CN106508675A - Seed tissue culture medium for root of subprostrate sophora - Google Patents
Seed tissue culture medium for root of subprostrate sophora Download PDFInfo
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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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Abstract
The invention provides a seed tissue culture medium for the root of subprostrate sophora, and relates to the technical field of tissue culture. The seed tissue culture medium is formed by mixing a basal culture solution and a mother solution with the volume ratio being 3-5:0.03-0.08. The mother solution mainly contains hymexazol, brassinolide, alpha-naphthylacetic sodium salt, pyrimethanil and diethyl aminoethyl hexanoate. The basal culture solution mainly contains pineapple waste hydrolysate, cane sugar, agar, lactoalbumin hydrolysate, ABT rooting powder, zeatin, kinetin, isopentenylaminopurine and gibberellin. According to the seed tissue culture medium disclosed by the invention, the using quantity of culture medium raw materials like saccharose in a traditional process is reduced through utilization of pineapple waste in the basal culture solution, and preparation cost of the culture medium is reduced. In addition, by adding the formula of the mother solution, rooting of strong seedlings can be promoted quickly, and the rooting percentage can reach 99% or above. Moreover, the average plant number reaches 20-25 after the strong seedlings take roots.
Description
【Technical field】
The present invention relates to tissue culture technology field, and in particular to a kind of root of subprostrate sophora seed tissue culture medium.
【Background technology】
Root of subprostrate sophora also known as Radix Sophorae Tonkinensiss, are the root of leguminous plant Sophora tonkinensis Gagnep..The effect of with heat-clearing and toxic substances removing, relieving sore throat and diminishing swelling, can
For treating the diseases such as pharyngalgia, gingival swelling and pain, cough due to lung-heat, jaundice due to damp-heat, nasopharyngeal carcinoma, cervical cancer, recent studiess find root of subprostrate sophora
Contained total alkali and oxymatrine have the function of promoting coronary flow to suppress tumor cell and enhance immunity.
Root of subprostrate sophora is mainly distributed on Guangxi, and also there is product in Yunnan, Guizhou, but yield is less.It is mainly derived from wild, Jing Guoduo
Year excavation, wild resource reduced year by year, some places of production oneself be on the verge of exhaustion or in exhausted state, distributional region reduces increasingly,
Landings are reduced year by year, and the development and application to root of subprostrate sophora in recent years is increasing year by year, causes serious confession should not
Ask.
Under naturalness, root of subprostrate sophora relies primarily on seed and is bred, however, often seed also immaturity is just most of
Come off, cause breeding coefficient low, in addition, it is larger by taking seed to carry out the obvious difficulty of artificial breeding, so provide one kind can
Root of subprostrate sophora seed is made for introduces a collection, shortening seedling raise period, it is ensured that after the survival rate of bud, increase bud meat thickness, guarantee are sprouted after sprouting
The glossy dark green cultural method of bud color is extremely urgent.
【The content of the invention】
In view of this, it is an object of the present invention to provide a kind of root of subprostrate sophora seed tissue culture medium, wherein, in basis training
The culture medium raw material consumption such as sucrose in traditional handicraft is reduced in nutrient solution to the utilization of pineapple bran, culture medium is saved and is prepared into
This, is in addition, the mother solution formula of the addition present invention, can rapidly promote strengthening seedling and rooting so that rooting rate reaches more than 99%, and
Plant mean elements after strengthening seedling and rooting reaches 20-25 bars.
In order to solve above-mentioned technical problem, following technical scheme is present invention employs：
A kind of root of subprostrate sophora seed tissue culture medium, the culture medium are mixed with mother solution by basic culture solution, they
Volume ratio is 3-5:0.03-0.08；
Every liter of mother solution is mainly contained：The hymexazol of 10-15mg, the brassin lactones of 10-20mg, 10-50mg α-
The diethyl aminoethyl hexanoate of Nafusaku, the pyrimethanil of 10-50mg and 20-50mg；Every liter of basic culture solution is mainly contained：30-46mg
Pineapple bran hydrolyzed solution, the sucrose of 5-10mg, the agar of 10-20mg, the lactoalbumin hydrolysate of 5-8mg, the ABT root-inducing powders of 2-8mg,
The zeatin of 1-4mg, the kinetins of 1-4mg, the iso-amylene amidopurin of 1-3mg and the gibberellins of 1-3mg.
