CN106818491A - A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide - Google Patents
A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide Download PDFInfo
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- CN106818491A CN106818491A CN201710226205.5A CN201710226205A CN106818491A CN 106818491 A CN106818491 A CN 106818491A CN 201710226205 A CN201710226205 A CN 201710226205A CN 106818491 A CN106818491 A CN 106818491A
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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
Abstract
The invention discloses a kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, including following raw material：Sugar, GINKGO BILOBA EXTRACT, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, conditioner, dispersant, synergist, stabilizer, antiseptic, water.Reticulate millettia tissue-cultured seedling culture medium of the invention, collocation is reasonable, comprehensive nutrition, quickly promoting root growth can give birth to stem, improves and transplants survival rate, while can also promote the accumulation of flavone compound and polysaccharide, increases economic efficiency.
Description
Technical field
The invention belongs to reticulate millettia planting technology field, and in particular to one kind is for improving reticulate millettia tissue-cultured seedling flavonoids
The culture medium of compound and polysaccharide.
Background technology
Reticulate millettia scientific name is Spatholobus suberectus Dunn, belongs to Dicotyledoneae pulse family Kadsura
Fruit of Chinese magnoliavine kind, is distributed mainly on the ground such as Fujian, Guangdong, Guangxi, Yunnan, is grown in the woods below 1800 meters of height above sea level.
The medicinal part of reticulate millettia is to dry root and rhizome, and multiple compounds are contained in reticulate millettia, and flavones ingredient is it
Main active component, additionally containing polysaccharide etc., cures mainly rheumatism tibialgia, irregular menstruation, blood deficiency chlorosis, paralysis and numbness, wind
Arthritis with fixed pain caused by dampness pain etc., is China's traditional Chinese medicine, is the raw material of many clinical applications, and market demand is huge, reticulate millettia impoverishment tolerant,
Strong adaptability, is also urban afforestation and stony desertification, the preferable liana kind of desertification treatment sometimes up to tens of rice.
In recent years because ecological influence and artificial unordered unrest are adopted, the protection of resource is not focused on so that wild chicken blood
Rattan quantity is fewer and feweri, and resource is increasingly reduced.To protect and developing this excellent herbal variety, people's medication demand is met,
Propagation method main at present is natural propagation, and tissue culture technology is simple to operate, and breeding potential is high, and right these natural propagation methods are not
The accumulation of reticulate millettia tissue-cultured seedling flavone compound and the raising of polysaccharide can be promoted.
The content of the invention
The present invention provides a kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, existing to solve
How culture medium improves the principle active component flavone compound of reticulate millettia and the content of polysaccharide in a organized way, increases economic efficiency
The problems such as.Reticulate millettia tissue-cultured seedling culture medium of the invention, collocation is reasonable, comprehensive nutrition, quickly promoting root growth can give birth to stem, improves and transplants
Survival rate, while can also promote the accumulation of flavone compound and polysaccharide, increases economic efficiency.
To solve above technical problem, the present invention uses following technical scheme：
A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, in units of weight portion, including with
Lower raw material：Sugared 35-53 parts, GINKGO BILOBA EXTRACT 12-19 parts, sylvite 2.2-2.9 parts, microcosmic salt 1.3-2.8 parts, nitrogen salt 1.1-1.9 parts, calcium
Salt 0.6-0.8 parts, magnesium salts 0.6-0.8 parts, salt compounded of iodine 0.2-0.4 parts, boron salt 0.2-0.4 parts, 0.2-0.4 parts of molybdenum salt, zinc salt 0.2-
0.4 part, manganese salt 0.2-0.4 parts, mantoquita 0.2-0.4 parts, molysite 0.2-0.4 parts, vitamin 0.1-0.25 parts, amino acid 0.08-
0.24 part, auxin 0.05-0.12 parts, gibberellin 0.06-0.09 parts, succimide 0.04-0.08 parts, the basic element of cell division
0.03-0.07 parts, kinetin 0.04-0.09 parts, conditioner 0.13-0.22 parts, dispersant 1.2-2.3 parts, synergist 0.02-
0.05 part, stabilizer 0.9-1.8 parts, antiseptic 1.5-2.6 parts, water 2000-3000 parts；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：Sodium carboxymethylcellulose 3-5 parts, modified starch 4-7 parts,
Polyacrylic acid potassium 2.5-4.6 parts, potassium alginate 1.2-3.4 parts；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 40-80 mesh sieve is crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 62-75 DEG C of water, solid-to-liquid ratio is 1 in extraction:12-
16（W/V）, microwave power is 200-800W, and extraction time is 15-30min, is extracted 2-3 times, filtering, merging filtrate after extraction,
Mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction 5.5-6 times of volume of cream
52-55 DEG C of water dissolved, extraction 2-3 time is carried out using water saturated n-butanol, continue to be entered with water saturated n-butanol
The multiple crystallization treatment of row, solid-to-liquid ratio is 1 in crystallization process:5.2-5.5（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is 21-
23MPa, extraction temperature is 51-54 DEG C, and extraction time is 25-28min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Reason agent, dispersant, synergist, water mixing, are 110-140W in microwave power, and temperature is 40-46 DEG C, and rotating speed is 200-300r/
22-32min is stirred under min, mixed liquor a is obtained；
S3：It it is 40-43 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is 100-
11-16min is stirred under 200r/min, mixed liquor b is obtained；
S4：The pH value of mixed liquor b obtained in step S3 is adjusted to 5.6-6.2, then using pulse vacuum pressure-steam sterilization
Device, is 134-136 DEG C in temperature, and pressure is the 1-1.4min that sterilized under 210-230KPa, is obtained for improving reticulate millettia tissue-cultured seedling
The culture medium of flavone compound and polysaccharide.
Preferably, the synergist is Octacide 264 diethyl aminoethyl hexanoate.
Preferably, the stabilizer is xanthans.
Preferably, the antiseptic is methyl-furfuryl fumarate.
Preferably, the extracting condition of Ginkgo Leaf is in step S12：Extracted using 70 DEG C of water, it is solid in extraction
Liquor ratio is 1:14（W/V）, microwave power is 600W, and extraction time is 22min, is extracted 3 times.
Preferably, the condition of extraction is in step S14：Extraction kettle pressure is 22MPa, and extraction temperature is 53 DEG C, during extraction
Between be 26min.
Preferably, temperature described in step S4 is 136 DEG C, and pressure is the 1minn that sterilized under 230KPa.
The effect of culture medium fractions of the present invention is as follows：
The accumulation being added with beneficial to polyoses content of sugar.
GINKGO BILOBA EXTRACT can promote accumulation of flavonoids.
The nutritive effect of potassium element can improve photosynthetic intensity, promote the formation of starch and sugar in crop body, increase
Pretend the resistance and resistance against diseases of thing, moreover it is possible to improve crop and nitrogen is absorbed.
Phosphorus is to form the indispensable element such as Nuclear extract, lecithin.P elements can accelerate cell division, promote root
System and overground part accelerate growth, promote bud differentiation, ripe ahead of time, improve fruit quality.
Nitrogen is the essential element that protein is constituted, and protein is the base substance in cellular plasm composition.Nitrogen energy
Promote the formation of protein and chlorophyll, make dark green leaf color, leaf area increase promotes the assimilation of carbon, is conducive to yield to increase, product
Matter improves.
The growth for lacking calcium plant meristem is slack-off, and, just in growing point and spire, when serious, leaf deforms for calcium deficiency and disease
And chlorosis, there is necrotic spot at the edge of leaf.
Magnesium is the constituent of chlorophyll.If lacking magnesium, blade loses green, it is impossible to manufacture nutriment.
Iodine is the indispensable element of plant growth, is described as biological element, wisdom element.
During boron compares in plant and concentrates on stem apex, the tip of a root, blade and floral organ, the content in dicotyledon is usually
Higher than monocotyledon.Boron can improve the nitrogenase activity of legume rhizobium, increase amount of nitrogen fixation, and during boron deficiency, root is stayed not to be sent out
Reach, influence amount of nitrogen fixation.Boron can also strengthen crop anti-adversity.
Molybdenum is nitrogen-fixing microorganism, particularly fixes necessary during air nitrogen with the rhizobium of legume symbiosis.Again simultaneously
Leaf photosynthesis intensity can be promoted.Plant is short and small when lacking molybdenum, and chlorosis between blade arteries and veins, leaf margin is dried-up, inward curl, into wilting
State.
Zinc can promote the crop to carry out photosynthesis, and it is the constituent of various enzymes, and stagnating occurs in crop growth during zinc-deficiency.
Contain manganese in plant chloroplast, manganese can promote germination and seedling period to grow, the light cooperation of crop is influenceed during manganese deficiency
With, respiration, nitrate nitrogen accumulation in vivo.Chlorosis yellow between blade arteries and veins is shown as, there is brown spot.
Copper participates in photosynthesis, respiration, nitrogen metabolism, the resistance of enhancing plant of plant.Plant strain growth is thin and weak when lacking copper, newly
Leaf is turned to be yellow, wilting withered, and blade tip curling is turned white, and has necrotic spot.
Iron is indispensable chlorophyll formation, transfer is difficult in plant body, so blade " chlorosis " is plant iron deficiency
Show, and this chlorosis is shown on young leaflet tablet first.
Vitamin has effect to taking root for plant, can promote plant establishment.
Amino acid is nutriment necessary to plant growth, special to plant growth due to amino acid characteristic in itself
It is that photosynthesis has unique facilitation, it can increase chlorophyll content of plant, improve the activity of enzyme, promote titanium dioxide
The infiltration of carbon, makes photosynthesis more vigorous, and to improving crop quality, the content for increasing Vc and sugar suffers from important function.
Auxin can promote cell elongation to grow.
Gibberellin, is the class plant hormone being widely present.Its chemical constitution belongs to Diterpenes acid, is derived by four ring skeletons
And obtain, the growth of leaf and bud can be stimulated.
Succimide is a kind of plant growth substance, can promote plant cell growth, improves the content of amino acid.
The basic element of cell division promotes the material of cytokinesis, promotes the differentiation and growth of Various Tissues, is have with plant growth
Synergy.
Kinetin is a kind of non-natural basic element of cell division, and chemical name is 6- glycosyls adenine phosphate (or N6- furfuryls
Adenine).It is water insoluble, it is dissolved in strong acid, alkali and glacial acetic acid；In addition to fissional effect is promoted, also with delaying
Excised leaf and cut flower withering, induced bud differentiation and development and increase the effect of stomatal aperture.
Conditioner can promote the development and beneficial microorganism activity of root system.
Dispersant can prevent flocculation, prevent sedimentation.
Synergist has the high energy plant growth regulator of wide spectrum and breakthrough effect.It can improve plant peroxidases
With the activity of nitrate reductase, the content quickening photosynthetic speed of chlorophyll is improved, promote the division and elongation of plant cell, promoted
The development of root system, adjusts the balance of internal nutrient.
Stabilizer can increase the stability of solution, colloid, solid, mixture.
Antiseptic is can to make certain micro-organisms within a certain period of time（Bacterium, fungi, saccharomycete, algae and virus etc.）
Growth or breeding be maintained at necessary chemical substance below horizontal.
The invention has the advantages that：
（1）GINKGO BILOBA EXTRACT can promote the accumulation of reticulate millettia flavone compound in the present invention；
（2）Sugar can promote the accumulation of reticulate millettia polysaccharide；Simultaneously in culture medium sugar-free, or sugar matches somebody with somebody effect with other nutrient media componentses 5
Under, it is little to the effect of reticulate millettia tissue-cultured seedling height growth；
（3）Culture medium of the invention is conducive to improving the transplanting survival rate of reticulate millettia tissue-cultured seedling, reaches 99%-100%.
Specific embodiment
For ease of more fully understanding the present invention, it is illustrated by following examples, these embodiments belong to of the invention
Protection domain, but do not limit the scope of the invention.
In embodiment, the culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, with weight
Part it is unit, including following raw material：It is sugared 35-53 parts, GINKGO BILOBA EXTRACT 12-19 parts, sylvite 2.2-2.9 parts, microcosmic salt 1.3-2.8 parts,
Nitrogen salt 1.1-1.9 parts, calcium salt 0.6-0.8 parts, magnesium salts 0.6-0.8 parts, salt compounded of iodine 0.2-0.4 parts, boron salt 0.2-0.4 parts, molybdenum salt
0.2-0.4 parts, zinc salt 0.2-0.4 parts, manganese salt 0.2-0.4 parts, mantoquita 0.2-0.4 parts, molysite 0.2-0.4 parts, vitamin 0.1-
0.25 part, amino acid 0.08-0.24 parts, auxin 0.05-0.12 parts, gibberellin 0.06-0.09 parts, succimide 0.04-
0.08 part, basic element of cell division 0.03-0.07 parts, kinetin 0.04-0.09 parts, conditioner 0.13-0.22 parts, dispersant 1.2-
2.3 parts, synergist 0.02-0.05 parts, stabilizer 0.9-1.8 parts, antiseptic 1.5-2.6 parts, water 2000-3000 parts；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：Sodium carboxymethylcellulose 3-5 parts, modified starch 4-7 parts,
Polyacrylic acid potassium 2.5-4.6 parts, potassium alginate 1.2-3.4 parts；
The synergist is Octacide 264 diethyl aminoethyl hexanoate；
The stabilizer is carragheen；
The antiseptic is methyl-furfuryl fumarate；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 40-80 mesh sieve is crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 62-75 DEG C of water, solid-to-liquid ratio is 1 in extraction:12-
16（W/V）, microwave power is 200-800W, and extraction time is 15-30min, is extracted 2-3 times, filtering, merging filtrate after extraction,
Mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction 5.5-6 times of volume of cream
52-55 DEG C of water dissolved, extraction 2-3 time is carried out using water saturated n-butanol, continue to be entered with water saturated n-butanol
The multiple crystallization treatment of row, solid-to-liquid ratio is 1 in crystallization process:5.2-5.5（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is 21-
23MPa, extraction temperature is 51-54 DEG C, and extraction time is 25-28min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Reason agent, dispersant, synergist, water mixing, are 110-140W in microwave power, and temperature is 40-46 DEG C, and rotating speed is 200-300r/
22-32min is stirred under min, mixed liquor a is obtained；
S4：It it is 40-43 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is 100-
11-16min is stirred under 200r/min, mixed liquor b is obtained；
S5：The pH value of mixed liquor b obtained in step S4 is adjusted to 5.6-6.2, then using pulse vacuum pressure-steam sterilization
Device, is 134-136 DEG C in temperature, and pressure is the 1-1.4min that sterilized under 210-230KPa, is obtained for improving reticulate millettia tissue-cultured seedling
The culture medium of flavone compound and polysaccharide.
Embodiment 1
A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, in units of weight portion, including with
Lower raw material：45 parts of sugar, 15 parts of GINKGO BILOBA EXTRACT, 2.5 parts of sylvite, 2 parts of microcosmic salt, 1.5 parts of nitrogen salt, 0.7 part of calcium salt, 0.7 part of magnesium salts, iodine
03 part of salt, 0.3 part of boron salt, 0.3 part of molybdenum salt, 0.3 part of zinc salt, 0.3 part of manganese salt, 0.3 part of mantoquita, 0.3 part of molysite, vitamin 0.18
Part, 0.18 part of amino acid, 0.08 part of auxin, 0.08 part of gibberellin, 0.06 part of succimide, 0.05 part of the basic element of cell division,
0.07 part of kinetin, 0.18 part of conditioner, 1.8 parts of dispersant, 0.04 part of synergist, 1.4 parts of stabilizer, 2 parts of antiseptic, water
2500 parts；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：4 parts of sodium carboxymethylcellulose, 6 parts of modified starch, poly- third
4.2 parts of olefin(e) acid potassium, 2.3 parts of potassium alginate；
The synergist is Octacide 264 diethyl aminoethyl hexanoate；
The stabilizer is carragheen；
The antiseptic is methyl-furfuryl fumarate；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 80 mesh sieve are crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 70 DEG C of water, solid-to-liquid ratio is 1 in extraction:14（W/
V）, microwave power is 600W, and extraction time is 22min, is extracted 3 times, filtering, merging filtrate after extraction, and mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction cream 6 times the 55 of volume
DEG C water dissolved, using water saturated n-butanol carry out extraction 2 times, continuation repeatedly crystallized with water saturated n-butanol
Treatment, solid-to-liquid ratio is 1 in crystallization process:5.5（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is
22MPa, extraction temperature is 53 DEG C, and extraction time is 26min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Agent, dispersant, synergist, water mixing are managed, is 130W in microwave power, temperature is 44 DEG C, and rotating speed is stirring under 200r/min
26min, is obtained mixed liquor a；
S3：It it is 42 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is under 100r/min
Stirring 14min, is obtained mixed liquor b；
S4：The pH value of mixed liquor b obtained in step S3 is adjusted to 5.8, then using pulse vacuum pressure steam sterilizer,
Temperature is 135 DEG C, and pressure is to sterilize 1.2min under 220KPa, is obtained for improving reticulate millettia tissue-cultured seedling flavone compound and many
The culture medium of sugar.
Embodiment 2
A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, in units of weight portion, including with
Lower raw material：Sugar 36 parts, 12 parts of GINKGO BILOBA EXTRACT, 2.2 parts of sylvite, 1.3 parts of microcosmic salt, nitrogen salt 1.1-2 parts, 0.6 part of calcium salt, magnesium salts 0.6
Part, 0.2 part of salt compounded of iodine, 0.2 part of boron salt, 0.2 part of molybdenum salt, 0.2 part of zinc salt, 0.2 part of manganese salt, 0.2 part of mantoquita, 0.2 part of molysite, dimension life
0.1 part of element, 0.08 part of amino acid, 0.05 part of auxin, 0.06 part of gibberellin, 0.04 part of succimide, the basic element of cell division
0.03 part, 0.04 part of kinetin, 0.13 part of conditioner, 1.2 parts of dispersant, 0.02 part of synergist, 0.9 part of stabilizer, antiseptic
1.5 parts, 2000 parts of water；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：3 parts of sodium carboxymethylcellulose, 4 parts of modified starch, poly- third
2.5 parts of olefin(e) acid potassium, 1.2 parts of potassium alginate；
The synergist is Octacide 264 diethyl aminoethyl hexanoate；
The stabilizer is carragheen；
The antiseptic is methyl-furfuryl fumarate；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 40 mesh sieve are crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 62 DEG C of water, solid-to-liquid ratio is 1 in extraction:12（W/
V）, microwave power is 200W, and extraction time is 30min, is extracted 2 times, filtering, merging filtrate after extraction, and mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction 5.5 times of volumes of cream
52 DEG C of water is dissolved, and extraction 3 times is carried out using water saturated n-butanol, and continuation is repeatedly tied with water saturated n-butanol
Crystalline substance treatment, solid-to-liquid ratio is 1 in crystallization process:5.5（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is
21MPa, extraction temperature is 51 DEG C, and extraction time is 25min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Agent, dispersant, synergist, water mixing are managed, is 110W in microwave power, temperature is 40 DEG C, and rotating speed is stirring under 200r/min
32min, is obtained mixed liquor a；
S3：It it is 40 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is under 100r/min
Stirring 16min, is obtained mixed liquor b；
S4：The pH value of mixed liquor b obtained in step S3 is adjusted to 5.6, then using pulse vacuum pressure steam sterilizer,
Temperature is 134 DEG C, and pressure is to sterilize 1.4min under 210KPa, is obtained for improving reticulate millettia tissue-cultured seedling flavone compound and many
The culture medium of sugar.
Embodiment 3
A kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, in units of weight portion, including with
Lower raw material：53 parts of sugar, 19 parts of GINKGO BILOBA EXTRACT, 2.9 parts of sylvite, 2.8 parts of microcosmic salt, 1.9 parts of nitrogen salt, 0.8 part of calcium salt, magnesium salts -0.8
Part, -0.4 part of salt compounded of iodine, -0.4 part of boron salt, 0.4 part of molybdenum salt, 0.4 part of zinc salt, 0.4 part of manganese salt, 0.4 part of mantoquita, 0.4 part of molysite, dimension
Raw 0.25 part of element, 0.24 part of amino acid, 0.12 part of auxin, 0.09 part of gibberellin, 0.08 part of succimide, the basic element of cell division
0.07 part, 0.09 part of kinetin, 0.22 part of conditioner, 2.3 parts of dispersant, 0.05 part of synergist, 1.8 parts of stabilizer, antiseptic
2.6 parts, 3000 parts of water；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：5 parts of sodium carboxymethylcellulose, 7 parts of modified starch, poly- third
4.6 parts of olefin(e) acid potassium, 3.4 parts of potassium alginate；
The synergist is Octacide 264 diethyl aminoethyl hexanoate；
The stabilizer is carragheen；
The antiseptic is methyl-furfuryl fumarate；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 60 mesh sieve are crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 70 DEG C of water, solid-to-liquid ratio is 1 in extraction:14（W/
V）, microwave power is 500W, and extraction time is 25min, is extracted 3 times, filtering, merging filtrate after extraction, and mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction 5.8 times of volumes of cream
53 DEG C of water is dissolved, and extraction 3 times is carried out using water saturated n-butanol, and continuation is repeatedly tied with water saturated n-butanol
Crystalline substance treatment, solid-to-liquid ratio is 1 in crystallization process:5.3（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is
23MPa, extraction temperature is 54 DEG C, and extraction time is 25min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Agent, dispersant, synergist, water mixing are managed, is 140W in microwave power, temperature is 46 DEG C, and rotating speed is stirring under 300r/min
22min, is obtained mixed liquor a；
S3：It it is 43 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is under 200r/min
Stirring 11min, is obtained mixed liquor b；
S4：The pH value of mixed liquor b obtained in step S3 is adjusted to 6.2, then using pulse vacuum pressure steam sterilizer,
Temperature is 136 DEG C, and pressure is the 1min that sterilized under 230KPa, is obtained for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide
Culture medium.
To verify the effect of culture medium of the present invention, by following experimental verification.
Every group of reticulate millettia stem with bud quantity 100 from the basically identical 2-3cm of specification, control group 1 with sugar, sylvite,
Microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin,
Gibberellin, succimide, the basic element of cell division, kinetin, conditioner, dispersant, synergist, stabilizer, antiseptic, water composition
Culture medium, control group 2 individually with GINKGO BILOBA EXTRACT as culture medium, other using embodiment 1-3 culture medium.After culture 110 days
Measurement reticulate millettia tissue-cultured seedling flavone compound average yield and length, the results are shown in Table 1.
The culture medium tissue culture reticulate millettia tissue-cultured seedling flavone compound average yield of table 1 and length situation
| Embodiment | Experiment tissue-cultured seedling number | The average yield of flavone compound（%） | Reticulate millettia tissue-cultured seedling length（cm） |
| 1 | 100 | 1.01 | 5.55 |
| 2 | 100 | 0.99 | 5.52 |
| 3 | 100 | 1.03 | 5.57 |
| Control group 1 | 100 | 0.76 | 5.68 |
| Control group 2 | 100 | / | It is almost unchanged |
As shown in Table 1, the reticulate millettia group of the reticulate millettia tissue-cultured seedling flavone compound average yield than control group 1 of embodiment 1-3
Training seedling flavone compound average yield 30.26%-35.53% high；The reticulate millettia tissue-cultured seedling length of embodiment 1-3 is than control group 1
The reticulate millettia short 1.94%-2.82% of tissue-cultured seedling length, reticulate millettia hardly grows control group 2, illustrates the presence of GINKGO BILOBA EXTRACT
Reticulate millettia tissue-cultured seedling flavone compound average yield can be greatly improved.
Every group of reticulate millettia stem with bud quantity 100 from the basically identical 2-3 cm of specification, control group with GINKGO BILOBA EXTRACT,
Sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc salt, manganese salt, mantoquita, molysite, vitamin, amino acid, life
Element long, gibberellin, succimide, the basic element of cell division, kinetin, conditioner, dispersant, synergist, stabilizer, antiseptic,
The culture medium of water composition, other use the culture medium of embodiment 1-3.Culture measures the polysaccharide average yield of tissue-cultured seedling after 110 days
And length, the results are shown in Table 2.
The culture medium tissue culture reticulate millettia tissue-cultured seedling average yield of table 2 and length situation
| Embodiment | Experiment tissue-cultured seedling number | Polysaccharide average yield（%） | Reticulate millettia tissue-cultured seedling length（cm） |
| 1 | 100 | 15.11 | 5.55 |
| 2 | 100 | 15.03 | 5.52 |
| 3 | 100 | 15.19 | 5.57 |
| Control group | 100 | 12.36 | 5.47 |
As shown in Table 2, the reticulate millettia tissue-cultured seedling polysaccharide average yield of embodiment 1-3 is put down than the reticulate millettia tissue-cultured seedling polysaccharide of control group
Equal yield 21.6%-22.9% high；The reticulate millettia tissue-cultured seedling length of embodiment 1-3 is shorter than the reticulate millettia tissue-cultured seedling length of control group
0.91%-1.83%, illustrates culture medium sugar-free, or sugar is matched somebody with somebody under effect with other nutrient media componentses 5, to reticulate millettia tissue culture height of seedling
Degree accretion is little.
Table 3 is embodiment 1-3 culture medium tissue culture reticulate millettia tissue culture transplantation of seedlings survival conditions.
The culture medium tissue culture reticulate millettia tissue culture transplantation of seedlings survival condition of table 3
| Embodiment | Experiment tissue-cultured seedling number | Tissue culture transplantation of seedlings survival number | Transplant survival rate（%） |
| 1 | 100 | 100 | 100 |
| 2 | 100 | 99 | 99 |
| 3 | 100 | 99 | 99 |
By table 3 it is recognised that with the average transplanting survival rate of culture medium tissue culture reticulate millettia tissue-cultured seedling of the invention as 99%-100%,
Illustrate that culture medium of the present invention is conducive to improving the transplanting survival rate of reticulate millettia tissue-cultured seedling.
The specific implementation of the invention is not to be limited to these illustrations for above content, is led for technology belonging to the present invention
For the those of ordinary skill in domain, without departing from the inventive concept of the premise, some simple deduction or replace can also be made,
The scope of patent protection that the claims of the present invention by being submitted to determine should be all considered as belonging to.
Claims (7)
1. a kind of culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, it is characterised in that with weight portion
It is unit, including following raw material：Sugared 35-53 parts, GINKGO BILOBA EXTRACT 12-19 parts, sylvite 2.2-2.9 parts, microcosmic salt 1.3-2.8 parts, nitrogen
Salt 1.1-1.9 parts, calcium salt 0.6-0.8 parts, magnesium salts 0.6-0.8 parts, salt compounded of iodine 0.2-0.4 parts, boron salt 0.2-0.4 parts, molybdenum salt 0.2-
0.4 part, zinc salt 0.2-0.4 parts, manganese salt 0.2-0.4 parts, mantoquita 0.2-0.4 parts, molysite 0.2-0.4 parts, vitamin 0.1-0.25
Part, amino acid 0.08-0.24 parts, auxin 0.05-0.12 parts, gibberellin 0.06-0.09 parts, succimide 0.04-0.08
Part, basic element of cell division 0.03-0.07 parts, kinetin 0.04-0.09 parts, conditioner 0.13-0.22 parts, dispersant 1.2-2.3 parts,
Synergist 0.02-0.05 parts, stabilizer 0.9-1.8 parts, antiseptic 1.5-2.6 parts, water 2000-3000 parts；
The sugar is fructose；
The sylvite is potassium sulfate；
The microcosmic salt is calcium phosphate；
The nitrogen salt is ammonium hydrogen carbonate；
The calcium salt is calcium glycine；
The magnesium salts is magnesium glycinate；
The salt compounded of iodine is KI；
The boron salt is potassium borate；
The molybdenum salt is potassium molybdate；
The zinc salt is zinc chloride；
The manganese salt is Mn-Gly；
The mantoquita is cupric glycinate；
The molysite is ferrous sulfate；
The vitamin is vitamin B1；
The amino acid is glycine；
The conditioner is humic acid；
The dispersant is in units of weight portion, including following raw material：Sodium carboxymethylcellulose 3-5 parts, modified starch 4-7 parts,
Polyacrylic acid potassium 2.5-4.6 parts, potassium alginate 1.2-3.4 parts；
The culture medium for improving reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its preparation method includes following step
Suddenly：
S1：The preparation of GINKGO BILOBA EXTRACT, comprises the following steps：
S11：Dry Folium Ginkgo is crushed and 40-80 mesh sieve is crossed, Ginkgo Leaf is obtained；
S12：Ginkgo Leaf obtained in step S11 is extracted using 62-75 DEG C of water, solid-to-liquid ratio is 1 in extraction:12-
16（W/V）, microwave power is 200-800W, and extraction time is 15-30min, is extracted 2-3 times, filtering, merging filtrate after extraction,
Mixing leaching liquor is obtained；
S13：Leaching liquor concentrate drying will be mixed obtained in step S12, extraction cream is obtained, be used in extraction 5.5-6 times of volume of cream
52-55 DEG C of water dissolved, extraction 2-3 time is carried out using water saturated n-butanol, continue to be entered with water saturated n-butanol
The multiple crystallization treatment of row, solid-to-liquid ratio is 1 in crystallization process:5.2-5.5（W/V）, crystal crude product is obtained；
S14：By crystal crude product obtained in step S13 by supercritical CO2Extraction, the condition of extraction：Extraction kettle pressure is 21-
23MPa, extraction temperature is 51-54 DEG C, and extraction time is 25-28min, and GINKGO BILOBA EXTRACT is obtained；
S2：By GINKGO BILOBA EXTRACT, sugar, sylvite, microcosmic salt, nitrogen salt, calcium salt, magnesium salts, salt compounded of iodine, boron salt, molybdenum salt, zinc obtained in step S1
Salt, manganese salt, mantoquita, molysite, vitamin, amino acid, auxin, gibberellin, succimide, the basic element of cell division, kinetin, tune
Reason agent, dispersant, synergist, water mixing, are 110-140W in microwave power, and temperature is 40-46 DEG C, and rotating speed is 200-300r/
22-32min is stirred under min, mixed liquor a is obtained；
S3：It it is 40-43 DEG C in temperature to stabilizer, antiseptic is added in mixed liquor a obtained in step S2, rotating speed is 100-
11-16min is stirred under 200r/min, mixed liquor b is obtained；
S4：The pH value of mixed liquor b obtained in step S3 is adjusted to 5.6-6.2, then using pulse vacuum pressure-steam sterilization
Device, is 134-136 DEG C in temperature, and pressure is the 1-1.4min that sterilized under 210-230KPa, is obtained for improving reticulate millettia tissue-cultured seedling
The culture medium of flavone compound and polysaccharide.
2. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that the synergist is Octacide 264 diethyl aminoethyl hexanoate.
3. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that the stabilizer is xanthans.
4. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that the antiseptic is methyl-furfuryl fumarate.
5. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that the extracting condition of Ginkgo Leaf is in step S12：Extracted using 70 DEG C of water, solid-to-liquid ratio is 1 in extraction:14
（W/V）, microwave power is 600W, and extraction time is 22min, is extracted 3 times.
6. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that the condition extracted in step S14 is：Extraction kettle pressure is 22MPa, and extraction temperature is 53 DEG C, and extraction time is 26min.
7. it is used to improve the culture medium of reticulate millettia tissue-cultured seedling flavone compound and polysaccharide, its feature according to claim 1
It is that temperature described in step S4 is 136 DEG C, pressure is the 1min that sterilized under 230KPa.
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