CN104823853B - A kind of method of dendrobe tissue culture bud fast breeding - Google Patents
A kind of method of dendrobe tissue culture bud fast breeding Download PDFInfo
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- CN104823853B CN104823853B CN201510234922.3A CN201510234922A CN104823853B CN 104823853 B CN104823853 B CN 104823853B CN 201510234922 A CN201510234922 A CN 201510234922A CN 104823853 B CN104823853 B CN 104823853B
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Abstract
The invention discloses a kind of method of dendrobe tissue culture bud fast breeding, the method carries out short-term immersion to dendrobe tissue culture bud using anion water to stimulate, then the light culture Nature enemy in sugar-free culture-medium, finally it is transferred in eutrophy culture medium, recover illumination condition, promote dendrobe tissue culture bud fast breeding, specifically include following operating procedure：（1）Anion water dipping pretreatment；（2）The hungry dark treatment of dendrobe tissue culture bud；（3）The Multiplying culture of dendrobe tissue culture bud.The present invention promotes dendrobe tissue culture bud fast breeding by short-term anion water combination light culture starvation Stress treatment, later stage eutrophy culture.The method is simple and easy to apply, with low cost, and dendrobe tissue culture bud propagation efficiency high, it is adaptable to which batch production is mass produced.
Description
Technical field
Patent of the present invention is related to plant tissue culture technical field, more particularly to a kind of method of dendrobe tissue culture bud fast breeding.
Background technology
Dendrobe tissue culture bud is intermediary of the stem of noble dendrobium by embryo seedling differentiation, is also to cultivate the crucial ring for forming test tube seedling
Section.Under common condition of culture, dendrobe tissue culture bud Proliferation, Differentiation speed is slow, and propagation efficiency is low.Addition hormone is commonly used at present
Means promote the Proliferation, Differentiation of tissue culture bud, but the addition of exogenous hormone, plant can be enriched to the carrying out of the growth of plant
In thing body, there is hidden danger in the security to eating.Seek the side for safely and effectively promoting dendrobe tissue culture bud fast breeding to break up
The problems such as method is to solving stem of noble dendrobium propagation during existing plant layout metaplasia is produced slow with differentiation, hormone residues is significant.
The content of the invention
Growing multiplication the invention aims to solve to be run into existing dendrobe tissue culture bud propagation production process is slow,
Exogenous hormone use, the problems such as production life cycle is long and a kind of method of dendrobe tissue culture bud fast breeding that provides, it is simple to operate, into
This is less expensive.
Realize that the technical solution of above-mentioned purpose is as follows：
A kind of method of dendrobe tissue culture bud fast breeding, it is characterised in that carry out as steps described below：
（1）Anion water dipping pretreatment：Dendrobe tissue culture bud is taken out from blake bottle, anion nutrient solution is added into extremely
Submergence, under the conditions of 25 DEG C, soaks 4-8 hours.
（2）The hungry dark treatment of dendrobe tissue culture bud：Dendrobe tissue culture bud after immersion is taken out, sugar culture-medium is transferred to
In, under the conditions of 25 DEG C, light culture 48-96 hours.
（3）The Multiplying culture of dendrobe tissue culture bud:From going to take out dendrobe tissue culture bud in sugar culture-medium, transfer into Multiplying culture
In base, in 25 ± 2 DEG C, 50 μm ol/ (m of temperature2S), under conditions of daily illumination 14 hours, intensity of illumination 1500-2000LX
Culture 30-40 days, dendrobe tissue culture bud can obtain fast breeding in this time.
The described stem of noble dendrobium is the one kind in Dendrobidium huoshanness, dendrobium candidum, Dendrobium Moniliforme.
Described dendrobe tissue culture bud is sprouted for Dendrobidium huoshanness, dendrobium candidum or Dendrobium Moniliforme seed in basic MS culture medium
Budlet.
Step（1）Described in anion water in anion concentration be not less than 200/cm3。
Step（2）Described in go sugar culture-medium be remove sucrose liquid MS medium.
Step（3）In proliferated culture medium, be sucrose containing 30g/L：Maltose ratio is 1:0.5~1:1.5 disaccharide is consolidated
Body MS culture mediums.
Principle summary：
Under anion refers to external condition, atoms outermost electronics break away from atomic nucleus, are discharged from atom, and escape
Free electron can be attached on some molecules or atom, form new negatively charged molecule.By ionizing what part water was obtained
Anion can increase ion concentration, promote absorption of the plant to moisture and nutriment, strengthen the matter transportation function of plant,
Adjust the equilibrium osmotic pressure of plant.Partial oxidation can be discharged in anion forming process, strengthens the absorption of root, so that
Reach the effect for promoting plant metabolism.
Short-term Hunger stress will not produce damage to body, and plant can be promoted compensatory growth phenomenon occur on the contrary.Work as the side of body
After compeling to release, the photosynthetic efficiency of plant and Nutrient Absorption ability is increased, growth rate can be carried significantly
It is high.
Compared with prior art, the method for the present invention has the advantages that：The present invention is by short-term anion
Reason generation is sprouted with reference to light culture stress to promote the dendrobe tissue culture bud to strengthen its metabolic vigor, improves the speed to photosynthetic response
Rate, reaches the purpose of synchronization, fast-growth and propagation.
1st, the present invention is sprouted using short-term anion and instead of tradition with reference to light culture stress and eutrophy cultural method
Dendrobe tissue culture bud is promoted to breed using hormone, it is to avoid exogenous hormone uses the harm to health.
2nd, the present invention is sprouted with reference to light culture stress and eutrophy cultural method using short-term anion, is significantly improved
Proliferation rate, dendrobe tissue culture bud fresh weight proliferation rate is up to more than 400% within 30-40 days, and the proliferation rate of bud, up to more than 110%, is approximately not locate
4 times of reason control group.Proliferation rate=（After treatment-before processing）/ before processing × 100%.
3rd, the present invention is sprouted with reference to light culture stress and eutrophy cultural method using short-term anion, simple to operate, into
This is less expensive, for enterprise's batch production, the large-scale production stem of noble dendrobium provide new method.
Specific embodiment
The present invention is further described with reference to specific embodiment.The features and advantages of the invention will be with
Describe and become apparent from, but these exemplary implementation methods are used merely to the explanation present invention, the scope of the present invention are not constituted
Any limitation.
Involved Dendrobidium huoshanness tissue culture bud, candidum tissue culturing bud, Dendrobium Moniliforme tissue culture bud are An Huikang in this patent
Er Xin Biology Pharmacy Co., Ltd produces；MS culture medium medicines are analyzes pure, from the bright bio tech ltd's purchase of Pood
Buy；Anion water dispenser is bought in Ge Hao electrolysis waters Co., Ltd, and anion concentration is not less than 200/cm3。
（1）Anion water dipping pretreatment：Dendrobe tissue culture bud is taken out from blake bottle, anion nutrient solution is added into extremely
Submergence, under the conditions of 25 DEG C, soaks 4-8 hours.
（2）The hungry dark treatment of dendrobe tissue culture bud：Dendrobe tissue culture bud after immersion is taken out, sugar culture-medium is transferred to
In, under the conditions of 25 DEG C, light culture 48-96 hours.
（3）The Multiplying culture of dendrobe tissue culture bud:From the dendrobe tissue culture bud that light culture takes out, in proliferated culture medium of transferring,
In 25 ± 2 DEG C, 50 μm ol/ (m of temperature2S), cultivated under conditions of daily illumination 14 hours, intensity of illumination 1500-2000LX
30-40 days, dendrobe tissue culture bud can obtain fast breeding in this time.Above is referred to tissue culture bud switch over operation all should be ultra-clean
Carried out in workbench.
Embodiment 1
A kind of method of Dendrobidium huoshanness tissue culture bud fast breeding, including following operating procedure：
Step 1：Anion water dipping pretreatment
Dendrobidium huoshanness tissue culture bud 20g, average 32/g of bud number are taken out from blake bottle, anion concentration is put into and is not less than
200/cm3The mL of anion water 200 in submerge, under the conditions of 25 DEG C, immersion treatment 4 hours.
Step 2：The hungry dark treatment of Dendrobidium huoshanness tissue culture bud
Dendrobidium huoshanness tissue culture bud after immersion is taken out, is transferred in desaccharification MS culture mediums, every bottle of switching about 4g, in temperature
At 25 ± 2 DEG C, light culture 96 hours.
Step 3：The Multiplying culture of Dendrobidium huoshanness tissue culture bud
From the Dendrobidium huoshanness tissue culture bud that light culture takes out, transfer into containing 30g/L（Sucrose：Maltose ratio is 1:0.5）'s
In disaccharide MS solid multiplication culture mediums, every bottle of about 4 g of switching, in 25 ± 2 DEG C, 50 μm ol/ (m of temperature2S), daily illumination 14
Hour, cultivate 35 days under conditions of intensity of illumination 1500LX.The Dendrobidium huoshanness tissue culture bud of propagation, every bottle of weight are taken out after 35 days
The g of average out to 21.47, rate of body weight gain is 436.75%, and the number that sprouts is average up to 77/g, and bud ratio is 140%.
Embodiment 2
A kind of method of candidum tissue culturing bud fast breeding, including following operating procedure：
Step 1：Anion water dipping pretreatment
Candidum tissue culturing bud 20g, average 41/g of bud number are taken out from blake bottle, anion concentration is put into and is not less than
200/cm3The mL of anion water 200 in submerge, under the conditions of 25 DEG C, the h of immersion treatment 6.
Step 2：The hungry dark treatment of candidum tissue culturing bud
Candidum tissue culturing bud after immersion is taken out, is transferred in the liquid MS medium of removal sucrose, every bottle of switching
About 4g, at 25 ± 2 DEG C of temperature, the h of light culture 72.
Step 3：The Multiplying culture of candidum tissue culturing bud
From the candidum tissue culturing bud that light culture takes out, transfer into containing 30 g/L（Sucrose：Maltose ratio is 1:1）It is double
In sugared MS solid multiplications culture medium, every bottle of switching about 4g, in 25 ± 2 DEG C, 50 μm ol/ (m of temperature2S), daily illumination 14 is small
When, cultivate 30 days under conditions of intensity of illumination 1800LX.The candidum tissue culturing bud of propagation, every bottle of weight average are taken out after 30 days
It is 22.12 g, rate of body weight gain is 453.00%, bud number is average up to 87/g, and bud ratio is 112.20%.
Embodiment 3
A kind of method of Dendrobium Moniliforme tissue culture bud fast breeding, including following operating procedure：
Step 1：Anion water dipping pretreatment
Dendrobium Moniliforme tissue culture bud 20g, average 38/g of bud number are taken out from blake bottle, anion concentration is put into and is not less than
200/cm3The mL of anion water 200 in submerge, under the conditions of 25 DEG C, the h of immersion treatment 4.
Step 2：The hungry dark treatment of Dendrobium Moniliforme tissue culture bud
Dendrobium Moniliforme tissue culture bud after immersion is taken out, is transferred in desaccharification MS culture mediums, every bottle of about 4 g of switching, in temperature
At 25 ± 2 DEG C of degree, the h of light culture 48.
Step 3：The Multiplying culture of Dendrobium Moniliforme tissue culture bud
From the Dendrobium Moniliforme tissue culture bud that light culture takes out, transfer into containing 30g/L（Sucrose：Maltose ratio is 1:1.5）'s
In disaccharide MS solid multiplication culture mediums, every bottle of about 4 g of switching, in 25 ± 2 DEG C, 50 μm ol/ (m of temperature2S), daily illumination 14
Cultivated 30 days under conditions of hour, intensity of illumination 2000LX.The Dendrobium Moniliforme tissue culture bud of propagation is taken out after 30 days, every bottle of weight is put down
20.37g is, rate of body weight gain is 409.25%, bud number is average up to 82/g, and bud ratio is 115.79%.
Claims (3)
1. a kind of method of dendrobe tissue culture bud fast breeding, it is characterised in that carry out as steps described below：
（1）Anion water dipping pretreatment：Dendrobe tissue culture bud is taken out from blake bottle, anion water to submergence, 25 is added into
Under the conditions of DEG C, soak 4-8 hours；
Anion concentration in described anion water is not less than 200/cm3；
（2）The hungry dark treatment of dendrobe tissue culture bud：Dendrobe tissue culture bud after immersion is taken out, is transferred in sugar culture-medium, 25
Under the conditions of DEG C, light culture 48-96 hours；
It is described to remove the liquid MS medium that sugar culture-medium is removal sucrose；
（3）The Multiplying culture of dendrobe tissue culture bud:From go in sugar culture-medium take out dendrobe tissue culture bud, in proliferated culture medium of transferring,
Cultivated 30-40 days under conditions of 25 ± 2 DEG C of temperature, daily illumination 14 hours, intensity of illumination 1500-2000LX, this time
Interior dendrobe tissue culture bud fast breeding；
The proliferated culture medium is sucrose containing 30g/L：Maltose ratio is 1:0.5~1:1.5 disaccharide solids MS culture mediums.
2. the method for a kind of dendrobe tissue culture bud fast breeding according to claim 1, it is characterised in that the described stem of noble dendrobium is
One kind in Dendrobidium huoshanness, dendrobium candidum, Dendrobium Moniliforme.
3. a kind of method of dendrobe tissue culture bud fast breeding according to claim 1, it is characterised in that described stem of noble dendrobium group
Training bud is the budlet that Dendrobidium huoshanness, dendrobium candidum or Dendrobium Moniliforme seed are sprouted in basic MS culture medium.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104429961A (en) * | 2014-12-08 | 2015-03-25 | 河源市裕森农林发展有限公司 | Rapid propagation tissue culture method for dendrobium officinale |
| CN104542270A (en) * | 2013-10-14 | 2015-04-29 | 厦门塔斯曼生物工程有限公司 | Rapid propagation method for medicinal dendrobium nobile |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104542270A (en) * | 2013-10-14 | 2015-04-29 | 厦门塔斯曼生物工程有限公司 | Rapid propagation method for medicinal dendrobium nobile |
| CN104429961A (en) * | 2014-12-08 | 2015-03-25 | 河源市裕森农林发展有限公司 | Rapid propagation tissue culture method for dendrobium officinale |
Non-Patent Citations (3)
| Title |
|---|
| Electrogenic machine and Nitrazyme soil conditioner: their influence on plants;Roxas, et al.;《Canopy International》;19821231;第8卷(第11期);第8-9页 * |
| 五种药用石斛快速繁殖的研究;卢文芸等;《种子》;20050531;第24卷(第5期);第23-28页，尤其是第23页摘要，第24页第1.2节 * |
| 植物组织培养电刺激效应研究进展;丁玉梅等;《生物学通报》;19981231;第33卷(第8期);第2-4页，尤其是第4页左栏第2段 * |
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