Preferably, every liter of mother solution is mainly contained：The hymexazol of 12mg, the brassin lactones of 15mg, the α-naphthalene of 30mg
The diethyl aminoethyl hexanoate of sodium acetate, the pyrimethanil of 30mg and 35mg；Every liter of basic culture solution is mainly contained：The pineapple bran hydrolysis of 38mg
Liquid, the sucrose of 7mg, the agar of 15mg, the lactoalbumin hydrolysate of 6mg, the ABT root-inducing powders of 5mg, the zeatin of 2mg, the excitement of 2mg
Element, the iso-amylene amidopurin of 2mg and the gibberellins of 1-3mg.
Preferably, the pineapple bran hydrolyzed solution is mainly obtained through following process：A. the Fructus Ananadis comosi obtained after squeeze the juice Fructus Ananadis comosi
Slag is dried, crushes, be subsequently adding ionic liquid to pineapple bran mass concentration be 3%-8%, then 90-120 DEG C,
Under conditions of 80-120r/min, stir process 60-120min obtains pineapple bran solution, adds 4-8 times of body of pineapple bran solution
Long-pending water, after 150-200r/min stir process 30-60min, under 3000-5000r/min, is centrifuged 10-20min, collects
Precipitation, will be deposited in 40-60 DEG C of vacuum drying 3-5h, and obtain regenerating pineapple bran；B. pineapple bran will be regenerated to add in right amount obtained by step a
Water is fully mixed, and is added；Described enzymolysis is to be in pH value
4.5-6.5, under conditions of temperature is 55-65 DEG C, enzymolysis time is 10-12h.
Preferably, the ionic liquid is guanidinium ionic liquid, alcaminess ionic liquid, glyoxaline ion liquid, season
Ammonium salt class ionic liquid, thiazoless ionic liquid, polynary amine ionic liquid, triazole ionic liquid, pyrrolin class ion
One kind in liquid, Thiazoling type ionic liquid and benzotriazole ionic liquid.
Preferably, the configuration of the culture medium is to match somebody with somebody to postpone basic culture solution and mother solution according to respective component, according to
Volume ratio is 3-5:0.03-0.08 mixes, then boils regulation pH, after concentrating laggard horizontal high voltage sterilizing.
Preferably, the pH=5.0-6.0 of the culture medium.
In sum, as a result of above-mentioned technical proposal, the invention has the beneficial effects as follows：
1st, a kind of root of subprostrate sophora seed tissue culture medium of the invention is improved on the formula of traditional culture medium, is added
Can be in the factor needed for root of subprostrate sophora is during strong plantlets and rootage, it has been investigated that using addition after the basic culture solution of the present invention originally
The mother solution formula of invention, using the teaching of the invention it is possible to provide the good condition taken root, promotion Seedling health are healthy and strong, and wherein diethyl aminoethyl hexanoate can promote root system
Development, adjusts the balance of internal nutrient；Brassin lactones are growth regulator, can growth regulation, hestening rooting；In tissue culture process
In often occur aetiolation, general two reasons, cytoactive is not enough, and dead and courses of infection is dead, hymexazol and phonetic mould
The addition of amine can effectively suppress the mycelial normal growth of pathogenic fungi or directly kill pathogenic bacteria, and plant can be promoted again to give birth to
It is long；The mother solution formula of the present invention can rapidly promote strengthening seedling and rooting, and rooting rate reaches the plant after more than 99%, and strengthening seedling and rooting
Mean elements reaches 20-25 bars.
