CN104604678B - A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method - Google Patents
A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method Download PDFInfo
- Publication number
- CN104604678B CN104604678B CN201510010242.3A CN201510010242A CN104604678B CN 104604678 B CN104604678 B CN 104604678B CN 201510010242 A CN201510010242 A CN 201510010242A CN 104604678 B CN104604678 B CN 104604678B
- Authority
- CN
- China
- Prior art keywords
- culture
- panax japonicus
- sugar
- seedling
- explant
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Landscapes
- Cultivation Of Plants (AREA)
Abstract
The invention discloses a kind of wild panax japonicus sugar-free open tissue and cultivate fast seedling-cultivating method, A, the stem of getting wild panax japonicus seedling or leaf, be seeded to through surface sterilizing treatments B, by the explant of processing of step A in the first culture base of panax japonicus sugar free tissue culture, the clear glass growth chamber that is placed in upper shed carries out just culture; C, by step B cultivate screen and use aseptic water washing without offspring, be cut into single seedling by what rinse well without offspring; D, by through step C process single seedling be seeded in panax japonicus sugar free tissue culture subculture medium, be placed in clear glass growth chamber and carry out subculture cultivation. In E, soil that the plantlet of transplant obtaining in D is mixed in fertile soil and vermiculite, cultivate in the usual way. Utilize the method for the invention, the differentiation rate of explant is more than 90%, and appreciation rate can reach 4.2, and pollution rate is few, acquisition be all can be for the qualified healthy and strong group training seedling of plantation, transplanting survival rate is more than 98%.
Description
Technical field
The present invention relates to field of plant tissue culture technique, is mainly that the sugar-free open tissue of wild panax japonicus is cultivated fast seedling growingMethod.
Background technology
Panax japonicus (PanaxjaponicusC.A.Meyer) is Araliaceae Panax herbaceos perennial, is China's Precious, Rare, EndangeredOne of famous and precious-seven class ‖ Chinese herbal medicines, the king ‖ of be described as-herbal medicine, have-‖ north medicine-ginseng tonic strong and-Nan medicine ‖ pseudo-ginseng blood-circulation-invigovatingEffect of the stasis of blood, has anti-inflammatory and antalgic, immunological regulation, resists myocardial ischemia, hypoglycemic, anti-ageing, antifatigue, anticancer and improve and learnPractise the multiple biologically actives such as memory function.
In recent years, be widely used in clinically because panax japonicus has multiple pharmacological effect, cause its demand to increase, for a long time withCause because panax japonicus medical value is high among the peoplely excavating in the mode of sacrificing future gains to satisfy present needs, this utilizes behavior just to make originallyRare wild panax japonicus resource becomes by endangered. So only rely on the supply of wild bulk drug, at quality and quantityOn be all difficult to meet the needs of domestic medical market. Therefore, carry out that panax japonicus is extensive, industrial seedling rearing is to preserve panax japonicusThe important foundation of germ plasm resource and exploitation is also the key of lengthening manufacturing chain.
Develop aspect at panax japonicus saponin class material, along with the extraction of panax japonicus saponin class material, the one-tenth increasingly of isolation technicsRipe, some medicine manufacturers fall over each other to develop oral liquid, capsule, compound buccal lozenge and extract the products such as intermediate, and Japan, easternThe foreign traders such as Europe also require to order panax japonicus saponin from China one after another, visible, greatly develop taking panax japonicus medicinal material as raw-material industryChain, has huge market prospects and economic benefit.
Most scholars have done greatly panax japonicus at aspects such as application among the people, chemical constitution study, Analysis on Biological Activity, cultivation managementsThe research work of amount, for deep level development utilization and the industrialized development of panax japonicus provide valuable scientific information, but will realizeThe sustainable use of panax japonicus plant resources, also has a series of work to do, and at present in the face of industrialized development is badly in need of solving seedlingProblem.
Quantity research confirms greatly, and well differentiated plant cell is still keeping the totipotency of Cell Differentiation. Based on plant cellTotipotency, set up plant culture system in vitro, can breed out at short notice a large amount of plant seedling, be solve current bambooThe key of contradiction between joint ginseng plasm resource protection and rational exploitation and utilization. In theory, each organ of plant, histocyteAnd protoplast can do explant. But in actual tissue incubation, need to select explant, because differentThe explant in source, the sensitiveness difference in vitro to envirment factor, growth potential is also different, cellular-restoring divisionThe ability that ability or callus break up is again also variant. If select badly, be just difficult to obtain the success of cultivating. Therefore,The selection of suitable explant is to guarantee whether successfully crucial tissue is cultivated.
At present, panax japonicus belongs to each position, as stem apex, and stipes, cortex, cotyledon, bennets etc. can successfully induce callus groupKnit. Research shows that panax japonicus seminal propagation speed is slow. The research of refined (2000) tried to gain shows, the blade in panax japonicus platymisciumInductivity as its callus of explant is the highest, reaches 100%. At present at application number: in 200610052487.3 patentThe explant adopting is exactly leaf and the stem of wild panax japonicus.
Plant tissue culture technique is since 20 beginnings of the century set up, and development in theoretical research and application technology, extensively shouldFor aspects such as the Fast-propagation of plant, breed improvement, genetic engineering breeding, germ plasm resource preservation, secondary metabolite productions,Produce huge economic benefit and social benefit, modern agriculture and medicine and other fields produced to profound influence. But these biographiesSystem tissue culture technology, the seedling poor growth of cultivating, seriously polluted, survival rate is low etc., can cause high cost, is unfavorable for group trainingThe large-scale commercial production of seedling, cannot meet market to the high-quality demand of hyte training seedling at a low price. Along with to Plant Tissue BreedingThe research that deepens continuously of technology, Sugar Free Plant Tissue Culture fast breeding technique will address these problems.
Tradition panax japonicus tissue is cultivated the sugar mainly relying in culture medium as carbon source, very easily causes the pollution of microorganism. In order to subtractThe pollution of few microorganism, must be used culture vessel sealing, little. Culture vessel is little, and depositing due to sugar in culture medium, cause blade top layer structural development poor, stomatal movement function is not strong, and blade is little, and chlorophyll content is low, finally suppresses and fallsThe low photosynthesis ability of plantlet. With environment in the container of plant self-sow difference great disparity, also cause plant physiology, shapeCertain disorder in state, the meeting that is difficult to adapt to is grown and is delayed or death, and what have there will be plant strain growth thin and delicate, and blade is unfoldedIt is poor to spend, and takes root bad, causes the higher death rate of domestication stage plant. Tradition tissue is cultivated taking fluorescent lamp as light source, plant pairIts light source availability is low.
For solving the technical bottleneck of panax japonicus industrialized development, we carried out taking panax japonicus stem and leaf as basic sugar-free openTissue is cultivated, with CO2Replace the carbon source of sugar as plant, set up plant regeneration system, to meet medicinal herb grower to panax japonicus kindThe demand of seedling.
Summary of the invention
The present invention will solve above-mentioned existing issue, shortens cultivation cycle, and reduction group training seedling production cost; Provide a kind of with open countryRaw panax japonicus stem and leaf are that basic sugar-free open tissue is cultivated fast seedling-cultivating method.
In order to achieve the above object, technical scheme of the present invention is:
A kind of wild panax japonicus sugar-free open tissue is cultivated method for culturing seedlings, and its step is as follows:
The choosing and disinfecting of A, explant: get stem or the leaf of wild panax japonicus seedling, after surface sterilizing is processed,Under aseptic condition, stem segment (1cm is long), leaf are cut into slices to (1cm × 1cm) as explant of organizing training use;
B, just for sugar free tissue culture: be just commissioned to train being seeded to panax japonicus sugar free tissue culture through the explant of processing of step ASupport in base, the clear glass growth chamber that is placed in upper shed carries out just culture;
C, screen without offspring: that will cultivate through step B screens without offspring, and selecting size is the nothing of 1 – 2.5cmOffspring is also used aseptic water washing, is cut into the mono-seedling of 1-2.5cm by what rinse well without offspring;
D, subculture sugar free tissue culture: single seedling of processing through step C is seeded to panax japonicus sugar free tissue culture subculture and cultivatesIn base, be placed in clear glass growth chamber and carry out subculture cultivation.
The transplanting of E, plant: be 5:1 by volume in fertile soil and vermiculite by the plantlet of transplant high 3～5cm obtaining in DIn the soil mixing, cultivate in the usual way.
The described panax japonicus sugar free tissue culture just formula of culture base is: the MS+GA of not sugary and agar3The leech of 0.8～1.2mg/L+6-BA2.0～3.5mg/L+ coconut milk, 100-150g/L+ active carbon, 200-300mg/L+0.25 millimeterStone 8-10g/L, PH is 5.8. Optimization formula is not sugary and MS culture medium+1.2mg/L or 1.0mg/L agarGA3The vermiculite of+3.0mg/L6-BA+130g/L coconut milk+250mg/L active carbon+10g/L0.25 millimeter, PH is 5.8; WhenWhen explant is stem, GA in culture medium3Content be 1.2mg/L; GA in culture medium in the time that explant is leaf3Content be1.0mg/L; The content of 6-BA is 3.0mg/L.
The cultivation cycle of step B 25-45 days, CO2Concentration 650 ± 50 μ L/L, 22 DEG C-27 DEG C of cultivation temperature, utilize LEDLamp carries out illumination, light quality ruddiness: blue light is 1:1-3:1, intensity of illumination 1500-2500Lx, light application time 10-16 hours/ day. Preferably condition of culture is CO2Concentration 650 μ L/L, 24 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness:Blue light is 2:1 intensity of illumination 2000Lx, light application time 13 hours/day.
The formula of the panax japonicus sugar free tissue culture subculture medium described in step D is: the MS+6-BA of not sugary and agarThe leech of 0.8～1.2mg/L+IBA2.7～3.3mg/L+ coconut milk, 100-150g/L+ active carbon, 300-600mg/L+0.25 millimeterStone 8-10g/L, pH is 5.8. Preferably subculture medium formula is: the MS culture medium+1.0mg/L of not sugary and agarThe vermiculite of 6-BA+3.0mg/LIBA+130g/L coconut milk+450mg/L active carbon+10g/L0.25 millimeter, PH is 5.8.
The cultivation cycle of step D 30-50 days, CO2Concentration 650 ± 50 μ L/L, 22 DEG C-27 DEG C of cultivation temperature, utilize LEDLamp carries out illumination, light quality ruddiness: blue light is 1:1-3:1, intensity of illumination 1500-2500Lx, light application time 10-16 hours/ day. Preferably condition of culture is CO2Concentration 650 μ L/L, 24 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness:Blue light is 2:1, intensity of illumination 2000Lx, light application time 15 hours/day.
Described upper shed clear glass growth chamber is provided with from the above 0.5cm of fluid nutrient medium liquid level place in incubatorAir inlet is provided with capnometer from the above 2cm of fluid nutrient medium liquid level place in incubator, and carbon dioxide is with emptyThe mist of gas enters in growth chamber by air inlet, by adjusting the ratio of carbon dioxide and air, makes titanium dioxideThe gas concentration lwevel that carbon detector detects is at 650 ± 50 μ L/L.
This panax japonicus sugar free tissue culture method of the present invention and traditional panax japonicus method for tissue culture advantage are:
(1) change of carbon source: in traditional plant tissue culture fast breeding technique, plantlet with sugar (as sucrose, glucose, reallySugar etc.) as main carbon source carry out heterotrophism or double health long, sugar is counted as requisite material in Plant Tissue Breeding. AndIn the present invention, use CO2Substitute the sole carbon source of sugar as plantlet growth, supply with plantlet by mandatory air exchange system rawLong required CO2, make it under artificial lighting, absorb CO2Carry out autophyting growth completely, avoided to a certain extent micro-Biological pollution.
(2) change of culture matrix: in traditional tissue is cultivated, conventionally use agar as culture matrix, it ventilativeProperty is poor, is unfavorable for movement and the absorption of moisture, gas and nutriment. The present invention uses vermiculite as culture matrix, this matrixThere is good gas permeability, can greatly improve rooting rate and the quality of rooting of plantlet, and compared with agar, this nothingMachine matrix relative low price, has saved cultivation cost.
(3) change of culture vessel: in traditional tissue is cultivated, due to the existence of sugar in culture medium, in order to prevent polluting,Can only use little culture vessel. The present invention does not use sugar and all kinds of organic substances in incubation, has greatly avoided pollutionGeneration, therefore select glass shell growth chamber that light transmission is good as culture vessel, both can Artificial Control CO2DenseDegree again can Artificial Control intensity of illumination, makes full use of light source.
(4) change of light source: in traditional tissue is cultivated, normally used fluorescent lamp is as light source, has that volume is large, the longevityOrder the shortcomings such as short, high, the wavelength of consuming energy is fixing, heating is high. Only affect one of key factor of growth and development of plants, light qualityGrowth, morphogenesis, photosynthesis, metabolism and gene expression to plant all have regulating and controlling effect. Novel illumination lightIts wavelength of light emitting diode (lightemittingdiode, LED) in source is the spectrum of and photomorphogenesis synthetic with Plant Light just in timeScope is coincide, and luminous energy effective rate of utilization can reach 80%～90%, and can realize control separately to different light medium and luminous intensity. ?Recent studies on discovery, the fluorescent lamp that the LED light source that light quality ratio and intensity of illumination are adjustable uses than common Plant Tissue Breeding more canEffectively promote the photosynthesis of test-tube plantlet and grow. In Plant Tissue Breeding, adopt LED that illumination is provided, regulation and control lightMatter and photosynthetic photon quanta flux density, not only can regulation and control group cultivate growing and morphogenesis of thing, shortens cultivation cycle,Can also save energy consumption, reduce production costs. In addition, LED also has that volume is little, the life-span is long, it is low to consume energy, wavelength is solidAdvantages such as determining, generate heat and be low, and can also carry out according to the growth needs of plant the accurate configuration of luminescent spectrum, realize conventional light sourceThe functions such as irreplaceable energy-saving and environmental protection and space-efficient utilization.
(5) subculture and process of rooting culture unite two into one, and there is no the generation of callus; Cultivation cycle has shortened more than 40%.
(6) reduce investment outlay, reduce production costs. Compared with traditional micro-propagation technique, seedling is produced integrated cost and is on average fallenLow by 30%. The simplification of group training production technology, flow process shortens, and the integrated level of technology and equipment improves, and has reduced operating technologyDifficulty and labor operation intensity, be easier to apply in large-scale production.
(7) utilize the method for the invention, the differentiation rate of explant is more than 90%, and appreciation rate can reach 4.2, and pollution rate is few,What obtain is all can be for the qualified healthy and strong group training seedling of plantation, and transplanting survival rate is more than 98%.
Detailed description of the invention
Below in conjunction with embodiment, the invention will be described further: but content of the present invention is not limited to this.
Embodiment 1:
(1), the choosing and disinfecting of explant: get the stem of wild panax japonicus seedling in 5～June as explant, soakedIn the beaker that bubble is 75% at alcohol volumn concentration, within 45 seconds, carry out sterilizing, then use aseptic water washing 2 times; At asepticUnder part, stem is cut into the long explant as group training use of 1cm;
(2), just for sugar free tissue culture: at the beginning of the explant of processing through step (1) is seeded to panax japonicus sugar free tissue cultureIn culture base, be placed in upper shed clear glass growth chamber and carry out just culture. Cultivation cycle is 35 days, CO2Concentration650 μ L/L, cultivation temperature is 24 DEG C, utilizes LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination is 2000Lx,Light application time 13 hours/day.
Described panax japonicus sugar free tissue culture just culture base is: the MS culture medium+1.2mg/L of not sugary and agarGA3The vermiculite of+3.0mg/L6-BA+130g/L coconut milk+250mg/L active carbon+10g/L0.25 millimeter, pH is 5.8.
Explant differentiation rate reaches 90-93%, is cultivating the length generating afterwards for 35 days without offspring at 1.4cm-2.2cm, and without callusGenerate.
Described upper shed clear glass growth chamber is provided with from the above 0.5cm of fluid nutrient medium liquid level place in incubatorAir inlet is provided with capnometer from the above 2cm of fluid nutrient medium liquid level place in incubator, and carbon dioxide is with emptyThe mist of gas enters in growth chamber by air inlet, by adjusting the ratio of carbon dioxide and air, makes titanium dioxideThe gas concentration lwevel that carbon detector detects is at 650 μ L/L.
(3), screen without offspring: that will cultivate through step (2) screens without offspring, selects size to reach 1.7-2.2cm'sWithout offspring and with aseptic water washing, be cut into single seedling by what rinse well without offspring;
(4), subculture sugar free tissue culture: single seedling of processing through step (3) is seeded to panax japonicus sugar free tissue culture subcultureIn culture medium, be placed in upper shed clear glass growth chamber and carry out subculture cultivation. Cultivation cycle is 40 days, CO2Concentration650 μ L/L, cultivation temperature is 24 DEG C, utilizes LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination is 2000Lx,Light application time 15 hours/day.
Described panax japonicus sugar free tissue culture subculture medium is: the MS culture medium+1.0mg/L of not sugary and agarThe vermiculite of 6-BA+3.0mg/LIBA+130g/L coconut milk+450mg/L active carbon+10g/L0.25 millimeter, pH is 5.8.
Cultivate 40 days afterwards average value-added coefficient be 3.9.
(5) transplanting of plant: in fertile soil and vermiculite be by volume by the plantlet of transplant high 3～5cm obtaining in (4)In the soil that 5:1 mixes, carry out routine and cultivate.
Embodiment 2:
A kind of wild panax japonicus sugar-free open tissue is cultivated method for culturing seedlings, comprises the following steps:
(1), the choosing and disinfecting of explant: get the leaf of wild panax japonicus seedling in 5～June as explant, soakedIn the beaker that bubble is 75% at alcohol volumn concentration, within 60 seconds, carry out sterilizing, then use aseptic water washing 2 times; At asepticUnder part, leaf is cut into the explant of 1cm × 1cm size as group training use;
(2), just for sugar free tissue culture: at the beginning of the explant of processing through step (1) is seeded to panax japonicus sugar free tissue cultureIn culture base, be placed in upper shed clear glass growth chamber and carry out just culture. Cultivation cycle is 35 days, CO2Concentration650 μ L/L, cultivation temperature is 24 DEG C, utilizes LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination is 2000Lx,Light application time 13 hours/day.
Described panax japonicus sugar free tissue culture just culture base is: the MS culture medium+1.0mg/L of not sugary and agarGA3The vermiculite of+3.0mg/L6-BA+130g/L coconut milk+250mg/L active carbon+10g/L0.25 millimeter, pH is 5.8.
Described upper shed clear glass growth chamber is provided with from the above 0.5cm of fluid nutrient medium liquid level place in incubatorAir inlet is provided with capnometer from the above 2cm of fluid nutrient medium liquid level place in incubator, and carbon dioxide is with emptyThe mist of gas enters in growth chamber by air inlet, by adjusting the ratio of carbon dioxide and air, makes titanium dioxideThe gas concentration lwevel that carbon detector detects is at 650 μ L/L.
After step (2), explant differentiation rate reaches 96%-98%, within 35 days, generates afterwards without the length of offspring and exists in cultivation1.5-2.5cm, and generate without callus.
(3), screen without offspring: that will cultivate through step (2) screens without offspring, selects size to reach 1.8-2.5cm'sWithout offspring and with aseptic water washing, be divided into single seedling of each 1.8-2.5cm by what rinse well without offspring;
(4), subculture sugar free tissue culture: single seedling of processing through step (3) is seeded to panax japonicus sugar free tissue culture subcultureIn culture medium, be placed in upper shed clear glass growth chamber and carry out subculture cultivation. Cultivation cycle is 40 days, CO2Concentration650 μ L/L, cultivation temperature is 24 DEG C, utilizes LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination is 2000Lx,Light application time 15 hours/day.
Described panax japonicus sugar free tissue culture subculture medium is: the MS culture medium+1.0mg/L of not sugary and agarThe vermiculite of 6-BA+3.0mg/LIBA+130g/L coconut milk+450mg/L active carbon+10g/L0.25 millimeter, pH is 5.8.
Cultivate 40 days afterwards average value-added coefficient be 4.2.
(5) transplanting of plant: in fertile soil and vermiculite be by volume by the plantlet of transplant high 3～5cm obtaining in (4)In the soil that 5:1 mixes, carry out routine and cultivate.
The plant that ruddiness is cultivated is similar to shade plant, and ruddiness can promote the growth of seedling, the seedling dry-matter accumulation of ruddiness processingMany, nourish and grow vigorous. Panax japonicus belongs to shade plant; So ruddiness will be more than blue light when illumination, intensity of illumination relatively a little less than,Be more conducive to the growth of group training seedling.
Show by above-mentioned experiment: in two groups of embodiment culture mediums of the present invention, pollution rate, within 1%, only has a small amount of bacterial plaque, not shadowRing the normal growth of group training seedling; About 75 days, reach the high 4 ㎝ left and right of stem in cultivation cycle, 4 of radicals, 5 of the numbers of blade canFor the qualified healthy and strong group training seedling of planting, in just for sugar free tissue culture taking leaf as explant is than the nothing obtaining taking stem as explantIt is healthy and strong that offspring is wanted, and may be because blade can better carry out photosynthesis, utilizes carbon source. Its group training transplantation of seedlings survival rate reachesTo more than 98%.
Claims (5)
1. wild panax japonicus sugar-free open tissue is cultivated a method for culturing seedlings, and its step is as follows:
A. the choosing and disinfecting of explant: get stem or the leaf of wild panax japonicus seedling, after surface sterilizing is processed, under aseptic condition, stem segment, leaf section are trained to the explant of use as group;
B. just for sugar free tissue culture: will be seeded to through the explant of processing of step A in the first culture base of panax japonicus sugar free tissue culture, the clear glass growth chamber that is placed in upper shed carries out just culture;
C. screen without offspring: by screening without offspring of cultivating through step B, select size be 1-2.5cm also use aseptic water washing without offspring, be cut into the mono-seedling of 1-2.5cm by what rinse well without offspring;
D. subculture sugar free tissue culture: single seedling of processing through step C is seeded in panax japonicus sugar free tissue culture subculture medium, is placed in clear glass growth chamber and carries out subculture cultivation;
E. the transplanting of plant: in the soil that the plantlet of transplant high 3～5cm obtaining in D is mixed for 5:1 by volume in fertile soil and vermiculite, cultivate in the usual way;
The described panax japonicus sugar free tissue culture just formula of culture base is: the MS+GA of not sugary and agar3Vermiculite 8-the 10g/L of 0.8～1.2mg/L+6-BA2.0～3.5mg/L+ coconut milk, 100-150g/L+ active carbon, 200-300mg/L+0.25 millimeter, pH is 5.8;
The cultivation cycle of step B 25-45 days, CO2Concentration 650 ± 50 μ L/L, 22 DEG C-27 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness: blue light is 1:1-3:1, intensity of illumination 1500-2500Lx, light application time 10-16 hours/days;
The formula of described panax japonicus sugar free tissue culture subculture medium is: the vermiculite 8-10g/L of MS+6-BA0.8～1.2mg/L+IBA2.7～3.3mg/L+ coconut milk 100-150g/L+ active carbon 300-600mg/L+0.25 millimeter of not sugary and agar, and pH is 5.8;
The cultivation cycle of step D 30-50 days, CO2Concentration 650 ± 50 μ L/L, 22 DEG C-27 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness: blue light is 1:1-3:1, intensity of illumination 1500-2500Lx, light application time 10-16 hours/days;
The clear glass growth chamber of described upper shed is provided with air inlet from the above 0.5cm of fluid nutrient medium liquid level place in incubator, in incubator, be provided with capnometer from the above 2cm of fluid nutrient medium liquid level place, the mist of carbon dioxide and air enters in growth chamber by air inlet, by adjusting the ratio of carbon dioxide and air, make the gas concentration lwevel of carbon dioxide detector detection at 650 ± 50 μ L/L.
2. method according to claim 1, is characterized in that: the described panax japonicus sugar free tissue culture just formula of culture base is: MS culture medium+1.2mg/L or the 1.0mg/LGA of not sugary and agar3The vermiculite of+3.0mg/L6-BA+130g/L coconut milk+250mg/L active carbon+10g/L0.25 millimeter, pH is 5.8; In the time that explant is stem, GA in culture medium3Content be 1.2mg/L; GA in culture medium in the time that explant is leaf3Content be 1.0mg/L; The content of 6-BA is 3.0mg/L.
3. method according to claim 1, it is characterized in that: the formula of described panax japonicus sugar free tissue culture subculture medium is: the vermiculite of MS culture medium+1.0mg/L6-BA+3.0mg/LIBA+130g/L coconut milk+450mg/L active carbon+10g/L0.25 millimeter of not sugary and agar, pH is 5.8.
4. method according to claim 1, is characterized in that: the condition of culture of step B is CO2Concentration 650 μ L/L, 24 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination 2000Lx, light application time 13 hours/day.
