pus, promoting granulation and the like, is mainly used for treating short breath, collapse, palpitation, spontaneous perspiration, body weakness, edema, chronic nephritis, dehydration, chronic diarrhea, qi deficiency, deficiency of qi and blood, sinking of middle-jiao, has the effects of resisting bacteria, diminishing inflammation, reducing blood pressure, promoting urination, detoxifying and benefiting gallbladder and the like, and is a common Chinese herbal medicine.
With the increasing enhancement of health care concept of people, the demand of astragalus membranaceus will increase year by year, but at present, the wild resources of astragalus membranaceus decrease year by year, the cultivars decrease, and the astragalus membranaceus is troubled by the problems of pesticide pollution, heavy metal residue and the like. Therefore, the application of modern biotechnology to increase the yield of astragalus, improve the quality of the astragalus and explore feasible ways of industrial production has important significance to the production of the astragalus, the astragaloside is a content detection index specified by Chinese pharmacopoeia, and if an artificial astragalus product for increasing the content of the astragaloside can be found, the astragalus has good development and application prospects.
Infection of plants by Agrobacterium rhizogenes (Agrobacterium rhizogenes) is a natural genetic transformation pattern in nature, which results in the production of hairy roots by integration and expression of a portion (T-DNA) of the root-causing plasmid (Ri) contained in the plant genome. The hairy root has the characteristics of rapid growth, multiple branches, no geotropism and hormone autotrophy, and the physiological, biochemical and genetic properties of the hairy root are stable. The hairy roots also keep certain biochemical functions of normal roots of plants, can efficiently produce secondary metabolites, and is particularly suitable for producing effective components of substituted plant roots. Since the first successful transformation of higher plants with Agrobacterium rhizogenes in 1997 Ackermann, more than 100 plant hairy root culture systems have been established to date, many of which are medicinal plants. For example, small-scale culture of hairy roots of Nicotiana citrifolia, Datura stramonium, belladonna, hyoscyami, Vinca rosea, Lithospermum erythrorhizon, Horseradish has been successful. The method for producing the effective secondary metabolites of the medicinal plants in a large scale by utilizing the hairy root culture technology has very attractive prospect, and has positive significance for protecting wild resources and ecological environment and realizing sustainable development. And the agrobacterium rhizogenes Ri plasmid is used for inducing the Mongolian milkvetch root to establish an asexual propagation system for hairy root culture, so that a feasible new way can be provided for the development, utilization and research of resources.
Disclosure of Invention
The invention aims to overcome the defects of different contents and different qualities of effective components in the astragalus mongholicus hairy roots, protect precious soil resources by using biotechnology, provide a method for industrially producing the astragalus mongholicus hairy roots in a large scale, and stably produce the astragalus mongholicus hairy roots in a high yield on the basis of ensuring the content of astragaloside.
In order to achieve the above objects, the present invention provides a method for mass production of hairy roots of Astragalus membranaceus, comprising the steps of:
step 1. preparation of Astragalus aseptic seedling
Soaking radix astragali seed in tap water for 20-24 hr, soaking in 70% ethanol for 1-3min, and washing with sterile water for 3-5 times; soaking in 1% mercuric chloride solution for 8-10 min; washing with sterile water for 5-10 times; inoculating to MS culture medium sterilized in advance, culturing at 25-30 deg.C and humidity of 50-60% to obtain sterile seedling of radix astragali;
step 2, strain activation and culture
Adding agrobacterium to a liquid YEB culture medium, culturing at 26-28 ℃ and 220-; then, culturing the bacterial liquid in 1/2MS liquid culture medium at the temperature of 26-28 ℃ and the rotation speed of 150-;
step 3 Material Pre-culture and Co-culture
Transferring the leaves, stems, hypocotyls and petioles of the aseptic seedlings to an MS culture medium added with phytohormone, culturing for 1-2 days in the dark, and inducing callus; then, fully contacting with the bacterial liquid in the step 2, and co-culturing for 30-40min at 50-60rpm and 25-28 ℃;
step 4, degerming
Adding the callus obtained in the step 3 to 1/2MS solid culture medium containing 500mg/L cefotaxime sodium (cef), and culturing at 25 ℃ in the dark; replacing the culture medium with new one week at intervals, and gradually reducing the concentration of cefotaxime sodium (cef);
separating the roots after the roots reach 3cm, placing the roots on 1/2MS solid culture medium containing 50mg/L cefotaxime sodium (cef), subculturing once every other week for three times, and then placing the roots on 1/2MS solid culture medium without cefotaxime sodium (cef) for culture;
step 5, enlarged cultivation of hairy root
The hairy roots of step 4 were transferred to 6,7-V liquid medium, expanded at 25-28 ℃ at 100-.
