CN102845302A - Tissue culture and rapid propagation method for Chrysanthemum morifolium - Google Patents
Tissue culture and rapid propagation method for Chrysanthemum morifolium Download PDFInfo
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- CN102845302A CN102845302A CN 201110173915 CN201110173915A CN102845302A CN 102845302 A CN102845302 A CN 102845302A CN 201110173915 CN201110173915 CN 201110173915 CN 201110173915 A CN201110173915 A CN 201110173915A CN 102845302 A CN102845302 A CN 102845302A
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Abstract
The invention provides a tissue culture and rapid propagation method for Chrysanthemum morifolium. The method includes selecting stem section with bud of the Chrysanthemum morifolium plant as explant, performing sterilization, and inoculating for culture; generating callus via cluster bud induction or inducting leaves of cluster buds, and finally performing propagation and adventitious bud differentiation to the callus; and performing root induction culture, and establishing vegetative propagation system of Chrysanthemum morifolium. The invention establishes tissue culture and rapid propagation system of Chrysanthemum morifolium for the first time, and provides technical foundation for development and utilization of further economic and ecological comprehensive values of Chrysanthemum morifolium.
Description
1. technical field: the present invention relates to a kind of technical method of organizing cultivation, relates in particular a kind of quick breeding method for tissue culture of monarch's feverfew.
2. technical background: monarch's feverfew is the high-quality Dendranthema morifolium Varieties that Beijing Forestry University cultivates through concentrating on studies of more than ten years.Its inflorescence diameter mostly is 1.5～4cm, presents oblate spheroid or sphere, and the periphery is multiple white or yellow ligulate flower, and central authorities are tubular flower.Monarch's feverfew gas delicate fragrance, sweet, little hardship of distinguishing the flavor of, Hua Xingxin, sweet, bitter is slightly cold, and has dispelling wind and heat from the body, and flat liver makes eye bright, and throat-clearing throat-moistening protects medical science effect and the medical values such as liver healthy tendency.In addition, the drink tea kind of monarch's feverfew or a kind of high-quality, it is unique high-quality chrysanthemum of drinking that must not use any agricultural chemicals, contain 17 seed amino acids and 8 kinds of mineral matters that body-care needs most, still all surpassing the conventional tea chrysanthemum aspect the health care in nutrition, mouthfeel, content of vitamin E exceeds the common chrysanthemum several times of drinking in its tire chrysanthemum, has good senile-resistant efficacy.Therefore, it also is described as the show of chrysanthemum, Hua Zhikui, and green treasured, as important flower economy kind, monarch's feverfew also has very large potentiality in the exploitation of the aspects such as edible, medicinal, health care.
3. summary of the invention: the method that the purpose of this invention is to provide a kind of monarch's feverfew tissue-culturing quick-propagation, the inventor is in the research of carrying out the cultivation of monarch's feverfew tissue, basic process from the cultivation of monarch's feverfew tissue, comprise the stages such as inducing clumping bud, Callus of Leaf are induced, differentiation adventitious buds, adventitious bud rooting, filter out optimal medium, make monarch's feverfew realize effective breeding, thereby reach purpose of the present invention.
Monarch's feverfew quick breeding method for tissue culture that the present invention proposes comprises following cultivation stage:
A) selection of stem with bud and sterilization: select indoor pot monarch feverfew plant as explant material to be selected, water every day once, with 2 ‰ NaClO thimerosal plant is carried out disinfection once per month.Choose healthy and strong stem without damage by disease and insect, the following 3-4 in clip top young tender stems with bud is as explant material.At first the stem with bud material is tentatively sterilized, behind 75% alcohol infiltrating material 30S, use rapidly aseptic water washing 4 times, and then with 3%-5%NaClO solution disinfection 5min (preferred 4%), the thimerosal that ceaselessly vibrates during sterilization makes it fully contact with material, use aseptic water washing 4 times after sterilization finishes, wait to inoculate.
