CN107306795A - A kind of efficient Euonymus fortunei regeneration culture medium group - Google Patents
A kind of efficient Euonymus fortunei regeneration culture medium group Download PDFInfo
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- CN107306795A CN107306795A CN201710673368.8A CN201710673368A CN107306795A CN 107306795 A CN107306795 A CN 107306795A CN 201710673368 A CN201710673368 A CN 201710673368A CN 107306795 A CN107306795 A CN 107306795A
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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 discloses a kind of efficient Euonymus fortunei regeneration culture medium group, it is made up of callus subculture medium, induced bud growth medium, regrowth subculture medium and regrowth root media.Wherein：NAA containing 0.5mg/L and 1~1.5mg/L BA in the callus subculture medium；NAA containing 0.1mg/L and 0.5mg/L BA in the induced bud growth medium；NAA containing 0.05mg/L and 1mg/L BA in the regrowth subculture medium.NAA containing 0.6mg/L in the regrowth root media.Callus subculture medium in culture medium group of the present invention can allow Euonymus fortunei callus comparatively fast to grow, culture has good differentiation, the callus of inducibility, and induced bud growth medium is up to 16.7% to Euonymus fortunei adventitious bud induction frequency, and regrowth subculture medium has significant facilitation to Euonymus fortunei ratoon growth, rear regrowth root media makes Euonymus fortunei have good rooting efficiency.
Description
Technical field
The present invention relates to plant hormone field, more particularly to a kind of Euonymus fortunei growth regulator and preparation method thereof.
Background technology
Plant hormone is the Auto-regulator produced by plant itself, and this kind of material is general to be risen in low concentration to plant growth
Adjustment effect, and site of action is generally moved on to by generating unit in plant.Generally acknowledged plant hormone has five major classes at present,
They are：Auxins, cytokinin, gibberellin class, abscisic acid and ethene.The mechanism of action of plant hormone is so
's：Hormone and the intracellular work(that certain is referred to as showing regulation metabolism after the protein of hormone receptor is combined in plant
Energy.Hormone receptor has very strong selectivity and affinity with hormone.Some acceptors are present with changing after on plasma membrane, being combined with auxin
Become proton pump vigor on plasma membrane, influence membrane permeability.Some acceptors are present with cytoplasm and nucleus, being influenceed after being combined with hormone
DNA, RNA and protein synthesis, and regulating and controlling effect is risen to the synthesis of specific enzymes.
There are various interactions between hormone：One is synergistic effect, such as GA3It is used in conjunction with strongly facilitating with auxin
Cambial cell division.To some apple varieties, only simultaneously using stenospermocarpy could being induced to be formed；Two be to promote to make
With external source GA3The synthesis of Endogenous auxin can be promoted, because the GA applied3The plain oxidizing ferment of tissue in-growth and peroxidating can be suppressed
The activity of thing enzyme, so that the decomposition of retarding of growing element.The exogenous auxin of high concentration promotes the generation of ethene；Three be with cooperation
With such as auxin can promote the formation of root restriction, and the basic element of cell division can induce the generation of bud.Carry out plant cell and tissue is trained
When supporting, must there are the auxin and the basic element of cell division that coordinate proper proportion just to show the totipotency of cell in culture medium, both
Long root sprouts again, as intact plant；Four be antagonism, and the auxin that for example plant tip is produced, which is transported downwards, can control side
The germination and growth of bud, shows apical dominance, such as lateral bud will be granted outside the basic element of cell division, can overcome the control of auxin, promotes
Sprouting of lateral bud grows.In another example GA3Alpha-amylase nucleus formation can be suppressed by ABA in induction barley grain aleurone.Conversely,
Germination inhibitors of the ABA to potato bud can be by GA3Offset.
Artificial synthesized there is physiologically active, the compound of similar plants hormone to be referred to as plant growth regulator or plant
Exogenous hormone.Their a small amount of apply can efficiently control growing for plant, increase crop yield, in agricultural and gardening
On be used widely.These plant growth regulator have following a few classes：Growth promoter, growth retardant, growth inhibitor,
Ethylene evolution agent, defoliant, drier.Forefathers are bloomed in the growth with class plant hormone to plant, as a result made a variation with induction etc.
Success is all achieved to aspect.Especially in terms of Plant Tissue Breeding, plant growth regulator is even more to be widely used.It is various to adjust
The reasonable employment of section agent is even more successfully to indicate as tissue cultures, in the fifties in last century, just has been established for GA3Effect
Mechanism.
Induction differentiation of the plant hormone to tissue is played a very important role.In Plant Tissue Breeding, people commonly use
Hormone have NAA, BA, IBA, TD etc..During explant evoked callus, the growths such as addition IAA, NAA are often required to
The hormone or growth regulator of plain class, and to be used with basic element of cell division parahormone or growth regulator proportioning.In callus
In induction differentiation adventitious bud or Multiple Buds squamous subculture, the often hormone of the cytokinin such as addition BA or growth in its culture medium
Conditioning agent, and to be used with auxins hormone or growth regulator proportioning.In regrowth rooting process, often add NAA,
The hormone or growth regulator of the auxins such as IBA are used alone.
Euonymus fortunei is the evergreen vine plant of Celastraceae, and stem is crawled, and has aerial root.Leaf is to life, and oval, limb is yellowish extremely
It is golden yellow.Cyme axillary, grey green, 4 pieces of petal, the month at florescence 5-6.The red or pale red of capsule Huang.9~October of fruiting period.
A mutation for Euonymus fortunei system winter creeper, property likes warm, moist environment, likes sunlight, also shade tolerant.Suitable raw temperature be 15 DEG C extremely
25 DEG C, grown suitable for loose, fertile, sandy loam.
Euonymus fortunei regenerative process includes callus squamous subculture, adventitious bud inducing growth, regrowth squamous subculture
And regeneration seedling rooting, but in the prior art still without preferable, the efficient regeneration culture medium group for being applied to Euonymus fortunei.
The content of the invention
It is an object of the invention to trained for above-mentioned the deficiencies in the prior art there is provided a kind of efficient Euonymus fortunei regeneration
Support base group.
The technical solution used in the present invention is：A kind of efficient Euonymus fortunei regeneration culture medium group, by callus
Subculture medium, induced bud growth medium, regrowth subculture medium and regrowth root media composition.Wherein：
NAA containing 0.5mg/L and 1~1.5mg/L BA in the callus subculture medium；
NAA containing 0.1mg/L and 0.5mg/L BA in the induced bud growth medium；
NAA containing 0.05mg/L and 1mg/L BA in the regrowth subculture medium.
NAA containing 0.6mg/L in the regrowth root media.
As the further improvement of such scheme, in the callus subculture medium NAA containing 0.5mg/L and
1mg/L BA.
As the further improvement of such scheme, in the callus subculture medium NAA containing 0.5mg/L and
1.5mg/L BA.
As the further improvement of such scheme, above-mentioned callus subculture medium, induced bud growth medium, again
Raw seedling subculture medium and regrowth root media are used as minimal medium using MS culture mediums.
The beneficial effects of the invention are as follows：Callus subculture medium in culture medium group of the present invention can allow floral leaf to help virtue
Rattan callus comparatively fast grows, and culture has good differentiation, the callus of inducibility, and induced bud growth medium is to flower
Leaf winter creeper adventitious bud induction frequency is up to 16.7%, and regrowth subculture medium have to Euonymus fortunei ratoon growth it is aobvious
The facilitation of work, rear regrowth root media makes Euonymus fortunei have good rooting efficiency.
Embodiment
The present invention is specifically described with reference to embodiment, in order to art personnel to the present invention
Understand.It is necessary that herein the present invention will be further described it is emphasized that embodiment is only intended to, it is impossible to be interpreted as to this
The limitation of invention protection domain, art person skilled in the art, the non-intrinsically safe made according to foregoing invention content to the present invention
The modifications and adaptations of property, should still fall within protection scope of the present invention.Simultaneously following mentioned raw materials are unspecified, are
Commercially available prod；The processing step or preparation method not referred in detail be processing step known to a person skilled in the art or
Preparation method.
Embodiment 1：Callus subculture medium
Using MS as minimal medium, the growth regulator of NAA and the BA proportioning of addition various concentrations, medium pH is adjusted to
5.8~7.0, sucrose 3%, agar 0.8%, BA concentration gradients be 1.0~5.0mg/L and NAA concentration gradients be 0.05~
0.5mg/L, which is prepared, sets up 16 kinds of callus subculture mediums, and it is numbered and is formulated as follows：
N11 culture mediums MS+NAA0.05mg/L+BA1.0mg/L
N12 culture mediums MS+NAA0.05mg/L+BA1.5mg/L
N13 culture mediums MS+NAA0.05mg/L+BA2.0mg/L
N14 culture mediums MS+NAA0.05mg/L+BA5.0mg/L
N21 culture mediums MS+NAA0.1mg/L+BA1.0mg/L
N22 culture mediums MS+NAA0.1mg/L+BA1.5mg/L
N23 culture mediums MS+NAA0.1mg/L+BA2.0mg/L
N24 culture mediums MS+NAA0.1mg/L+BA5.0mg/L
N31 culture mediums MS+NAA0.2mg/L+BA1.0mg/L
N32 culture mediums MS+NAA0.2mg/L+BA1.5mg/L
N33 culture mediums MS+NAA0.2mg/L+BA2.0mg/L
N34 culture mediums MS+NAA0.2mg/L+BA5.0mg/L
N41 culture mediums MS+NAA0.5mg/L+BA1.0mg/L
N42 culture mediums MS+NAA0.5mg/L+BA1.5mg/L
N43 culture mediums MS+NAA0.5mg/L+BA2.0mg/L
N44 culture mediums MS+NAA0.5mg/L+BA5.0mg/L
IIA types and II type Euonymus fortunei callus are selected, 5~10mm fritter is cut on superclean bench,
It is inoculated on above-mentioned 16 kinds of callus subculture mediums, every kind of 8 bottles of inoculation, every bottle of 3~4 callus lines are placed in culture
Cultivated in base, condition of culture is 25 DEG C or so of temperature, and illumination 12h/d, light intensity 1600Lx, training objective 20d grow every 3d to it
Gesture is observed.
To the weight of Euonymus fortunei callus before experiment, color, volume is recorded.Inoculated and cultured is carried out, is seen
Examine callus growth situation.In Euonymus fortunei callus tissue culture 20d, its upgrowth situation is observed, as a result such as
Shown in table 1 below.
The callus squamous subculture upgrowth situation of table 1
As can be seen from Table 1, when BA concentration is constant, NAA concentration is more high more is conducive to the subculture of callus to break up；When
When NAA concentration is constant, BA concentration in the range of 1.0~2.0mg/L is more high more is conducive to callus growth.When BA concentration reaches
During 5.0mg/L, callus growth slows down and browning death occurs.When NAA and BA coordinates, synergistic effect can be produced
Really, when NAA is 0.5mg/L and BA is 1.0 or callus growth is fast during 1.5mg/L, short texture is that yellow is preferably more
Injured tissue type.
Embodiment 2：Induced bud growth medium
Using MS as minimal medium, the growth regulator of NAA and the BA proportioning of addition various concentrations, medium pH is adjusted to
5.8~7.0, sucrose 3%, agar 0.8%, BA0.1~1.0mg/L and NAA0~1.0mg/L, which are prepared, sets up 25 kinds of induced bud lifes
Long culture medium, it is numbered and is formulated as follows：
B01 culture mediums MS
B02 culture mediums MS+BA0.1mg/L
B03 culture mediums MS++BA0.2mg/L
B04 culture mediums MS+BA0.5mg/L
B05 culture mediums MS+BA1mg/L
B11 culture mediums MS+NAA0.1mg/L
B12 culture mediums MS+NAA0.1mg/L+BA0.1mg/L
B13 culture mediums MS+NAA0.1mg/L+BA0.2mg/L
B14 culture mediums MS+NAA0.1mg/L+BA0.5mg/L
B15 culture mediums MS+NAA0.1mg/L+BA1mg/L
B21 culture mediums MS+NAA0.2mg/L+BA0mg/L
B22 culture mediums MS+NAA0.2mg/L+BA0.1mg/L
B23 culture mediums MS+NAA0.2mg/L+BA0.2mg/L
B24 culture mediums MS+NAA0.2mg/L+BA0.5mg/L
B25 culture mediums MS+NAA0.2mg/L+BA1mg/L
B31 culture mediums MS+NAA0.5mg/L+BA0mg/L
B32 culture mediums MS+NAA0.5mg/L+BA0.1mg/L
B33 culture mediums MS+NAA0.5mg/L+BA0.2mg/L
B34 culture mediums MS+NAA0.5mg/L+BA0.5mg/L
B35 culture mediums MS+NAA0.5mg/L+BA1mg/L
B41 culture mediums MS+NAA1mg/L+BA0mg/L
B42 culture mediums MS+NAA1mg/L+BA0.1mg/L
B43 culture mediums MS+NAA1mg/L+BA0.2mg/L
B44 culture mediums MS+NAA1mg/L+BA0.5mg/L
B45 culture mediums MS+NAA1mg/L+BA1mg/L
IIA types and II type Euonymus fortunei callus are selected, 5~10mm fritter is cut on superclean bench,
It is inoculated on above-mentioned 25 bottles of induced bud growth mediums, it is every kind of to be at least inoculated with 8 bottles, 3~4 pieces every bottle.Condition of culture is ibid.And
After inoculated and cultured 20d, adventitious bud induction frequency is counted, and its growing state is observed in 20 days, adventitious buds differentiation and life
Shown in long situation table 2 below.
Adventitious bud induction frequency %=budding callus block number/in inoculation callus block number × 100%.
The callus adventitious bud growing state of table 2
According to table 2, in B11~B15 as can be seen that when NAA concentration in 0.1mg/L, have 4 kinds of culture mediums (B12, B13,
B14, B15) adventitious bud successfully has been induced, highest when BA concentration is 0.5mg/L, is 16.7%.
In B21~B25, concentration is 0.2mg/L in NAA, also has 4 kinds of culture mediums also successfully to induce sprout, but its
Inductivity is decreased obviously, and only 7% or so.And raising NAA concentration again after, callus could not induce sprout, callus
Structural transformation is 1 type.It is understood that concentration relatively low (0.1~0.5mg/L) is more effective to callus induction.
When NAA concentration increases, the inductivity of bud also declines rapidly.It is demonstrated experimentally that Euonymus fortunei callus is to indefinite
It is more taller than BA to NAA susceptibility in the Induction Process of bud.So reduction or refinement NAA two kinds of different culture medias it
Between multiple can more favorably find can be with the culture medium of high rate.
Embodiment 3：Regrowth subculture medium
Using MS as minimal medium, the growth regulator of NAA and the BA proportioning of addition various concentrations, medium pH is adjusted to
5.8~7.0, sucrose 3%, agar 0.8%, BA (0~2.0mg/L) and NAA (0~1.0mg/L) prepare set up 8 kinds of regrowths after
For culture medium, it is numbered and is formulated as follows：
MS makees blank control
D1 culture mediums MS+BA0.5mg/L
D2 culture mediums MS+BA1.0mg/L
D3 culture mediums MS+BA1.5mg/L
D4 culture mediums MS+BA2.0mg/L
D5 culture mediums MS+BA0.1mg/L+NAA0.01mg/L
D6 culture mediums MS+BA0.1mg/L+NAA0.05mg/L
D7 culture mediums MS+BA0.1mg/L+NAA0.1mg/L
The culture medium plant division of adventitious bud will be grown on superclean bench, above-mentioned 8 kinds of regrowth subculture mediums are inoculated into
On.It is every kind of to connect 8 bottles, every bottle of 3~4 sprouts.Condition of culture is ibid.And after 20d, the upgrowth situation of adventitious bud is observed, it is again
Raw seedling growing state is as shown in table 3 below.
The growing state of the regrowth squamous subculture of table 3
As can be seen from Table 3, the growth of regrowth needs BA supply, with the BA concentration increase of addition, regrowth
Growth accelerates, after BA concentration reaches 1.5mg/L, grows slack-off.When illustrating concentration relatively low (0.5~1.0mg/L), contribute to
Ratoon growth.Too high (more than 1.0mg/L) can then hinder it to grow.NAA's and BA plays the role of resistance to seedling with credit union, and
When NAA concentration is 0.05mg/L, inhibition level is minimum, will not produce substantially.
Embodiment 4：Regrowth root media
Using MS as minimal medium, the growth regulator of the NAA proportionings of addition various concentrations, medium pH is adjusted to 5.8
~7.0, sucrose 3%, agar 0.8%, NAA0~1.2mg/L, which is prepared, sets up 5 kinds of regrowth root medias, and it is numbered and is formulated
It is as follows：
E0 culture mediums MS
E1 culture mediums MS+NAA0.6mg/L
E2 culture mediums MS+NAA0.8mg/L
E3 culture mediums MS+NAA1mg/L
E4 culture mediums MS+NAA1.2mg/L
The regrowth cutting plant division segmentation cutting that squamous subculture is obtained, changes 5 kinds of above-mentioned regrowth root medias,
Every kind of to connect 8 bottles, every bottle of 3~4 sprouts, condition of culture ibid, allows it to carry out experiment of taking root.Taking root for regrowth is observed after inoculation
After situation, 20d, the rooting rate of regrowth is counted, its situation of taking root is as shown in table 4 below.Regrowth rooting rate %=rooted seedling numbers
Amount/experiment plant division number × 100%.
The regrowth culture of rootage situation of table 4
It can be obtained by table 4, NAA plays the role of to promote Euonymus fortunei to take root well.And when NAA concentration is 0.6mg/L,
To its rooting effect most preferably.Well developed root system and many two grades of root systems, with increasing for concentration, although NAA can also promote floral leaf
Winter creeper takes root, but the root system born is substantially changed into thick and short.Two grades of root systems are also drastically reduced.So NAA pairs of low concentration
The root induction of Euonymus fortunei is advantageously.
Above-described embodiment is the preferred embodiments of the present invention, all with similar technique of the invention and the equivalence changes made,
The protection category of the present invention all should be belonged to.
Claims (4)
1. a kind of efficient Euonymus fortunei regeneration culture medium group, by callus subculture medium, induced bud growth medium,
Regrowth subculture medium and regrowth root media composition, it is characterised in that：Contain in the callus subculture medium
There are NAA and 0.5mg/ containing 0.1mg/L in 0.5mg/L NAA and 1~1.5mg/L BA, the induced bud growth medium
NAA containing 0.05mg/L and 1mg/L BA, the regrowth root media in L BA, the regrowth subculture medium
In the NAA containing 0.6mg/L.
2. a kind of efficient Euonymus fortunei regeneration culture medium group according to claim 1, it is characterised in that：The callus group
Knit the BA of the NAA containing 0.5mg/L and 1mg/L in subculture medium.
3. a kind of efficient Euonymus fortunei regeneration culture medium group according to claim 1, it is characterised in that：The callus group
Knit the BA of the NAA containing 0.5mg/L and 1.5mg/L in subculture medium.
4. a kind of efficient Euonymus fortunei regeneration culture medium group according to claim 1, it is characterised in that：The callus group
Subculture medium, induced bud growth medium, regrowth subculture medium and regrowth root media are knitted with MS culture mediums
It is used as minimal medium.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111670807A (en) * | 2020-04-27 | 2020-09-18 | 江苏农林职业技术学院 | Tissue culture method of euonymus alatus |
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| CN106797890A (en) * | 2017-03-14 | 2017-06-06 | 河南红枫种苗股份有限公司 | A kind of culture medium and its method for reducing leather leaf winged euonymus tissue culture pollution rate |
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- 2017-08-08 CN CN201710673368.8A patent/CN107306795A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106797890A (en) * | 2017-03-14 | 2017-06-06 | 河南红枫种苗股份有限公司 | A kind of culture medium and its method for reducing leather leaf winged euonymus tissue culture pollution rate |
Non-Patent Citations (1)
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| 徐萌: "扶芳藤组织培养再生体系的建立", 《中国优秀博硕士学位论文全文数据库（硕士）农业科技辑》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111670807A (en) * | 2020-04-27 | 2020-09-18 | 江苏农林职业技术学院 | Tissue culture method of euonymus alatus |
| CN111670807B (en) * | 2020-04-27 | 2022-03-11 | 江苏农林职业技术学院 | Tissue culture method of euonymus alatus |
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