5. method according to claim 1, is characterized in that: the condition of culture of step D is CO2Concentration 650 μ L/L, 24 DEG C of cultivation temperature, utilize LED lamp to carry out illumination, light quality ruddiness: blue light is 2:1, intensity of illumination 2000Lx, light application time 15 hours/day.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510010242.3A CN104604678B (en) | 2015-01-09 | 2015-01-09 | A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510010242.3A CN104604678B (en) | 2015-01-09 | 2015-01-09 | A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104604678A CN104604678A (en) | 2015-05-13 |
| CN104604678B true CN104604678B (en) | 2016-05-04 |
Family
ID=53139554
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510010242.3A Expired - Fee Related CN104604678B (en) | 2015-01-09 | 2015-01-09 | A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104604678B (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106520664A (en) * | 2016-11-15 | 2017-03-22 | 天津市博爱生物药业有限公司 | Synthetic medium for cells of panax japonicus |
| CN106954551A (en) * | 2017-04-28 | 2017-07-18 | 上海离草科技有限公司 | Sugar Free Plant Tissue Culture special culture media |
| CN107318649B (en) * | 2017-06-29 | 2019-06-28 | 河北农业大学 | Culture medium and culture method for reducing browning rate of walnut explants |
| CN116724890A (en) * | 2023-06-15 | 2023-09-12 | 德兴市中医研究院试验培训基地 | Open cultivation method for plant tissue culture seedlings |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH08140672A (en) * | 1994-11-16 | 1996-06-04 | Eniwa Res Business Park Kk | Culture of hairy roots of tochiba ninjin and production of panax saponin |
| CN100407906C (en) * | 2006-07-10 | 2008-08-06 | 浙江省农业科学院 | Tissue-culturing quick propagation method of wild rhizoma panacis japonici |
| CN102084813A (en) * | 2010-12-23 | 2011-06-08 | 福建永安天奇健金线莲生态实业有限公司 | Method for culturing sugar-free tissue of jewel orchid |
| CN104186331A (en) * | 2014-09-12 | 2014-12-10 | 南京通泽农业科技有限公司 | Quick propagation method of panax japonicus C.A . Mey. var. |
-
2015
- 2015-01-09 CN CN201510010242.3A patent/CN104604678B/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| CN104604678A (en) | 2015-05-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102119655B (en) | Natural light rapid breeding method for dendrobium officinale | |
| CN101822214A (en) | Method for breeding minitype potato seeds by directly using potato stem segments | |
| CN104604678B (en) | A kind of wild panax japonicus sugar-free open tissue is cultivated fast seedling-cultivating method | |
| CN104099287B (en) | The acquisition of high yield OPC Vitis davidii Foex callus and subculture keeping method | |
| CN102577976A (en) | Simple tissue culture method for broussonetia papyrifera | |
| CN103120126A (en) | Method for carrying out anoectochilus formosanus tissue culture propagation by using intermittent immersion bioreactor | |
| CN101720670A (en) | Rapid breeding method for pinellia tuber tissue culture | |
| CN103975834A (en) | In-house production technology for hydroponic swamp cabbages | |
| CN1328951C (en) | Dendrobe protocorm hormoneless cultivation method | |
| CN103098712B (en) | Davallia mariesii breeding method | |
| CN107041309A (en) | A kind of method of fast breeding Chinese tallow tree plant | |
| CN102948325A (en) | Cordyceps militaris efficient quick cultivation technology | |
| CN106258960A (en) | A kind of orchid seed germination quick-breeding method | |
| CN103609452B (en) | Tissue-culture rapid propagation method of cymbidium | |
| CN103461119B (en) | PIECA ASPERATA somatic embryo occurs and plant regeneration method | |
| CN103858760A (en) | Manufacturing method of test-tube type anoectohilus formosanus | |
| CN105432470A (en) | Tissue culture method for dendrobium moniliforme | |
| CN110432154A (en) | A kind of cultural method of spinulose tree fern tissue | |
| CN110089432A (en) | Paper mulberry sugar-free tissue culture method based on environmental factor regulation | |
| CN105766338A (en) | Seedling culture method of Abelmoschus esculentus | |
| KR20080073388A (en) | Mass production of bulblet via somatic embryogenic cell culture in lily | |
| CN105494105B (en) | A kind of peony tissue culture vessel seedling technology | |
| CN107821162A (en) | A kind of large-scale method for producing of babysbreath Plug seedling | |
| CN103125387A (en) | Method for producing lily bulb by using plant tissue culture technology | |
| CN102823499A (en) | Factorized breeding method of blueberry tissue-cultured seedlings |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20160504 Termination date: 20200109 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |