CN106386493A - Immature-embryo culture method for red-peel peanuts - Google Patents
Immature-embryo culture method for red-peel peanuts Download PDFInfo
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- CN106386493A CN106386493A CN201610857781.5A CN201610857781A CN106386493A CN 106386493 A CN106386493 A CN 106386493A CN 201610857781 A CN201610857781 A CN 201610857781A CN 106386493 A CN106386493 A CN 106386493A
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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
- A01H4/001—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
- A01H4/008—Methods for regeneration to complete plants
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Abstract
The invention belongs to the field of plant-tissue culture methods, and particularly relates to an immature-embryo culture method for red-peel peanuts. The immature-embryo culture method based on the existing peanut tissue culture comprises the following steps: disinfection and soaking of seeds, induced culture of clustered shoots, culture of strong tissue culture seedlings, rooting culture of seedlings, and transplantation after seedling hardening of complete plants. The immature-embryo culture method has the advantages that complete peanut plants can be obtained within 15 weeks by carrying out tissue culture on peanuts, so the period for tissue culture is greatly shortened, mass propagation in short time can be achieved, and the significance for propagation of rare varieties and the current peanut transgenosis is very important; secondly, culture conditions can be artificially controlled, and are not limited by regions and seasons, so that convenience is brought for stable culture production, and foundation is laid for development of tissue culture and genetic transformation of peanuts.
Description
Technical field
The invention belongs to method for plant tissue culture field is and in particular to a kind of method of immature embryo culture of red skin peanuts.
Background technology
Peanut is industrial crops important in the world and oil crops.China's peanut yield ranks first in the world, cultivated area
Occupy the second in the world, very important status is had on world's peanut production.Traditional breeding method can effectively improve peanut and produce
The characteristic of the aspects such as amount, drought resisting, but genetic engineering breeding more can be effectively to some beneficial purpose proterties than traditional breeding way
Carry out retaining, screen.The hereditary basis of peanut is weaker, and available efficient resource is limited, carries out peanut using technique for gene engineering
Genetic transformation, it is critical only that and set up an efficient receptor system, that is, need to set up efficient peanut plant tissue cultures
Method.
Peanut is as a kind of big grain legume, the existing report successfully realizing peanut Regeneration in Vitro both at home and abroad：
McKently etc., to carry scutellum and to scutellum for explant induction Multiple Buds, finds that band scutellum is the most suitable explant,
Grow up to whole plant through 258d, but in the studies above, explant process of drawing materials is complicated and larger to seed demand；
Charleen and Venkatachala, respectively with cotyledon Leaflet as explant, successfully induces embryo by somatic embryo Regeneration Ways
Callus, but there is a problem of breaking up strain difficulty；Mroginski etc. is with the immature leaflet of 3-5d seedling age as explant
Induction Multiple Buds simultaneously obtain whole plant；Tiwari etc. is with immature embryo leaflet for explant research pre-incubation time, culture medium
And the impact that genotype regenerates to cotyledon Leaflet, whole process regeneration rate up to 80%, take 4 months, but above-mentioned be outer with cotyledon Leaflet
The regenerating system of implant still has the problem that strain is difficult and repeatability is low, and regeneration frequency is low and cycle length simultaneously
Problem is also to be solved, and research simultaneously shows that plant Regeneration in Vitro has very strong dependence to genotype.
Peanut fruit contains protein, fat, carbohydrate, vitamin A, vitamin B6, vitamin E, vitamin K, Yi Jikuang
The nutrient contents such as material calcium, phosphorus, iron, the amino acid containing 8 kinds of needed by human body and unrighted acid, containing lecithin, choline, recklessly
The materials such as radish element, crude fibre.[4-5] fat content is 44%-45%, and protein content is 24-36%, and sugar content is 20% about.
Containing abundant vitamin B2, PP, A, D, E, calcium and iron etc..And contain the multivitamin such as thiamine, riboflavin, niacin,
There is anti-aging, blood coagulation hemostasis, promote to develop, strengthen memory, the effect of prevention of tumor.
Carry out peanut using method for tissue culture and expand numerous having the advantage that：First, can short time amount reproduction, to breeding
Rare Kinds and current peanut transgenic work tool are of great significance；Secondly, take up room little, be not subject to area,
Season limit；In addition, condition of culture is artificially controlled, condition is homogeneous, is easy to stably carry out culture production.
Content of the invention
It is an object of the invention to：For above-mentioned deficiency, the invention provides the cultural method of red skin peanuts, lead to
The cultural method crossing the present invention carries out tissue cultures to peanut, can obtain complete peanut plant, substantially reduce in 15 weeks
Cycle needed for tissue culture, can short time amount reproduction, to breed Rare Kinds and current peanut transgenic work have very
Important meaning；Secondly, condition of culture is artificially controlled, is not subject to area, season limit, is easy to stably carry out culture production, is
The development of peanut tissue cultures and genetic transformation is laid a good foundation.
To achieve these goals, the technical solution used in the present invention is as follows：
A kind of method of immature embryo culture of red skin peanuts, its technical scheme comprises the following steps：
（1）First surface sterilization is carried out to peanut seed, then soak seed, obtain aseptic peanut cotyledon Leaflet；
（2）Take step（1）The aseptic peanut cotyledon Leaflet obtaining, is inoculated on inducing clumping bud culture medium, cultivates 20-25d, induction
Cotyledon Leaflet produces Multiple Buds or Multiple Buds tissue block, obtains plantlet in vitro；
（3）Cut step（2）The plantlet in vitro obtaining, is inoculated into culture 15-20d on strong seedling culture base A, and 7-10d changes and once trains
Foster base, the long plantlet in vitro to more than 1.5cm is divided into individual plant, moves to culture 20-25d on strong seedling culture base B, and 8-11d changes one
Subculture, obtains seedling；
（4）By step（3）The long seedling to 2-3cm length, transfers to culture 20-25d on root media, obtains complete plant
Strain；
（5）By step（4）Described complete plant is transplanted after hardening.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（1）Described surface sterilization refers to：
Peanut maturation pod is removed pericarp, chooses full grains, the seed of smooth surface, blot surface moisture after being rinsed with water, successively
Be dipped in 1-3min in the ethanol solution that percent by volume is 75%, mass percent be 0.1% mercuric chloride solution in 9-13min,
Use rinsed with sterile water 4-8 time again；Described seed soaking is that the peanut seed through surface sterilization is peelled off kind of a skin, is soaked in 6- in sterilized water
10h.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（2）Described inducing clumping bud culture
Base is MS+0.1-0.3mg/L NAA+5-7mg/L6-BA, the solid medium of pH=5.8.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（3）Strong seedling culture base A is MS+
The solid medium of 0.1-0.3mg/LNAA+1-4mg/L6-BA, pH=5.8；Described strong seedling culture base B is MS+0.1-0.3mg/
The solid medium of LNAA+3-6mg/L6-BA+1-3mg/LGA3, pH=5.8.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（4）Described root media is 1/
The solid medium of 2MS+2-4mg/LIBA+0.1-0.5mg/LNAA, pH=5.8, its thickness is 2-2.5cm.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（4）Described complete plant is length
Go out at least 4 roots and main root length is more than 2cm.
Further, the method for immature embryo culture of described a kind of red skin peanuts, described Fiber differentiation, strong seedling culture, takes root
The temperature of culture is 25 ± 1 DEG C, and light application time is 16h/d, and intensity of illumination is 800-1200Lx.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（3）If plantlet in vitro is in strong seedling culture
Also do not grow to 1.5cm after culture 20d on base A, then excise plantlet in vitro bottom and contact the dark brown color part being formed, bottom with culture medium
Stem end submerge culture medium 2-3mm culture, change strong seedling culture base A, be cultivated for 5-7d.
Further, the method for immature embryo culture of described a kind of red skin peanuts, in step（4）Culture of rootage to after 25 days,
If situation of taking root still does not reach requirement, need in the former culture epibasal tier thickness of falling 0.6-1cm root media again, newly adding culture medium needs
With cool to 34-36 DEG C of water-bath, tissue culture bottle is tilted slowly to pour into along bottle wall.
