CN107360973B - A method of induction flowering dogwood callus formation - Google Patents
A method of induction flowering dogwood callus formation Download PDFInfo
- Publication number
- CN107360973B CN107360973B CN201710686445.3A CN201710686445A CN107360973B CN 107360973 B CN107360973 B CN 107360973B CN 201710686445 A CN201710686445 A CN 201710686445A CN 107360973 B CN107360973 B CN 107360973B
- Authority
- CN
- China
- Prior art keywords
- flowering dogwood
- callus
- induction
- stem
- flowering
- 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.)
- Active
Links
Classifications
-
- 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
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a kind of methods that induction flowering dogwood callus is formed, comprising: using the stem of flowering dogwood as explant, is inoculated on culture medium after disinfection and carries out illumination cultivation；The wherein formula of the culture medium are as follows: basal medium+6-BA 1-2mg/L+NAA 0.1-0.3mg/L+ sucrose 30-40g/L+ agar 6-9g/L.The present invention explores actively and effective flowering dogwood stem tissue cultures new method, and culture medium, condition of culture, sterilization method are optimized, and can induce in a short time and obtain a large amount of callus.
Description
Technical field
The present invention relates to tissue cultures more particularly to a kind of methods that induction flowering dogwood callus is formed.
Background technique
Flowering dogwood, Cornaceae, Lai wood category, also known as great Hua Dendronenthamia japonica var.chinensis originate in eastern North America, the flowering dogwood four seasons
Scenery is had nothing in common with each other, and spring petal-shaped bract is in full bloom, the splendid blade of summer and autumn colored, the berry of Hua Houyou salmon pink, is high
The landscape plant and oilseed plant, wide market, China of shelves are still in the stage of introducing a fine variety.Flowering dogwood is generally broadcast using seed
Kind, cottage method breeding.This patent carries out tissue culture propagation to flowering dogwood, by cultivating flowering dogwood organ-tissue,
Promote calli induction, to cultivate the nursery stock of more flowering dogwoods, this is the important of flowering dogwood tissue cultures
Approach.
Flowering dogwood is new introduced variety, thus it is domestic seldom to the research of the tissue cultures related fields of flowering dogwood,
But the research for thering are a small number of scholars to carry out some tissue cultures for the other plant that Lai wood belongs to, but breeding coefficient compares
It is low.
Summary of the invention
Goal of the invention: being directed to the problems of the prior art, and the present invention provides a kind of induction flowering dogwood callus and formed
Method, using stem as explant, induction obtains a large amount of callus in a short time.
Technical solution: the method that induction flowering dogwood callus of the present invention is formed, comprising: with flowering dogwood
Stem is explant, is inoculated on culture medium after disinfection and carries out illumination cultivation；The wherein formula of the culture medium are as follows: basal medium
+ 6-BA 1-2mg/L+NAA 0.1-0.3mg/L+ sucrose 30-40g/L+ agar 6-9g/L.
The explant the preparation method comprises the following steps: acquisition flowering dogwood current-year branch tender stem, shearing growth 1-3cm stem
Section.
The method of the disinfection includes: after cleaning explant, first to use 70-75% ethanol postincubation 30-50s, and sterile water is clear
It is carried out disinfection processing with 0.1-0.2% mercuric chloride again after washing, the processing time is 3-6min, finally clean with aseptic water washing, removal
Residual moisture.
The time of ethanol postincubation is preferably 30s, and the processing time of mercuric chloride is preferably 5min.
The formula of culture medium is preferred are as follows: basal medium+6-BA 1.5-2mg/L+NAA 0.1-0.2mg/L+ sucrose 30-
40g/L+ agar 6-9g/L.
It is further preferred that the formula of culture medium are as follows: basal medium+6-BA 2mg/L and NAA 0.2mg/L+sucrose
35g/L+ agar 7g/L.
The basal medium is WPM.
The temperature of the illumination cultivation is 23-27 DEG C, light application time 16-17h/d, illuminance 2000-2200lx.
The utility model has the advantages that the present invention explores actively and effective flowering dogwood stem tissue cultures new method, to culture medium, training
The condition of supporting, sterilization method are optimized, and can induce in a short time and obtain a large amount of callus.
Detailed description of the invention
Fig. 1 is the flowering dogwood stem explant that embodiment 1 is being cleaned；
Fig. 2 is the partial medium that embodiment 1 prepares sterilizing；
Fig. 3 is that embodiment 1 is just being inoculated into the stem explant on culture medium；
Fig. 4 is that embodiment 1 is placed on the inoculation seedling cultivated in culturing room；
Fig. 5 is the inoculation seedling that embodiment 1 pollutes；
Fig. 6 is the inoculation seedling that embodiment 1 has formed a large amount of callus.
Specific embodiment
Combined with specific embodiments below, the present invention is furture elucidated, it should be understood that these embodiments are merely to illustrate the present invention
Rather than limit the scope of the invention, after the present invention has been read, those skilled in the art are to various equivalences of the invention
The modification of form falls within the application range as defined in the appended claims.
Embodiment 1
1, acquire and clean explant
Flowering dogwood current-year branch tender stem is acquired, the stem section for being about 1-3cm is cut into, is put into ceramic whiteware cylinder few with having added
The water for measuring washing powder eluriates 2min, rinses 2h under tap water after foam is eluriated completely.Surface dirt and soil will be eliminated
Explant be placed on superclean bench and disinfect.
2, carry out disinfection processing
It is then primary with sterile water wash first with 75% ethanol postincubation 30-50s, then carried out disinfection place with 0.1% mercuric chloride
Reason, the processing time is 3-6min, then with aseptic water washing 3 times or more, remaining moisture is drawn with aseptic paper.
3, inoculation work is carried out
The stem of sanitized is placed on aseptic paper, is inoculated on culture medium with tweezers, culture medium select respectively WPM,
MS, 1/2MS, B5, N6, select the concentration of different 6-BA and NAA: 6-BA concentration is respectively 0,0.5,1,2mg/L, and NAA's is dense
Degree is 0,0.02,0.2,2mg/L respectively, studies the influence for generating callus under hormon various concentration to explant.Often
One 30 bottles of processing inoculation, is repeated 3 times, every test tube bottle places 1 explant.
4, condition of culture is controlled
All culture mediums add sucrose 35g/L, agar 7g/L, and adjusting pH value is 5.8 or so, under 121 DEG C of high pressures
Sterilize 25min.Explant after inoculation is placed in culturing room, illumination 16h/d, illuminance is about 2000lx, and temperature control exists
(25±2)℃。
5, result and analysis
5.1 obtain the most suitable disinfecting time of flowering dogwood stem
The influence of the different disinfection way of table 1
By table 1 we it can be found that using alcohol impregnate 30s, mercuric chloride impregnate this disinfection way of 5min it is best, survival
Rate is 89%, and the death rate and pollution rate are minimum compared with other processing modes, therefore, impregnates 30s using alcohol, mercuric chloride impregnates
This disinfection way of 5min reduces pollution rate most beneficial for survival rate is improved.
5.2 obtain flowering dogwood most suitable culture medium in callus forming process
It is analyzed by this following data, we are it can be concluded that WPM and N6 formed flowering dogwood stem callus
Influence is more more significant than other culture mediums, and wherein WPM is better than N6 again, it may thus be appreciated that production of the WPM to the callus of flowering dogwood stem
Coming into force, (table 3 is size (being shown in Table 2, the unit cm) progress that 1 half a month forms callus during tissue culture after inoculation to fruit
It is being obtained after being counted with SPSS24 as a result, from Table 2, it can be seen that MS, 1/2MS, B5 three distinguish not after investigation
Obviously, B5,1/2MS, N6, WPM difference are equally also unobvious and in subset 2, but unobvious do not represent is not distinguished, from number
As can be seen that WPM is highest in all culture mediums in value, therefore from the point of view of optimization, WPM culture medium is than other
Culture medium effect is more preferable.) more significant than other culture mediums.