Further, the astragalus seeds are selected from one of astragalus mongholicus seeds or astragalus membranaceus seeds;
further, the composition per liter of the MS medium is as follows:
macroelements: KNO 34-6 mM, Ca (NO3) 2.4H 2O 4-6mM, MgSO 4.7H 2O 1.5.5-2.5 mM, KH2PO40.8-1.2mM,
trace elements: h3BO 344-48 μ M, MnCl 2.4H2O 8-10 μ M, ZnSO 4.7H2O 0.6-0.8 μ M, CuSO 4.5H2O 0.2-0.4 μ M, Na2MoO 4.2H2O 0.1-0.3 μ M, EDTA-Fe-Na 40-60 μ M;
organic matter: inositol 80-100. mu.M, sucrose 100-150. mu.M, and hydrolyzed casein 300-500. mu.M.
Further, the 1/2MS culture medium is composed by reducing the dosage of macroelements in the MS culture medium by half.
Further, the agrobacterium strain is selected from ATCC15834, LBA 9402.
Further, the plant hormone in the step 3 is one of IBA and NAA, and the content is 0.1-0.3 mg/L.
Further, the composition of the liquid YEB medium was as follows: dissolving yeast powder (1g), beef extract (5g), peptone (5g), sucrose (5g) and MgSO4 & 7H2O (0.5g) in 1L distilled water, adjusting pH to 7.4 with sodium hydroxide, autoclaving at 121 deg.C for 20 min;
further, the composition and preparation method of the 6,7-V culture medium are as follows:
A. preparation of component mother liquor
(1) Macroelement mother liquor: the mother liquor contains 16g KNO3, 2g (NH4)2SO4, 5g MgSO4 & 7H2O, 3g NaH2PO4, 4g KCl and 0.4g Na2HPO4 per liter;
(2) CaCl2 mother liquor: the mother liquor contains 4g of CaCl2 & 2H2O per liter
(3) And (3) a microelement mother solution: the mother liquor contains 4g of MnSO4 & H2O, 1.5g of ZnSO4 & 7H2O, 5g H3BO3, 0.05g of KI, 0.25g of NaMoO4 & 2H2O, 0.25g of CuSO4 & 5H2O and 0.25g of CoCl2 & 6H2O per liter
(4) Mother liquor of iron salt: the mother liquor contains 1.86g of EDTA-Na2 and 1.39g of FeSO 4.7H 2O per liter
(5) Vitamin mother liquor: the mother liquor contains nicotinic acid 0.125g, vitamin B1 0.05g, vitamin B6 0.05g, and inositol 10g per liter
B. The preparation method of the 6,7-V liquid culture medium comprises the following steps: taking 50mL of macroelement mother liquor, 50mL of CaCl2 mother liquor, 1mL of microelement mother liquor, 10mL of ferric salt mother liquor, 10mL of vitamin mother liquor and 30g of cane sugar; dissolving in 1L distilled water to adjust pH to 5.8.
Preferably, 7g of agar powder is added in the preparation of 6,7-V solid medium.
Further, the step-wise reduction of the concentration of cefotaxime sodium (cef) in the step 4 is performed according to the manners of 400mg/L, 300mg/L, 200mg/L and 100 mg/L.
The invention has the following beneficial effects: according to the method, the astragalus mongholicus hairy roots are produced by agrobacterium induction, parameters such as a culture medium, culture temperature and culture time are adjusted and optimized, and the astragalus mongholicus hairy roots which can be stably proliferated and have stable properties are screened out, wherein the hairy roots have high astragalus mongholicus methyl glycoside content. The invention can stably and highly produce the hairy roots of the astragalus on the basis of ensuring the content of the astragaloside, and has the advantages of short production period, uniform production quality and adaptation to the growth condition at normal temperature.
Drawings
FIG. 1 is a diagram showing the activation of strains in the method for mass production of hairy roots of Astragalus membranaceus according to the present invention
FIG. 2 is a diagram of the pre-cultivation of materials in the method for mass production of hairy roots of Astragalus membranaceus according to the present invention
FIG. 3 is a diagram of co-cultivation of materials in the method for mass production of hairy roots of Astragalus membranaceus according to the present invention
FIG. 4 is a diagram showing the material sterilization process in the method for mass production of hairy root of Astragalus membranaceus according to the present invention
FIG. 5 is an enlarged culture diagram of the method for mass production of hairy roots of Astragalus membranaceus according to the present invention
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention. In addition, the technical features involved in the embodiments of the present invention described below may be combined with each other as long as they do not conflict with each other.
Preparing an MS culture medium, wherein each liter of the MS culture medium comprises the following components:
macroelements: KNO 34-6 mM, Ca (NO3) 2.4H 2O 4-6mM, MgSO 4.7H 2O 1.5.5-2.5 mM, KH2PO40.8-1.2mM,
trace elements: h3BO 344-48 μ M, MnCl 2.4H2O 8-10 μ M, ZnSO 4.7H2O 0.6-0.8 μ M, CuSO 4.5H2O 0.2-0.4 μ M, Na2MoO 4.2H2O 0.1-0.3 μ M, EDTA-Fe-Na 40-60 μ M;
organic matter: inositol 80-100. mu.M, sucrose 100-150. mu.M, and hydrolyzed casein 300-500. mu.M.
Meanwhile, the 1/2MS culture medium is prepared by halving the dosage of macroelements in the MS culture medium;
the method for preparing the liquid YEB culture medium is as follows: dissolving yeast powder (1g), beef extract (5g), peptone (5g), sucrose (5g) and MgSO4 & 7H2O (0.5g) in 1L distilled water, adjusting pH to 7.4 with sodium hydroxide, autoclaving at 121 deg.C for 20 min;
the composition and preparation method of the 6,7-V culture medium are as follows:
A. preparation of component mother liquor
(1) Macroelement mother liquor: the mother liquor contains 16g KNO3, 2g (NH4)2SO4, 5g MgSO4 & 7H2O, 3g NaH2PO4, 4g KCl and 0.4g Na2HPO4 per liter;
(2) CaCl2 mother liquor: the mother liquor contains 4g of CaCl2 & 2H2O per liter
(3) And (3) a microelement mother solution: the mother liquor contains 4g of MnSO4 & H2O, 1.5g of ZnSO4 & 7H2O, 5g H3BO3, 0.05g of KI, 0.25g of NaMoO4 & 2H2O, 0.25g of CuSO4 & 5H2O and 0.25g of CoCl2 & 6H2O per liter
(4) Mother liquor of iron salt: the mother liquor contains 1.86g of EDTA-Na2 and 1.39g of FeSO 4.7H 2O per liter
(5) Vitamin mother liquor: the mother liquor contains nicotinic acid 0.125g, vitamin B1 0.05g, vitamin B6 0.05g, and inositol 10g per liter
B. The preparation method of the 6,7-V liquid culture medium comprises the following steps: taking 50mL of macroelement mother liquor, 50mL of CaCl2 mother liquor, 1mL of microelement mother liquor, 10mL of ferric salt mother liquor, 10mL of vitamin mother liquor and 30g of cane sugar; dissolving in 1L distilled water to adjust pH to 5.8.
Example 1
A method for mass production of hairy roots of Astragalus membranaceus comprises the following steps:
step 1. preparation of Astragalus aseptic seedling
Soaking Mongolian radix astragali seed in tap water for 24 hr, soaking in 70% ethanol for 3min, and washing with sterile water for 5 times; soaking in 1% mercuric chloride solution for 10 min; washing with sterile water for 5 times; inoculating to MS culture medium sterilized in advance, culturing at 25 deg.C and humidity of 60% to obtain radix astragali aseptic seedling;
step 2, strain activation and culture
Agrobacterium ATCC15834 was added to liquid YEB medium and cultured at 26 ℃ and-250 rpm to an OD600 value of 0.6; then, culturing the bacterial liquid in 1/2MS liquid culture medium, and culturing for 30min at 26 ℃ and 150 rpm;
step 3 Material Pre-culture and Co-culture
Transferring the leaves of the aseptic seedlings to an MS culture medium added with 0.3mg/L phytohormone IBA, culturing for 2 days in the dark, and inducing callus; then, fully contacting with the bacterial liquid in the step 2, and co-culturing for 40min at the temperature of 25 ℃ at 60 rpm;
step 4, degerming
Adding the callus obtained in the step 3 to 1/2MS solid culture medium containing 500mg/L cefotaxime sodium (cef), and culturing at 25 ℃ in the dark; replacing new culture medium at intervals of one week, and gradually reducing the concentration of cefotaxime sodium (cef) in the order of 400mg/L, 300mg/L, 200mg/L and 100 mg/L;
separating the roots after the roots reach 3cm, placing the roots on 1/2MS solid culture medium containing 50mg/L cefotaxime sodium (cef), subculturing once every other week for three times, and then placing the roots on 1/2MS solid culture medium without cefotaxime sodium (cef) for culture;
step 5, enlarged cultivation of hairy root
The hairy root of step 4 was transferred to 6,7-V liquid medium, expanded at 25 ℃, 100rpm, and subcultured every 18 days.
Example 2
A method for mass production of hairy roots of Astragalus membranaceus comprises the following steps:
step 1. preparation of Astragalus aseptic seedling
Soaking Astragalus membranaceus seed in tap water for 20 hr, soaking in 70% ethanol for 2min, and washing with sterile water for 5 times; soaking in 1% mercuric chloride solution for 8 min; washing with sterile water for 5 times; inoculating to MS culture medium sterilized in advance, culturing at 30 deg.C and humidity of 60% to obtain radix astragali aseptic seedling;
step 2, strain activation and culture
Adding agrobacterium LBA9402 into liquid YEB culture medium, culturing at 26 deg.C and 220rpm until OD600 value is 0.5; then, culturing the bacterial liquid in 1/2MS liquid culture medium, culturing for 30min at 28 ℃ and 180 rpm;
step 3 Material Pre-culture and Co-culture
Transferring the leafstalk of the aseptic seedling to an MS culture medium added with phytohormone NAA with the content of 0.2mg/L, culturing for 2 days in the dark, and inducing callus; then, fully contacting with the bacterial liquid in the step 2, and co-culturing for 30min at 50rpm and 25 ℃;
step 4, degerming
Adding the callus obtained in the step 3 to 1/2MS solid culture medium containing 500mg/L cefotaxime sodium (cef), and culturing at 25 ℃ in the dark; replacing new culture medium at intervals of one week, and gradually reducing the concentration of cefotaxime sodium (cef) in the order of 400mg/L, 300mg/L, 200mg/L and 100 mg/L;
separating the roots after the roots reach 3cm, placing the roots on 1/2MS solid culture medium containing 50mg/L cefotaxime sodium (cef), subculturing once every other week for three times, and then placing the roots on 1/2MS solid culture medium without cefotaxime sodium (cef) for culture;
step 5, enlarged cultivation of hairy root
The hairy root of step 4 was transferred to 6,7-V liquid medium, expanded at 25 ℃, 100rpm, and subcultured every 20 days.
Determination of astragaloside in hairy roots: the content of astragaloside in the examples 1-2, wild roots and field roots was determined by HPLC according to the test conditions of the version of the reference Chinese pharmacopoeia 2015, and the results of the related experiments are shown in Table 1:
table 1:
| numbering | Example 1 | Example 2 | Root of wild rootstalk | Root for field cultivation |
| Content of Astragalus glycoside (mg/g) | 2.36 | 2.25 | 2.30 | 1.87 |
From the results shown in Table 1, the method for mass production of Astragalus membranaceus hairy root of the present invention can obtain hairy root with high astragaloside content, which is similar to that of wild Astragalus membranaceus root.
It will be understood by those skilled in the art that the foregoing is only a preferred embodiment of the present invention, and is not intended to limit the invention, and that any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the scope of the present invention.
Claims (9)
1. A method for producing astragalus membranaceus hairy roots in a large scale is characterized by comprising the following steps: the method comprises the following steps:
step 1. preparation of Astragalus aseptic seedling
Soaking radix astragali seed in tap water for 20-24 hr, soaking in 70% ethanol for 1-3min, and washing with sterile water for 3-5 times; soaking in 1% mercuric chloride solution for 8-10 min; washing with sterile water for 5-10 times; inoculating to MS culture medium sterilized in advance, culturing at 25-30 deg.C and humidity of 50-60% to obtain sterile seedling of radix astragali;
step 2, strain activation and culture
Adding agrobacterium to a liquid YEB culture medium, culturing at 26-28 ℃ and 220-; then, culturing the bacterial liquid in 1/2MS liquid culture medium at the temperature of 26-28 ℃ and the rotation speed of 150-;
step 3 Material Pre-culture and Co-culture
Transferring the leaves, stems, hypocotyls and petioles of the aseptic seedlings to an MS culture medium added with phytohormone, culturing for 1-2 days in the dark, and inducing callus; then, fully contacting with the bacterial liquid in the step 2, and co-culturing for 30-40min at 50-60rpm and 25-28 ℃;
step 4, degerming
Adding the callus obtained in the step 3 to 1/2MS solid culture medium containing 500mg/L cefotaxime sodium (cef), and culturing at 25 ℃ in the dark; replacing the culture medium with new one week at intervals, and gradually reducing the concentration of cefotaxime sodium (cef);
separating the roots after the roots reach 3cm, placing the roots on 1/2MS solid culture medium containing 50mg/L cefotaxime sodium (cef), subculturing once every other week for three times, and then placing the roots on 1/2MS solid culture medium without cefotaxime sodium (cef) for culture;
step 5, enlarged cultivation of hairy root
The hairy roots of step 4 were transferred to 6,7-V liquid medium, expanded at 25-28 ℃ at 100-.
2. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the radix astragali seed is selected from one of Mongolian radix astragali seed or membrane pod radix astragali seed.
3. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the composition per liter of the MS medium was as follows:
macroelements: KNO 34-6 mM, Ca (NO3) 2.4H 2O 4-6mM, MgSO 4.7H 2O 1.5.5-2.5 mM, KH2PO40.8-1.2mM,
trace elements: h3BO 344-48 μ M, MnCl 2.4H2O 8-10 μ M, ZnSO 4.7H2O 0.6-0.8 μ M, CuSO 4.5H2O 0.2-0.4 μ M, Na2MoO 4.2H2O 0.1-0.3 μ M, EDTA-Fe-Na 40-60 μ M;
organic matter: inositol 80-100. mu.M, sucrose 100-150. mu.M, and hydrolyzed casein 300-500. mu.M.
4. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the 1/2MS culture medium is composed by halving the amount of macroelements in the MS culture medium.
5. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the agrobacterium strain is selected from ATCC15834, LBA 9402.
6. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the plant hormone in the step 3 is one of IBA and NAA, and the content is 0.1-0.3 mg/L.
7. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the preparation method of the liquid YEB culture medium comprises the following steps: dissolving yeast powder 1g, beef extract 5g, peptone 5g, sucrose 5g, MgSO4 & 7H2O 0.5g in 1L distilled water, adjusting pH to 7.4 with sodium hydroxide, and autoclaving at 121 deg.C for 20 min.
8. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the composition and preparation method of the 6,7-V culture medium are as follows:
A. preparation of component mother liquor
(1) Macroelement mother liquor: the mother liquor contains 16g KNO3, 2g (NH4)2SO4, 5g MgSO4 & 7H2O, 3g NaH2PO4, 4g KCl and 0.4g Na2HPO4 per liter;
(2) CaCl2 mother liquor: the mother liquor contains 4g of CaCl2 & 2H2O per liter
(3) And (3) a microelement mother solution: the mother liquor contains 4g of MnSO4 & H2O, 1.5g of ZnSO4 & 7H2O, 5g H3BO3, 0.05gKI, 0.25g of NaMoO4 & 2H2O, 0.25g of CuSO4 & 5H2O and 0.25g of CoCl2 & 6H2O per liter
(4) Mother liquor of iron salt: the mother liquor contains 1.86g of EDTA-Na2 and 1.39g of FeSO 4.7H 2O per liter
(5) Vitamin mother liquor: the mother liquor contains 0.125g nicotinic acid, 0.05g vitamin B1, 0.05g vitamin B6 and 10g inositol per liter;
B. the preparation method of the 6,7-V liquid culture medium comprises the following steps: taking 50mL of macroelement mother liquor, 50mL of CaCl2 mother liquor, 1mL of microelement mother liquor, 10mL of ferric salt mother liquor, 10mL of vitamin mother liquor and 30g of cane sugar; dissolving in 1L distilled water to adjust pH to 5.8.
9. The method for mass production of hairy roots of astragalus membranaceus as claimed in claim 1, wherein: the step-wise reduction of the concentration of cefotaxime sodium (cef) in the step 4 is performed according to the modes of 400mg/L, 300mg/L, 200mg/L and 100 mg/L.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111587785A (en) * | 2020-05-06 | 2020-08-28 | 内蒙古自治区生物技术研究院 | Method for culturing astragalus membranaceus hairy roots for promoting accumulation of flavonoids |
| CN116114495A (en) * | 2023-02-24 | 2023-05-16 | 青岛大学 | Method for infecting sterile black pine seedlings by pine wood nematodes |
| CN116711741A (en) * | 2023-07-27 | 2023-09-08 | 内蒙古多肽科技有限责任公司 | Application of microbial preparation in astragalus membranaceus planting |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003030822A2 (en) * | 2001-10-05 | 2003-04-17 | Wu, Bertolotti, Jy-Yuan | Pharmaceutical composition having prophylactic effects on lamivudine-related disease relapse and drug resistance and methods of using the same |
| CN1421528A (en) * | 2002-12-17 | 2003-06-04 | 上海中医药大学 | Method of raising astragalus polysaccharide content in hairy root of astragalus |
| CN102229945A (en) * | 2011-05-20 | 2011-11-02 | 陕西科技大学 | Cultivation method for inducing scutellaria baicalensis hairy root based on agrobacterium rhizogenes infection |
| CN103451146A (en) * | 2013-09-11 | 2013-12-18 | 南京泽朗农业发展有限公司 | Rapid propagation method of astragalus membranaceus calluses and astragalus membranaceus suspension cells |
| CN105087635A (en) * | 2015-08-14 | 2015-11-25 | 贵州大学 | Method for inducing hairy roots of anisodus acutangulus C.Y.Wu et C.Chen |
-
2020
- 2020-03-02 CN CN202010135220.0A patent/CN111226793A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003030822A2 (en) * | 2001-10-05 | 2003-04-17 | Wu, Bertolotti, Jy-Yuan | Pharmaceutical composition having prophylactic effects on lamivudine-related disease relapse and drug resistance and methods of using the same |
| CN1421528A (en) * | 2002-12-17 | 2003-06-04 | 上海中医药大学 | Method of raising astragalus polysaccharide content in hairy root of astragalus |
| CN102229945A (en) * | 2011-05-20 | 2011-11-02 | 陕西科技大学 | Cultivation method for inducing scutellaria baicalensis hairy root based on agrobacterium rhizogenes infection |
| CN103451146A (en) * | 2013-09-11 | 2013-12-18 | 南京泽朗农业发展有限公司 | Rapid propagation method of astragalus membranaceus calluses and astragalus membranaceus suspension cells |
| CN105087635A (en) * | 2015-08-14 | 2015-11-25 | 贵州大学 | Method for inducing hairy roots of anisodus acutangulus C.Y.Wu et C.Chen |
Non-Patent Citations (4)
| Title |
|---|
| 李巧峡主编: "《发育生物学实验指导》", 31 August 2019, 甘肃科学技术出版社 * |
| 李雅丽等: ""用农杆菌Ri诱导蒙古黄芪发根培养的研究"", 《云南植物研究》 * |
| 杜旭红: ""两种丹参毛状根的快速培养及其生物反应器初步研究"", 《中国优秀硕士学位论文全文数据库 农业科技辑》 * |
| 焦骄: ""黄芪毛状根培养体系的优化及其主要活性成分生物合成的诱导调控研究"", 《中国博士学位论文全文数据库 农业科技辑》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111587785A (en) * | 2020-05-06 | 2020-08-28 | 内蒙古自治区生物技术研究院 | Method for culturing astragalus membranaceus hairy roots for promoting accumulation of flavonoids |
| CN116114495A (en) * | 2023-02-24 | 2023-05-16 | 青岛大学 | Method for infecting sterile black pine seedlings by pine wood nematodes |
| CN116711741A (en) * | 2023-07-27 | 2023-09-08 | 内蒙古多肽科技有限责任公司 | Application of microbial preparation in astragalus membranaceus planting |
| CN116711741B (en) * | 2023-07-27 | 2023-11-24 | 内蒙古多肽科技有限责任公司 | Application of microbial preparation in astragalus membranaceus planting |
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Application publication date: 20200605 |