B) the inducing clumping bud stage: the stem with bud after will sterilizing is inoculated on the inducing culture, normally cultivate 15-20 days after, can produce indefinite bud.Described inducing culture adds 6-BA 3.0mg.L for take the MS medium as minimal medium
-1, NAA0.5mg.L
-1Other adds agar 7g.L
-1, sucrose 25g.L
-1
C) the aseptic blade that forms in the stem with bud inducing clumping bud the callus induction stage of blade: with b) is cut into the fritter about 0.5cm * 0.5cm, and the explant as the blade adventitious shoot regeneration is induced is inoculated in the callus inducing medium.The normal cultivation after 15-20 days produces a large amount of callus.Described inducing culture is: as minimal medium, add 6-BA 5.0mg.L with the MS medium
-1, NAA 1.0mg.L
-1, other adds agar 7g.L
-1, sucrose 25g.L
-1
D) fritter that the good callus that induces adventitious bud inducing differential period: with c) is cut into 1.5cm * 1.5cm is inoculated in the differential medium, and normally cultivating has indefinite bud to produce in callus after 15-20 days.Above-mentioned differential medium is: as minimal medium, add 6-BA 3.0mg.L with the MS medium
-1, NAA0.5mg.L
-1, other adds agar 7g.L
-1, sucrose 25g.L
-1
E) the culture of rootage stage: the indefinite bud cutting that inducing clumping bud and Calli Differentiation are formed is inoculated in the root media into about the little stem section of band axillalry bud of 1.5-2.0cm, generates adventive root after 25-30 days.Above-mentioned root media is: the additional 1.0mg.L take 1/2MS as minimal medium
-1IBA.
Normal condition of cultivating is in the above-mentioned steps: 25 ± 2 ℃ of room temperatures, and relative air humidity about 60%, intensity of illumination is 2000-3000lx, the photoperiod is that light is cultivated 16h, secretly cultivated 8h.
4. description of drawings:
The growth of accompanying drawing 1 blank
Accompanying drawing 2 inducing clumping buds
Accompanying drawing 3 differentiation adventitious buds
Accompanying drawing 4 adventitious bud inducings are taken root
5. embodiment:
Following examples are used for explanation the present invention, but are not used for limiting the scope of the invention.
Embodiment 1
The stem with bud of choosing indoor pot monarch feverfew plant is explant, with 75% alcohol to its 15-30S that sterilizes (preferred 30S), behind the aseptic water washing 4 times material is carried out preliminarily pasteurized, use again 3%-5%NaClO solution disinfection 5min (preferred 5%), carry out pasteurised completely behind the aseptic water washing 4 times and process.After the method sterilization, the explant pollution rate is lower than 5%.Survival rate all is higher than more than 90%.
Stem with bud after the sterilization is inoculated in the MS medium of additional different growth regulators, every bottle graft kind 2-3, place 25 ± 2 ℃ of room temperatures, relative air humidity 60%, intensity of illumination 2000-3000lx cultivates under light cultivation/16h, the dark cultivation/8h condition.After 1-2 days, axillalry bud expands rapidly sprouts into young tender vanelets, and the explant more than 95% is finished and started growth, and after cultivating through 30 days, statistics (table 1) shows, MS+3.0mg.L
-16-BA+0.5mg.L
-1The inducing clumping bud effect is best among the NAA, and growth coefficient can reach 5.5.
The different growth regulators of table 1 are on the impact of stem with bud inducing clumping bud
Embodiment 2
With the aseptic blade that forms in the inducing clumping bud as explant, be cut into the fritter about 0.5cm * 0.5cm, be inoculated in the inducing culture that adds different growth regulators and cultivate, condition of culture is 25 ± 2 ℃ of room temperatures, relative air humidity 60%, intensity of illumination 2000-3000lx, light cultivation/16h, dark cultivation/8h.After 15 days, begin that in the place that blade contacts with medium a small amount of callus growth is arranged, larger near the local callus growth amount of petiole.In the time of 15-30 days, callus growth is rapid, presents yellow green or absinthe-green graininess projection, the callus consolidation, and growing state is better.Statistics (table 2) shows, MS+5.0mg.L
-16-BA+1.0mg.L
-1Callus induction rate is the highest in the NAA medium, reaches 100%, and the callus amount is more.
The MS medium of the different growth regulators of table 2 is on the impact of callus induction
With the light green color of inducing, consolidation, well-grown callus are inoculated in the MS differential medium of additional different growth regulators, and after 20 days, the part callus begins to carry out differentiation adventitious buds, and statistics (table 3) shows, MS+3.0mg.L
-16-BA+0.5mg.L
-1NAA medium adventitious bud induction frequency can reach 28.6%, and the medium that coefficient of differentiation is the highest is MS+2.0mg.L
-16-BA+0.2mg.L
-1NAA, coefficient of differentiation are 2.3.
The different growth regulators of table 3 are on the impact of differentiation adventitious buds
Embodiment 3
Indefinite bud cutting being inoculated in the 1/2MS root media that has added different growth hormone conditioning agents with the little stem section of axillalry bud into about 1.5-2.0cm with Multiple Buds and callus induction, 1 indefinite bud of every bottle graft kind, place 25 ± 2 ℃ of room temperatures, relative air humidity 60%, intensity of illumination 2000-3000lx cultivates under light cultivation/16h, the dark cultivation/8h condition.Add up its rooting rate after 40 days, result's (table 4) is as follows, add the rooting rate that the medium of archusia can Effective Raise monarch feverfew, improved the quality of root, wherein, IBA and NAA all can make the rooting rate of monarch's feverfew reach more than 80%, aspect quality of rooting, the processing root that adds the IBA hormone is more sturdy, and the root quality is high, and the root growth that has added the NAA hormone is frail, and growing way is poor.Analysis-by-synthesis, take 1/2MS as minimal medium, additional 1.0mg.L
-1IBA is the optimal medium of monarch's feverfew adventitious bud rooting, and rooting rate can reach 86.2%, and the number of taking root is 31.
The impact of 1/2MS medium on taking root of the different growth regulators of table 4
The above only is optimal way of the present invention; should be pointed out that the common laborer for the art, under the principle prerequisite that does not break away from the present technique invention; can also make some suitable improvements and modifications, these improvements and modifications also should be considered as in the protection domain of the present invention.
Claims (6)
1. the method for monarch's feverfew tissue-culturing quick-propagation, it is characterized in that may further comprise the steps: inducing clumping bud propagation is carried out in inoculation behind the stem with bud sterilization, the fritter that the aseptic blade that again Multiple Buds is generated is cut into 0.5cm * 0.5cm carries out callus induction and adventitious bud inducing, and the indefinite bud that above-mentioned two kinds of methods are obtained carries out root induction.
2. quick breeding method for tissue culture according to claim 1, it is characterized in that, described stem with bud method for disinfection and sterilization comprises: infiltrate behind the stem with bud explant 30S aseptic water washing 4 times with 75% alcohol, again with aseptic water washing behind 3%-5%NaClO solution impregnation stem with bud explant and the thimerosal 5min that vibrates 4 times.
3. quick breeding method for tissue culture according to claim 1 is characterized in that, described stem with bud inducing clumping bud medium is: MS+3.0mg.L
-16-BA+0.5mg.L
-1NAA.
4. quick breeding method for tissue culture according to claim 1 is characterized in that, the callus inducing medium of described aseptic blade is MS+5.0mg.L
-16-BA+1.0mg.L
-1NAA; The adventitious bud inducing differential medium is: MS+3.0mg.L
-16-BA+0.5mg.L
-1NAA.
5. quick breeding method for tissue culture according to claim 1 is characterized in that, described adventitious bud rooting medium is: 1/2MS+1.0mg.L
-1IBA.
6. quick breeding method for tissue culture according to claim 1 is characterized in that, medium all adds 7g.L
-1Agar, 25g.L
-1Sucrose, PH are 5.8-6.0, and 121 ℃ of 20min of autoclaving, condition of culture are 25 ± 2 ℃ of room temperatures, relative air humidity about 60%, and intensity of illumination is 2000-3000lx, the photoperiod is that light is cultivated 16h and the dark 8h of cultivation.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103141384A (en) * | 2013-02-27 | 2013-06-12 | 上海交通大学 | Rapid tissue culture propagation method of pot chrysanthemum cultivars |
| CN108142288A (en) * | 2017-11-21 | 2018-06-12 | 山东农业大学 | A kind of method that cultured in vitro improves chrysanthemum for tea use flower sugariness |
| CN108849512A (en) * | 2018-07-16 | 2018-11-23 | 中国科学院合肥物质科学研究院 | A kind of method of spun gold emperor chrysanthemum tissue culture regeneration |
| CN112167058A (en) * | 2020-09-29 | 2021-01-05 | 四川云辰园林科技有限公司 | Rapid propagation method for improved plant tissue culture |
-
2011
- 2011-06-27 CN CN 201110173915 patent/CN102845302A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103141384A (en) * | 2013-02-27 | 2013-06-12 | 上海交通大学 | Rapid tissue culture propagation method of pot chrysanthemum cultivars |
| CN108142288A (en) * | 2017-11-21 | 2018-06-12 | 山东农业大学 | A kind of method that cultured in vitro improves chrysanthemum for tea use flower sugariness |
| CN108849512A (en) * | 2018-07-16 | 2018-11-23 | 中国科学院合肥物质科学研究院 | A kind of method of spun gold emperor chrysanthemum tissue culture regeneration |
| CN112167058A (en) * | 2020-09-29 | 2021-01-05 | 四川云辰园林科技有限公司 | Rapid propagation method for improved plant tissue culture |
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