CN107211896A - The culture medium and method of a kind of open country daisy_petal part tissue cultures - Google Patents
The culture medium and method of a kind of open country daisy_petal part tissue cultures Download PDFInfo
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- CN107211896A CN107211896A CN201710628207.7A CN201710628207A CN107211896A CN 107211896 A CN107211896 A CN 107211896A CN 201710628207 A CN201710628207 A CN 201710628207A CN 107211896 A CN107211896 A CN 107211896A
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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
-
- 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
Abstract
The present invention relates to technical field of tissue culture, the culture medium and method of more particularly to a kind of open country daisy_petal part tissue cultures.The culture medium of the open country chrysanthemum tissue cultures includes inducing culture, differential medium and root media.Inducing culture is：The MS culture mediums of the BA+1mg/L NAA+30g/L sucrose+8g/L agar of 2.0mg/L 6；Differential medium is the MS culture mediums of the BA+0.2mg/L NAA+30g/L sucrose+8g/L agar of 0.5mg/L 6；Root media is the 1/2MS culture mediums of 0.05mg/L NAA+30g/L sucrose+8g/L agar.The culture medium prescription and method provided using the present invention is remarkably improved open country chrysanthemum callus induction rate, differentiation rate and rooting of vitro seedling rate.
Description
Technical field
The present invention relates to field of plant tissue culture technique, the culture medium of specifically a kind of open country daisy_petal part tissue cultures and
Method.
Background technology
Outdoor cropping chrysanthemum is that a Xiao side for Northeast Forestry University is taught on the basis of introducing region is by the small chrysanthemum of chrysanthemum and Beijing, warp
A new ground for crossing seed selection after natural hybrization, Seeds of First Post-flight plants small chrysanthemum Breeds.Open country chrysanthemum is close because spending more, and the florescence is longer,
Plant is short, easily breeds and is widely used in the greening of north city autumn.Since the advent of the world is just low with its unique plant
It is short, bloom dense, resistance to low level management has the advantages such as environment, social and economic benefits concurrently and received much concern.
Chrysanthemum generally carries out expanding numerous using asexual reproduction methods such as plant division, cuttages for a long time.But traditional propagation method
Cycle is long, and floor space is big, and labour cost is high, and breeding coefficient is low.With sharply increasing for the market demand, traditional breeding side
The need for method can not meet people.The features such as plant tissue culture technique has fast reproduction speed, propagation efficiency high, Ke Yi
Shorter time and spatially produce substantial amounts of test tube seedling.And in the selection of explant petal not only virus quantity is few, and
Its tissue cultures has the advantages that materials, and easily, easily sterilization, breeding coefficient are high.Therefore, research and development are using open country daisy_petal part as material
Tissue-culturing rapid propagation experimental technique, it is significant to the factorial praluction of open country chrysanthemum.
The content of the invention
In view of this, the invention provides a kind of culture medium of open country daisy_petal part tissue cultures and method.The culture medium is matched somebody with somebody
Inductivity, differentiation rate and the rooting rate of open country chrysanthemum can be significantly improved.
In order to realize foregoing invention purpose, the present invention provides following technical scheme：
The invention provides a kind of culture medium of open country chrysanthemum tissue cultures, the culture medium includes inducing culture, differentiation training
Support base and root media；
The in vitro breeding method of chrysanthemum of the present invention follows these steps to realize
The petal for taking open country chrysanthemum ' powder tail feather ' is explant, and 1h is cleaned through flowing water.Under sterile conditions, petal is cut into length
8mm fritter, is seeded on inducing culture.
Inducing culture is the MS cultures of 0.5~2.0mg/L 6-BA, 1.0mg/L NAA, 30g/L sucrose and 8g/L agar
Base, pH value is 5.8.
It will be inoculated into by the callus of induced synthesis in differential medium.Differential medium is 0~1.5mg/L 6-
The MS culture mediums of BA, 0.2mg/L NAA, 30g/L sucrose and 8g/L agar, pH value is 5.8.
Seedling inoculation after differentiation culture is subjected to culture of rootage into root media.Root media is 0.05
The 1/2MS culture mediums of~0.20mg/LNAA, 30g/L sucrose and 8g/L agar, pH value is 5.8.
6-BA is 6-benzyladenine, is autosynthetic calls's mitogen, with suppress leaves of plants inner chlorophyll, nucleic acid,
Breaks down proteins, protect green anti-old；Amino acid, auxin, inorganic salts etc. a variety of efficiency such as allocate and transport to treatment site, and agricultural are used extensively
With garden crop from germinateing to harvesting each stage.
Methyl α-naphthyl acetate, abbreviation NAA is the auxin analog in a plant growth regulators, is usually used in cuttage breeding
When the root of hair powder that uses or rooting agent in.It can also be used for Plant Tissue Breeding.It is to promote cell division with expanding that it, which is acted on, is lured
Lead to form adventitious root increase fruit setting, prevent shedding, change female, male flower ratio etc..Tender epidermis that can be through blade, branch, seed enters
Enter into plant, with nutrition stream transporting to complete stool.
Above-mentioned 2 kinds of hormones are added in culture medium by the present invention with special ratios, and inducing culture, differential medium is made
And root media, when carrying out tissue cultures to open country daisy_petal part explant, it is remarkably improved the induction point of open country chrysanthemum ' powder tail feather '
Rate, rooting rate, effect are better than the kinds of culture medium of existing report.
A kind of optimal method is found out by carrying out experiment to above technical scheme, for claim.
Brief description of the drawings
Upgrowth situation after Fig. 1 open countrys daisy_petal part inoculation 3d；
The callus formed after Fig. 2 inoculations 10d；
The adventitious bud formed after Fig. 3 inoculations 25d；
The seedling of adventitious buds differentiation after Fig. 4 inoculations 35d；
Fig. 5 open countrys chrysanthemum breaks up seedling rooting culture；
Basin culture in Fig. 6 rooted seedlings.
Embodiment
The present invention is described in further detail with reference to specific example below, and introduces the correlative study knot of the present invention
Really.
1. to present invention work by taking open country chrysanthemum ' powder tail feather ' petal of gardens institute of Northeast Forestry University nursery Experimental Base as an example
It is further to describe in detail.
Using open country daisy_petal part as explant, after the petal of selection is cleaned into 1h through flowing water, with filter paper by unnecessary moisture
Blot.After 2% sodium hypochlorite sterilizing 5min, with aseptic water washing 6 times, petal is cut into the fritter for being about 8mm, is seeded in
On the inducing culture of different ratio, every kind of processing is inoculated with after 30 explants, 20d the callus induction situation that counts.
1) using MS culture mediums as minimal medium, auxin NAA 0.2mgL are added-1, because of the different designs of 6-BA concentration
4 groups of experiments.
Influence of the difference 6-BA concentration of table 1 to open country chrysanthemum ' powder tail feather ' Callus induction rate
2) using MS culture mediums as minimal medium, the callus switching that upper step is obtained is trained in the differentiation of induction Multiple Buds
Support on base (each processing inoculation 30 bottles), because of 4 groups of experiments of different designs of hormone concentration and species.Culture 2 weeks or so, observation
Adventitious bud is grown successively to callus surface (see Fig. 3).
Influence of the difference 6-BA concentration of table 2 to the callus bud ratio of open country chrysanthemum ' powder tail feather '
3) using 1/2MS culture mediums as minimal medium, because of 3 kinds of different culture of rootage of different designs of auxin concentration
Base.The high seedlings of 4cm are scaled off, root media (each 30 bottles of processing inoculation) are transferred to, 7d or so observations find most of
Test tube seedling root starts to differentiate rootlet and dashed forward, after the root that about 3~4cm length is seen after 7d (see Fig. 5).
Influence of the difference NAA concentration of table 3 to the rooting of vitro seedling rate of open country chrysanthemum ' powder tail feather '
Condition of culture：Culture medium is using MS as basic culture, culture medium 30mgL containing sucrose-1, agar 8mgL-1, training
Support base and pH value is adjusted to 5.8 before autoclaving；Cultivation temperature is 25 DEG C, and intensity of illumination is 2000lx, and light application time is 12h/
d。
In test tube height of seedling 4cm, and bottleneck is opened when sending out roots, taken out after hardening 3d, clean the culture medium of root, transplanted
To soil：Vermiculite：Perlite=4：1：In 1 culture matrix.Place at backlight, temperature is maintained at 20 DEG C.
2. experimental result
1) as shown in Table 1, No. 4 culture mediums, i.e. MS+2.0mg/L 6-BA+0.2mg/L NAA, callus induction rate is most
Height, and callus is loosely organized, upgrowth situation is good.
2) from table 2 it can be seen that No. 2 culture mediums, i.e. MS+0.5mg/L 6-BA+0.2mg/L NAA, differentiation rate highest, are inductions
The optimal medium of Multiple Buds.
3) from table 3 it can be seen that No. 3 culture mediums, i.e. 1/2MS+0.05mg/L NAA, preferably, rooting rate reaches condition of rooting
100%, and it is well developed root system, sturdy.
Claims (8)
1. a kind of culture medium of open country daisy_petal part tissue cultures, it is characterised in that：The culture medium of the open country chrysanthemum tissue cultures includes
Inducing culture, differential medium and root media；
The inducing culture is：Contain 0.5~2.0mg/L 6-BA, 1.0mg/L NAA, 30g/L sucrose and 8g/L agar
MS culture mediums, pH value is 5.8；
The differential medium is：Containing 0~1.5mg/L 6-BA, the MS of 0.2mg/L NAA, 30g/L sucrose and 8g/L agar
Culture medium, pH value is 5.8；
The root media is the 1/2MS culture mediums of 0.05~0.20mg/L NAA, 30g/L sucrose and 8g/L agar, pH value
For 5.8.
2. culture medium according to claim 1, it is characterised in that the inducing culture is 2.0mg/L 6-BA+1mg/L
The MS culture mediums of NAA+30g/L sucrose+8g/L agar；
The differential medium is：The MS culture mediums of 0.5mg/L 6-BA+0.2mg/L NAA+30g/L sucrose+8g/L agar；
The root media is：The 1/2MS culture mediums of 0.05mg/L NAA+30g/L sucrose+8g/L agar.
3. a kind of method for tissue culture of open country chrysanthemum, it is characterised in that comprise the following steps：
Open country daisy_petal part explant is seeded on inducing culture；It will turn to be inoculated into differentiation by the callus of induced synthesis
In culture medium；The seedling inoculation broken up after culture is subjected to culture of rootage into root media, tissue-cultured seedling is obtained.
4. the method for tissue culture according to right 3, it is characterised in that described be seeded in open country daisy_petal part explant lures
Also include before leading on culture medium, open country daisy_petal part is carried out disinfection after processing, petal is cut into long 8mm fritter, revealed
Ground chrysanthemum explant.
5. the method for tissue culture according to right 4, it is characterised in that described to disinfect specially：By open country chrysanthemum
Valve is sterilized with 2% sodium hypochlorite after 5min, with aseptic water washing 6 times.
6. the method for tissue culture according to any one of right 3 to 5, it is characterised in that the open country chrysanthemum tissue cultures bar
Part is that periodicity of illumination is 12h/d, and intensity of illumination is 2000lx, and cultivation temperature is 25 DEG C.
7. the method for tissue culture according to any one of right 3 to 6, it is characterised in that described to obtain also wrapping after tissue-cultured seedling
The step of including hardening.
8. the method for tissue culture according to any one of right 7, it is characterised in that the hardening is：Treat tissue culture height of seedling 4cm
And open culture bottleneck when sending out roots, taken out after 3d, wash the culture medium of root attachment off, dry and transplanted after moisture to soil：
Vermiculite：Perlite=4：1：In 1 culture matrix.Place at backlight, temperature is maintained at 20 DEG C.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108174845A (en) * | 2017-11-16 | 2018-06-19 | 海盐县凌特生物科技有限公司 | Cliff chrysanthemum bud breaks up the preparation method of culture solution |
| CN111903515A (en) * | 2020-04-21 | 2020-11-10 | 北京农业生物技术研究中心 | Method for inducing callus of 'Yutai I' petal |
| CN113115710A (en) * | 2021-05-25 | 2021-07-16 | 浙江大学 | Culture medium for inducing cut flower chrysanthemum petal callus and application |
| CN115968788A (en) * | 2022-07-28 | 2023-04-18 | 中国农业大学 | Chrysanthemum mutation breeding method based on tissue culture by using petals |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108174845A (en) * | 2017-11-16 | 2018-06-19 | 海盐县凌特生物科技有限公司 | Cliff chrysanthemum bud breaks up the preparation method of culture solution |
| CN111903515A (en) * | 2020-04-21 | 2020-11-10 | 北京农业生物技术研究中心 | Method for inducing callus of 'Yutai I' petal |
| CN111903515B (en) * | 2020-04-21 | 2021-12-24 | 北京农业生物技术研究中心 | Method for inducing callus of 'Yutai I' petal |
| CN113115710A (en) * | 2021-05-25 | 2021-07-16 | 浙江大学 | Culture medium for inducing cut flower chrysanthemum petal callus and application |
| CN115968788A (en) * | 2022-07-28 | 2023-04-18 | 中国农业大学 | Chrysanthemum mutation breeding method based on tissue culture by using petals |
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