MS in inducing clumping bud culture medium described above, strong seedling culture base, root media is international formula, 6-
BA is the basic element of cell division, IBA and NAA is auxin, and GA3 is gibberellic acid.
In sum, the present invention, due to employing such scheme, has the advantages that：Culture side by the present invention
Method carries out tissue cultures to peanut, can obtain complete peanut plant, substantially reduce the cycle needed for tissue culture in 15 weeks, can be short
Time amount reproduction, to breed Rare Kinds and current peanut transgenic work tool be of great significance；Secondly, training
Foster condition is artificially controlled, is not subject to area, season limit, is easy to stably carry out culture production, is peanut tissue cultures and heredity
The development of conversion is laid a good foundation, and social benefit and scientific research value are notable.
Specific embodiment
With reference to the method for immature embryo culture of embodiment red skin peanuts a kind of to the present invention, it is described further.The present invention
The various raw materials being used are conventional commercial product, all can be commercially available by market.
Embodiment 1：
A kind of method of immature embryo culture of red skin peanuts, its technical scheme comprises the following steps：
（1）Peanut maturation pod is removed pericarp, chooses full grains, the seed of smooth surface, after being rinsed with water, blot surface water
Point, it is dipped in 1min in the ethanol solution that percent by volume is 75% successively, mass percent is in 0.1% mercuric chloride solution
9min, then with rinsed with sterile water 4 times, then peanut seed is peelled off kind of a skin, it is soaked in 6h in sterilized water, obtain aseptic peanut embryo little
Leaf；
（2）Take step（1）The aseptic peanut cotyledon Leaflet obtaining, is inoculated on inducing clumping bud culture medium, described inducing clumping bud
Culture medium is MS+0.1mg/L NAA+5mg/L6-BA, the solid medium of pH=5.8, cultivates 20d, and induction cotyledon Leaflet produces clump
Sprout or Multiple Buds tissue block, obtain plantlet in vitro；
（3）Cut step（2）The plantlet in vitro obtaining, is inoculated into culture 15-20d, described strong seedling culture base A on strong seedling culture base A
For MS+0.1mg/LNAA+1mg/L6-BA, the solid medium of pH=5.8,7d changes a subculture；To grow to more than 1.5cm
Plantlet in vitro be divided into individual plant, move on strong seedling culture base B, described strong seedling culture base B be MS+0.1mg/LNAA+3mg/L6-BA+
The solid medium of 1mg/LGA3, pH=5.8, cultivates 20-25d, and 8d changes a subculture, obtains seedling；If plantlet in vitro is strong
Also do not grow to 1.5cm after culture 20d in seedling culture medium A, then excise plantlet in vitro bottom and contact the pitchy portion being formed with culture medium
Point, bottom stem end submerge culture medium 2-3mm culture, change strong seedling culture base A, be cultivated for 5-7d；
（4）By step（3）The long seedling to 2-3cm length, transfers on root media, and described root media is 1/2MS+
The solid medium of 2mg/LIBA+0.1mg/LNAA, pH=5.8, its thickness is 2-2.5cm, cultivates 20-25d, grows at least 4
Root and main root length are more than 2cm, obtain complete plant；Culture of rootage is to after 25 days, if situation of taking root still does not reach requirement, needs
Newly to add culture medium and need to use cool to 34-36 DEG C of water-bath, by tissue culture in the former culture epibasal tier thickness of falling 0.6-1cm root media again
Bottle tilts slowly to pour into along bottle wall；
（5）By step（4）Described complete plant is transplanted after hardening.
Further, described Fiber differentiation, strong seedling culture, the temperature of culture of rootage are 25 ± 1 DEG C, and light application time is 16h/
D, intensity of illumination is 900Lx.
Through method of immature embryo culture, 200 aseptic peanut seeds can obtain 191 plants of complete peanuts in 15 weeks to red skin peanuts
Plant, 191 plants of plantlet in vitro enter takes root the stage, and rooting rate is 100%, and to moving to root media 25 days, 99% tissue culture
Seedling all bears the root of more than 3, and root length, more than 2cm, reaches transplanting and requires, transplanting survival rate reaches more than 92%.
And common cultural method, the at most available 153 plants of complete peanut plants of 200 aseptic peanut seeds, Er Qiexu
The time wanted, considerably beyond 15 weeks, enters and takes root the stage, only 43% plantlet in vitro bears the root of more than 3, and root length typically exists
0.5-1.2cm, reaches and transplants requirement, and transplanting survival rate is less than 50%.
Embodiment 2
A kind of method of immature embryo culture of red skin peanuts, its technical scheme comprises the following steps：
（1）Peanut maturation pod is removed pericarp, chooses full grains, the seed of smooth surface, after being rinsed with water, blot surface water
Point, it is dipped in 3min in the ethanol solution that percent by volume is 75% successively, mass percent is in 0.1% mercuric chloride solution
13min, then with rinsed with sterile water 8 times, then peanut seed is peelled off kind of a skin, it is soaked in 10h in sterilized water, obtain aseptic peanut embryo
Leaflet；
（2）Take step（1）The aseptic peanut cotyledon Leaflet obtaining, is inoculated on inducing clumping bud culture medium, described inducing clumping bud
Culture medium is MS+0.3mg/L NAA+7mg/L6-BA, the solid medium of pH=5.8, cultivates 25d, and induction cotyledon Leaflet produces clump
Sprout or Multiple Buds tissue block, obtain plantlet in vitro；
（3）Cut step（2）The plantlet in vitro obtaining, is inoculated into culture 15-20d, described strong seedling culture base A on strong seedling culture base A
For MS+0.3mg/LNAA+4mg/L6-BA, the solid medium of pH=5.8,10d changes a subculture；To grow to 1.5cm with
On plantlet in vitro be divided into individual plant, move on strong seedling culture base B, described strong seedling culture base B be MS+0.3mg/LNAA+6mg/L6-BA
The solid medium of+3mg/LGA3, pH=5.8, cultivates 20-25d, and 11d changes a subculture, obtains seedling；If plantlet in vitro exists
Also do not grow to 1.5cm after culture 20d on strong seedling culture base A, then excise plantlet in vitro bottom and contact the pitchy being formed with culture medium
Part, bottom stem end submerge culture medium 2-3mm culture, change strong seedling culture base A, be cultivated for 5-7d；
（4）By step（3）The long seedling to 2-3cm length, transfers on root media, and described root media is 1/2MS+
The solid medium of 4mg/LIBA+0.5mg/LNAA, pH=5.8, its thickness is 2-2.5cm, cultivates 20-25d, grows at least 4
Root and main root length are more than 2cm, obtain complete plant；Culture of rootage is to after 25 days, if situation of taking root still does not reach requirement, needs
Newly to add culture medium and need to use cool to 34-36 DEG C of water-bath, by tissue culture in the former culture epibasal tier thickness of falling 0.6-1cm root media again
Bottle tilts slowly to pour into along bottle wall；
（5）By step（4）Described complete plant is transplanted after hardening.
Further, described Fiber differentiation, strong seedling culture, the temperature of culture of rootage are 25 ± 1 DEG C, and light application time is 16h/
D, intensity of illumination is 1200Lx.
Through method of immature embryo culture, 200 aseptic peanut seeds can obtain 195 plants of complete peanuts in 15 weeks to red skin peanuts
Plant, 195 plants of plantlet in vitro enter takes root the stage, and rooting rate is 100%, and to moving to root media 20 days, 99% tissue culture
Seedling all bears the root of more than 3, and root length, more than 2cm, reaches transplanting and requires, transplanting survival rate reaches more than 93%.
And common cultural method, the at most available 153 plants of complete peanut plants of 200 aseptic peanut seeds, Er Qiexu
The time wanted, considerably beyond 15 weeks, enters and takes root the stage, only 43% plantlet in vitro bears the root of more than 3, and root length typically exists
0.5-1.2cm, reaches and transplants requirement, and transplanting survival rate is less than 50%.
Embodiment 3
A kind of method of immature embryo culture of red skin peanuts, its technical scheme comprises the following steps：
（1）Peanut maturation pod is removed pericarp, chooses full grains, the seed of smooth surface, after being rinsed with water, blot surface water
Point, it is dipped in 2min in the ethanol solution that percent by volume is 75% successively, mass percent is in 0.1% mercuric chloride solution
11min, then with rinsed with sterile water 6 times, then peanut seed is peelled off kind of a skin, it is soaked in 8h in sterilized water, obtain aseptic peanut embryo
Leaflet；
（2）Take step（1）The aseptic peanut cotyledon Leaflet obtaining, is inoculated on inducing clumping bud culture medium, described inducing clumping bud
Culture medium is MS+0.2mg/L NAA+6mg/L6-BA, the solid medium of pH=5.8, cultivates 23d, and induction cotyledon Leaflet produces clump
Sprout or Multiple Buds tissue block, obtain plantlet in vitro；
（3）Cut step（2）The plantlet in vitro obtaining, is inoculated into culture 15-20d, described strong seedling culture base A on strong seedling culture base A
For MS+0.2mg/LNAA+2.5mg/L6-BA, the solid medium of pH=5.8,9d changes a subculture；To grow to 1.5cm with
On plantlet in vitro be divided into individual plant, move on strong seedling culture base B, described strong seedling culture base B be MS+0.2mg/LNAA+4.5mg/L6-
The solid medium of BA+2mg/LGA3, pH=5.8, cultivates 20-25d, and 9d changes a subculture, obtains seedling；If plantlet in vitro
Also do not grow to 1.5cm after culture 20d on strong seedling culture base A, then excise plantlet in vitro bottom and contact the dark brown of formation with culture medium
Color part, bottom stem end submerge culture medium 2-3mm culture, change strong seedling culture base A, be cultivated for 5-7d；
（4）By step（3）The long seedling to 2-3cm length, transfers on root media, and described root media is 1/2MS+
The solid medium of 3mg/LIBA+0.3mg/LNAA, pH=5.8, its thickness is 2-2.5cm, cultivates 20-25d, grows at least 4
Root and main root length are more than 2cm, obtain complete plant；Culture of rootage is to after 25 days, if situation of taking root still does not reach requirement, needs
Newly to add culture medium and need to use cool to 34-36 DEG C of water-bath, by tissue culture in the former culture epibasal tier thickness of falling 0.6-1cm root media again
Bottle tilts slowly to pour into along bottle wall；
（5）By step（4）Described complete plant is transplanted after hardening.
Further, described Fiber differentiation, strong seedling culture, the temperature of culture of rootage are 25 ± 1 DEG C, and light application time is 16h/
D, intensity of illumination is 1000Lx.
Through method of immature embryo culture, 200 aseptic peanut seeds can obtain 192 plants of complete peanuts in 15 weeks to red skin peanuts
Plant, 192 plants of plantlet in vitro enter takes root the stage, and rooting rate is 100%, and to moving to root media 22 days, 99% tissue culture
Seedling all bears the root of more than 3, and root length, more than 2cm, reaches transplanting and requires, transplanting survival rate reaches more than 94%.
And common cultural method, the at most available 153 plants of complete peanut plants of 200 aseptic peanut seeds, Er Qiexu
The time wanted, considerably beyond 15 weeks, enters and takes root the stage, only 43% plantlet in vitro bears the root of more than 3, and root length typically exists
0.5-1.2cm, reaches and transplants requirement, and transplanting survival rate is less than 50%.
In sum, tissue cultures are carried out to peanut by the cultural method of the present invention, can obtain complete in 15 weeks
Peanut plant, substantially reduces the cycle needed for tissue culture, can short time amount reproduction, to breeding Rare Kinds and current peanut
Transgenosis work tool is of great significance；Secondly, condition of culture is artificially controlled, is not subject to area, season limit, is easy to stable
Ground carries out culture and produces, and is that the development of peanut tissue cultures and genetic transformation is laid a good foundation, and social benefit and scientific research value show
Write.
The foregoing is only the preferred embodiment of invention, not in order to limit the present invention, all spirit in the present invention
Within principle, any modification, equivalent substitution and improvement made etc., should be included within the scope of the present invention.
Claims (9)
1. a kind of method of immature embryo culture of red skin peanuts is it is characterised in that its technical scheme comprises the following steps：
（1）First surface sterilization is carried out to peanut seed, then soak seed, obtain aseptic peanut cotyledon Leaflet；
（2）Take step（1）The aseptic peanut cotyledon Leaflet obtaining, is inoculated on inducing clumping bud culture medium, cultivates 20-25d, induction
Cotyledon Leaflet produces Multiple Buds or Multiple Buds tissue block, obtains plantlet in vitro；
（3）Cut step（2）The plantlet in vitro obtaining, is inoculated into culture 15-20d on strong seedling culture base A, and 7-10d changes and once trains
Foster base, the long plantlet in vitro to more than 1.5cm is divided into individual plant, moves to culture 20-25d on strong seedling culture base B, and 8-11d changes one
Subculture, obtains seedling；
（4）By step（3）The long seedling to 2-3cm length, transfers to culture 20-25d on root media, obtains complete plant
Strain；
（5）By step（4）Described complete plant is transplanted after hardening.
2. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（1）Described
Surface sterilization refers to：Peanut maturation pod is removed pericarp, chooses full grains, the seed of smooth surface, blot after being rinsed with water
Surface moisture, be dipped in 1-3min in the ethanol solution that percent by volume is 75% successively, mass percent be 0.1% mercury chloride molten
9-13min in liquid, then with rinsed with sterile water 4-8 time；Described seed soaking is that the peanut seed through surface sterilization is peelled off kind of a skin, soaks
6-10h in sterilized water.
3. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（2）Described
Inducing clumping bud culture medium is MS+0.1-0.3mg/L NAA+5-7mg/L6-BA, the solid medium of pH=5.8.
4. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（3）Strong sprout
Culture medium A is MS+0.1-0.3mg/LNAA+1-4mg/L6-BA, the solid medium of pH=5.8；Described strong seedling culture base B is MS
The solid medium of+0.1-0.3mg/LNAA+3-6mg/L6-BA+1-3mg/LGA3, pH=5.8.
5. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（4）Described
Root media is 1/2MS+2-4mg/LIBA+0.1-0.5mg/LNAA, the solid medium of pH=5.8, and its thickness is 2-
2.5cm.
6. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（4）Described
Complete plant is to grow at least 4 roots and main root length to be more than 2cm.
7. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：Described Fiber differentiation,
Strong seedling culture, the temperature of culture of rootage are 25 ± 1 DEG C, and light application time is 16h/d, and intensity of illumination is 800-1200Lx.
8. according to claim 1 or 5 a kind of method of immature embryo culture of red skin peanuts it is characterised in that：In step（3）If
Plantlet in vitro is not also grown to 1.5cm after cultivating 20d on strong seedling culture base A, then excise plantlet in vitro bottom and contact formation with culture medium
Dark brown color part, bottom stem end submerge culture medium 2-3mm culture, change strong seedling culture base A, be cultivated for 5-7d.
9. a kind of red skin peanuts according to claim 1 method of immature embryo culture it is characterised in that：In step（4）Take root
After cultivating 25 days, if situation of taking root still does not reach requirement, need in the former culture epibasal tier thickness of falling 0.6-1cm culture of rootage again
Base, newly adds culture medium and need to use cool to 34-36 DEG C of water-bath, tissue culture bottle is tilted slowly to pour into along bottle wall.
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|---|---|---|---|---|
| CN110367124A (en) * | 2019-08-29 | 2019-10-25 | 淮北师范大学 | A method of building peanut cotylcdon regenerating system |
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| CN102577981A (en) * | 2012-03-23 | 2012-07-18 | 山东省花生研究所 | Method for strengthening and rooting tissue culture seedlings of transgenic peanuts |
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| 张月婷等: "花生胚小叶高效再生体系的建立", 《中国油料作物学报》 * |
| 马彩霞等: "一种快速、高效花生植株再生体系的建立", 《植物生理学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110367124A (en) * | 2019-08-29 | 2019-10-25 | 淮北师范大学 | A method of building peanut cotylcdon regenerating system |
| CN110367124B (en) * | 2019-08-29 | 2021-04-09 | 淮北师范大学 | Method for constructing peanut cotyledon regeneration system |
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