2nd, in root of subprostrate sophora seed tissue culture medium of the invention, basic culture solution is added with pineapple bran, using ionic liquid
Regeneration Treatment is carried out to which, is then hydrolyzed with cellulase, in pineapple bran, mainly contain the compositions such as cellulose, reducing sugar,
Cellulose solution after dissolving separates out can Cellulose precipitates after adding water, and realize the regeneration of cellulose, and regenerative process can remove
Lignin component, reduces the inhibitory action to subsequent enzymatic hydrolysis process, while regenerative process can make part hydrogen in cellulosic molecule
Bond fission, the degree of polymerization, degree of crystallinity are reduced, and make the configuration of surface of cellulose change, surface area increase, and then the contact with enzyme
Area increases, and the utilization to pineapple bran reduces the culture medium raw material consumption such as sucrose in traditional handicraft, saves culture medium preparation
Cost, while the problem of environmental pollution that the pineapple bran produced during can solve the problem that juice production causes.
3rd, culture medium preparation process is simple of the invention, easy to operate, with great dissemination.
【Specific embodiment】
The following examples can help those skilled in the art that the present invention is more fully understood, but cannot be with any
Mode limits the present invention.
Embodiment 1
A kind of root of subprostrate sophora seed tissue culture medium, is 3 by volume ratio:0.03 basic culture solution is mixed with mother solution,
Wherein, every liter of mother solution is mainly contained：The hymexazol of 10mg, the brassin lactones of 10mg, the α-naphthaleneacidsodium of 10mg, 10mg it is phonetic
The diethyl aminoethyl hexanoate of mould amine and 20mg；Every liter of basic culture solution is mainly contained：The pineapple bran hydrolyzed solution of 30mg, the sucrose of 5mg, 10mg
Agar, the lactoalbumin hydrolysate of 5mg, the ABT root-inducing powders of 2mg, the zeatin of 1mg, the kinetins of 1mg, 1mg isoamyl enamino it is fast
The gibberellins of purine and 1mg；
The configuration of culture medium is to match somebody with somebody to postpone basic culture solution and mother solution according to respective component, is 3 according to volume ratio:
0.03 mixing, then regulation pH is boiled, after concentrating laggard horizontal high voltage sterilizing, the PH=5.0 of culture medium.
Pineapple bran hydrolyzed solution in basic culture solution is mainly obtained through following process：A. pineapple bran is dried, powder
Broken, the mass concentration for being subsequently adding ionic liquid to pineapple bran is 3%, then under conditions of 90 DEG C, 80r/min, at stirring
Reason 60min, obtains pineapple bran solution, adds the water of 4 times of volumes of pineapple bran solution, after 150r/min stir process 30min,
Under 3000r/min, 10min is centrifuged, collects precipitation, 40 DEG C of vacuum drying 3h will be deposited in, obtain regenerating pineapple bran；B. will step
Regenerating pineapple bran obtained by rapid a adds suitable quantity of water fully to mix, and adds
Liquid；Enzymolysis be pH value be 4.5, temperature be 55 DEG C under conditions of, enzymolysis time is 10h；
Ionic liquid is guanidinium ionic liquid.
Embodiment 2
A kind of root of subprostrate sophora seed tissue culture medium, is 4 by volume ratio:0.05 basic culture solution is mixed with mother solution,
Wherein, every liter of mother solution is mainly contained：The hymexazol of 12mg, the brassin lactones of 15mg, the α-naphthaleneacidsodium of 30mg, 35mg it is phonetic
The diethyl aminoethyl hexanoate of mould amine and 30mg；Every liter of basic culture solution is mainly contained：The pineapple bran hydrolyzed solution of 38mg, the sucrose of 7mg, 15mg
Agar, the lactoalbumin hydrolysate of 7mg, the ABT root-inducing powders of 5mg, the zeatin of 2mg, the kinetins of 2mg, 2mg isoamyl enamino it is fast
The gibberellins of purine and 2mg；
The configuration of culture medium is to match somebody with somebody to postpone basic culture solution and mother solution according to respective component, is 4 according to volume ratio:
0.05 mixing, then regulation pH is boiled, after concentrating laggard horizontal high voltage sterilizing, the PH=5.5 of culture medium.
Pineapple bran hydrolyzed solution in basic culture solution is mainly obtained through following process：A. pineapple bran is dried, powder
Broken, the mass concentration for being subsequently adding ionic liquid to pineapple bran is 3%-8%, then under conditions of 105 DEG C, 100r/min,
Stir process 80min, obtains pineapple bran solution, adds the water of pineapple bran 4-8 times of volume of solution, in 180r/min stir process
After 45min, under 4000r/min, 15min is centrifuged, collects precipitation, 50 DEG C of vacuum drying 4h will be deposited in, obtain regenerating Fructus Ananadis comosi
Slag；B. pineapple bran will be regenerated obtained by step a adds suitable quantity of water fully to mix, and adds
Trailing plants slag hydrolyzed solution；Hydrolysis be pH value be 5.5, temperature be 60 DEG C under conditions of, enzymolysis time is 11h；
Ionic liquid is glyoxaline ion liquid.
Embodiment 3
A kind of root of subprostrate sophora seed tissue culture medium, is 5 by volume ratio:0.08 basic culture solution is mixed with mother solution,
Wherein, every liter of mother solution is mainly contained：The hymexazol of 15mg, the brassin lactones of 20mg, the α-naphthaleneacidsodium of 50mg, 50mg it is phonetic
The diethyl aminoethyl hexanoate of mould amine and 50mg；Every liter of basic culture solution is mainly contained：The pineapple bran hydrolyzed solution of 46mg, the sucrose of 10mg, 20mg
Agar, the lactoalbumin hydrolysate of 8mg, the ABT root-inducing powders of 8mg, the zeatin of 4mg, the kinetins of 4mg, the isoamyl enamino of 3mg
The gibberellins of purine and 3mg；
The configuration of culture medium is to match somebody with somebody to postpone basic culture solution and mother solution according to respective component, is 5 according to volume ratio:
0.08 mixing, then regulation pH is boiled, after concentrating laggard horizontal high voltage sterilizing, the PH=6.0 of culture medium.
Pineapple bran hydrolyzed solution in basic culture solution is mainly obtained through following process：A. pineapple bran is dried, powder
Broken, the mass concentration for being subsequently adding ionic liquid to pineapple bran is 8%, then under conditions of 120 DEG C, 120r/min, stirring
120min is processed, pineapple bran solution is obtained, is added the water of pineapple bran 4-8 times of volume of solution, in 200r/min stir process
After 60min, under 5000r/min, 20min is centrifuged, collects precipitation, 60 DEG C of vacuum drying 3-5h will be deposited in, obtain regenerating spinach
Trailing plants slag；B. pineapple bran will be regenerated obtained by step a adds suitable quantity of water fully to mix, and adds after cellulase is digested and is obtained
Pineapple bran hydrolyzed solution；Enzymolysis be pH value be 6.5, temperature be 65 DEG C under conditions of, enzymolysis time is 12h；
Ionic liquid is pyrrolin class ionic liquid.
Compliance test result
In order to further illustrate the using value of root of subprostrate sophora seed tissue culture medium of the present invention, implement people and select disease-free,
Without incompleteness, the close root of subprostrate sophora seed of size, after sterilizing, is prepared in being aseptically respectively connected to following 2 groups
The culture medium for obtaining, each processes and is inoculated with 10 bottles, 10 per bottle, cultivation temperature (27 ± 1) DEG C, intensity of illumination 2500-3000lx,
Light application time 12h/d.Record sprouts the time, and bud growing state the results are shown in Table 1；
First group：Remove mother solution, other modes are carried out in strict accordance with embodiment 2；
Second group：Carry out in strict accordance with embodiment 2.
The growing state of 1 root of subprostrate sophora seed of table
| Test group | Germination percentage % | Sprout the time (h) | Bud color | Bud meat thickness (cm) |
| First group | 75 | 70 | It is light yellow-yellowish green-yellowish green | 0.09 |
| Second group | 100 | 41 | Light yellow-light green-green | 0.13 |
As seen from the above table, by adopt the present invention technical scheme (embodiment 2), germination percentage reach 100%, averagely
Time and bud color sprout also superior to removing first group of mother solution, and bud thickness is full.
Although above the present invention is described in detail with a general description of the specific embodiments,
On the basis of the present invention, it can be made some modifications or improvements, this will be apparent to those skilled in the art.Cause
This, these modifications or improvements, belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Claims (6)
1. a kind of root of subprostrate sophora seed tissue culture medium, it is characterised in that the culture medium mixed with mother solution by basic culture solution and
Into their volume ratio is 3-5:0.03-0.08；
Every liter of mother solution is mainly contained：The hymexazol of 10-15mg, the brassin lactones of 10-20mg, the α-naphthalene second of 10-50mg
The diethyl aminoethyl hexanoate of sour sodium, the pyrimethanil of 10-50mg and 20-50mg；Every liter of basic culture solution is mainly contained：The spinach of 30-46mg
Trailing plants slag hydrolyzed solution, the sucrose of 5-10mg, the agar of 10-20mg, the lactoalbumin hydrolysate of 5-8mg, the ABT root-inducing powders of 2-8mg, 1-
The zeatin of 4mg, the kinetins of 1-4mg, the iso-amylene amidopurin of 1-3mg and the gibberellins of 1-3mg.
2. a kind of root of subprostrate sophora seed tissue culture medium according to claim 1, it is characterised in that every liter of mother solution is main
Contain：The amine of the hymexazol of 12mg, the brassin lactones of 15mg, the α-naphthaleneacidsodium of 30mg, the pyrimethanil of 30mg and 35mg is fresh
Ester；Every liter of basic culture solution is mainly contained：The pineapple bran hydrolyzed solution of 38mg, the sucrose of 7mg, the agar of 15mg, the water of 6mg
Solution lactoprotein, the ABT root-inducing powders of 5mg, the zeatin of 2mg, the kinetins of 2mg, the iso-amylene amidopurin of 2mg and 1-3mg's
Gibberellins.
3. a kind of root of subprostrate sophora seed tissue culture medium according to claim 1, it is characterised in that the pineapple bran hydrolyzed solution
It is main to obtain through following process：A. pineapple bran is dried, crushed, the quality for being subsequently adding ionic liquid to pineapple bran is dense
Spend for 3%-8%, then under conditions of 90-120 DEG C, 80-120r/min, stir process 60-120min obtains pineapple bran molten
Liquid, adds the water of pineapple bran 4-8 times of volume of solution, after 150-200r/min stir process 30-60min, in 3000-
Under 5000r/min, 10-20min is centrifuged, collects precipitation, 40-60 DEG C of vacuum drying 3-5h will be deposited in, obtain regenerating pineapple bran；
B. pineapple bran will be regenerated obtained by step a adds suitable quantity of water fully to mix, and adds
Slag hydrolyzed solution；
Described enzymolysis be pH value be 4.5-6.5, temperature be 55-65 DEG C under conditions of, enzymolysis time is 10-12h.
4. a kind of root of subprostrate sophora seed tissue culture medium according to claim 1, it is characterised in that the configuration of the culture medium
It is to match somebody with somebody to postpone basic culture solution and mother solution according to respective component, is 3-5 according to volume ratio:0.03-0.08 mixes, then boils
PH is adjusted, after concentrating laggard horizontal high voltage sterilizing.
5. a kind of root of subprostrate sophora seed tissue culture medium according to claim 1, it is characterised in that the pH=of the culture medium
5.0-6.0。
6. a kind of root of subprostrate sophora seed tissue culture medium according to claim 3, it is characterised in that the ionic liquid is guanidine
It is salt ion liquid, alcaminess ionic liquid, glyoxaline ion liquid, ion liquid of quaternaries, thiazoless ionic liquid, many
First amine ionic liquid, triazole ionic liquid, pyrrolin class ionic liquid, Thiazoling type ionic liquid and BTA
One kind in class ionic liquid.
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| CN108456098A (en) * | 2018-01-26 | 2018-08-28 | 宜州市横山泰顺种养专业合作社 | A kind of red heart honey shaddock fertilizer and preparation method thereof |
| CN108293877A (en) * | 2018-02-11 | 2018-07-20 | 平南县德湖种养农民专业合作社 | A kind of Monstera deliciosa seed tissue culture medium |
| CN111357568A (en) * | 2020-04-28 | 2020-07-03 | 广西壮族自治区农业科学院 | Edible fungus cultivation base material and preparation method thereof |
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