Table 2
Influence of the different culture medium of table 3 to callus effect is formed
As a result
DuncanA, b
The class mean in similar subset has been displayed.
Based on the mean value observed.
Error term is mean value side (mistake)=.005.
A. harmomic mean sample size=16.000 are used.
B.Alpha=.05.
5.3 obtain and spend more strain wood most suitable 6-BA concentration in callus forming process
It is equally analyzed by this following data, we can be found that the 6-BA of four kinds of concentration to the callus for spending more strain wood stem
The influence difference organized the formation of is smaller, but is cured compared to the 6-BA of other concentration, the 6-BA of 2mg/L to strain wood stem is spent more
The influence that injured tissue is formed is more significant.Know that the 6-BA of 2mg/L is best to telling on for the callus for spending more strain wood stem.
Influence of the 6-BA of 4 various concentration of table to callus effect is formed
As a result
DuncanA, b
The class mean in similar subset has been displayed.
Based on the mean value observed.
Error term is mean value side (mistake)=.005.
A. harmomic mean sample size=20.000 are used.
B.Alpha=.05.
5.4 obtain and spend more strain wood most suitable NAA concentration in callus forming process
It is analyzed in the same old way by this following data, we can be found that NAA that concentration is 0.2mg/L and 2mg/L to spending more
The influence that the callus of the wooden stem of strain is formed is more significant relative to the NAA culture medium of other concentration, and wherein concentration is 0.2mg/L
NAA to spend more strain wood stem callus formed influence be better than 2mg/L NAA, it may thus be appreciated that the NAA of 0.2mg/L is to more
Telling on for the callus of flower strain wood stem is best.
Influence of the NAA of 5 various concentration of table to callus effect is formed
As a result
DuncanA, b
The class mean in similar subset has been displayed.
Based on the mean value observed.
Error term is mean value side (mistake)=.005.
A. harmomic mean sample size=20.000 are used.
B.Alpha=.05.
5.5 obtain and spend more strain wood most suitable hormone ratio in callus forming process
6 NAA of table and 6-BA main effect are examined
The inspection of effect between main body
Dependent variable: result
The side a.R=.283 (the adjustment side R=.179)
By table 6, it can be concluded that, hormone NAA is later more most beneficial for promoting the callus for spending more strain wood stem to be formed
Flower strain wood bud differentiation is laid a good foundation.
By comparing above, we may safely draw the conclusion, and ratio is WPM+2mg/L 6-BA+0.2mg/L NAA, sucrose
The culture medium of 35g/L, agar 7g/L are the most advantageous for the calli induction for spending more the stem of strain wood.
To sum up, minimal medium, the 6-BA concentration, NAA concentration of strain wood stem callus formation are spent more in this test to promotion
It is screened, the disinfection treatment mode impregnated with 0.1% mercuric chloride of 5min is impregnated by 75% alcohol of 30s, is tentatively obtained
Most suitable initial culture base is WPM+2mg/L 6-BA+0.2mg/L NAA, sucrose 35g/L, agar 7g/L.
Claims (4)
1. a kind of method that induction flowering dogwood callus is formed characterized by comprising using the stem of flowering dogwood as explant
Body is inoculated on culture medium after disinfection and carries out illumination cultivation；The wherein formula of the culture medium are as follows: basal medium+6-BA
1.5-2mg/L+NAA 0.1-0.2mg/L+ sucrose 30-40g/L+ agar 6-9g/L；
The method of the disinfection includes: after cleaning explant, 70-75% ethanol postincubation 30-50s first to be used, after sterile water wash
It is carried out disinfection processing with 0.1-0.2% mercuric chloride again, the processing time is 3-6min, and finally clean with aseptic water washing, removal is remaining
Moisture；The basal medium is WPM.
2. the method that induction flowering dogwood callus according to claim 1 is formed, which is characterized in that the explant
The preparation method comprises the following steps: acquisition flowering dogwood current-year branch tender stem, shearing growth 1-3cm stem section.
3. the method that induction flowering dogwood callus according to claim 1 is formed, which is characterized in that the illumination training
Feeding temperature is 23-27 DEG C.
4. the method that induction flowering dogwood callus according to claim 1 is formed, which is characterized in that light application time is
16-17h/d, illuminance 2000-2200lx.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710686445.3A CN107360973B (en) | 2017-08-11 | 2017-08-11 | A method of induction flowering dogwood callus formation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710686445.3A CN107360973B (en) | 2017-08-11 | 2017-08-11 | A method of induction flowering dogwood callus formation |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107360973A CN107360973A (en) | 2017-11-21 |
| CN107360973B true CN107360973B (en) | 2019-09-10 |
Family
ID=60310146
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710686445.3A Active CN107360973B (en) | 2017-08-11 | 2017-08-11 | A method of induction flowering dogwood callus formation |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107360973B (en) |
-
2017
- 2017-08-11 CN CN201710686445.3A patent/CN107360973B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN107360973A (en) | 2017-11-21 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103931492B (en) | The tissue culture fast seedling-cultivating method of apple rootstock M9 | |
| CN104099287B (en) | The acquisition of high yield OPC Vitis davidii Foex callus and subculture keeping method | |
| CN102232360B (en) | Method for establishing in-vitro rapid propagation system of anubias barteri var. barteri, anubias barteri var. nana, ficus henryi and anubias hastifolia | |
| CN102640705B (en) | Fast propagation method for aquatic plant lotus flowers | |
| CN105474992A (en) | Chinese chestnut summer truffle mycorrhizal seedling culture method based on symbiotic relation | |
| CN104855294B (en) | A kind of Caulis Akebiae rapid propagation method | |
| CN104686361B (en) | The induction of a kind of Fructus Vitis viniferae embryo callus and cultural method | |
| CN107155875B (en) | A kind of allopolyploid induction production method of hybrid Chinese pennisetum | |
| CN100394845C (en) | In-bottle production method of detoxified small seed ball of east lily | |
| CN105475067A (en) | Chinese chestnut tuber indicum mycorrhizal seedling culture method based on symbiotic relation | |
| CN106069772A (en) | A kind of sword-leaved cymbidium tissue culture quick propagation culturing method | |
| CN105766655A (en) | Establishing method of Gleditsia vestita Chun et Howex B.G.Li.tissue culture regeneration system | |
| CN107360973B (en) | A method of induction flowering dogwood callus formation | |
| CN105104201B (en) | A kind of Sa Wanabai primary tissue culture method | |
| CN107466863B (en) | A kind of Beijing Sorbus alnifloria method for tissue culture | |
| CN105918132A (en) | Rapid breeding method of clerodendrum trichotomum thunb | |
| CN109329059A (en) | A kind of method of straw short-tube lycoris tissue cultures | |
| CN107372114B (en) | A kind of abductive approach of the callus of flowering dogwood bud origin | |
| CN108633742A (en) | A kind of China fir Stem tip induction culture medium and abductive approach | |
| CN101268744B (en) | Method for innoculating soybean with phytophthora sojae | |
| CN107683771A (en) | A kind of method of the efficient sterile culture of tree peony embryo | |
| CN107494270B (en) | The tissue culture and rapid propagation method of gold leaf common bluebeard | |
| CN110226514A (en) | The method for obtaining salt-tolerant mutant based on ARTP breeding instrument mutagenesis alfalfa | |
| CN109997692A (en) | Blue or green money willow callus induction and subculture multiplication medium and its cultural method | |
| CN110432150A (en) | A kind of method of the acquisition and fast breeding of lamiophlomis rotata aseptic seedling |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |