CN106376461B - The method of China fir tissue-cultured seedling culture of rootage and hardening - Google Patents
The method of China fir tissue-cultured seedling culture of rootage and hardening Download PDFInfo
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- CN106376461B CN106376461B CN201610755011.XA CN201610755011A CN106376461B CN 106376461 B CN106376461 B CN 106376461B CN 201610755011 A CN201610755011 A CN 201610755011A CN 106376461 B CN106376461 B CN 106376461B
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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
- A01H4/008—Methods for regeneration to complete plants
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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
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Abstract
The present invention relates to the method for a kind of China fir tissue-cultured seedling culture of rootage and hardening, this method includes the steps that selecting Tube plantlets, inoculation of taking root, culture of rootage, seal cap hardening of taking root and hardening of uncapping.This method will take root to be combined with hardening, in culturing room after short-term culture of rootage, carries out take root seal cap, hardening of uncapping culture, and tissue-cultured seedling is made to hardening on one side take root while.This method can significantly shorten the time of China fir tissue culture seedling rooting and hardening, to make, growing-seedling period shortening more significant than conventional method, root long is suitable for, nursery effect significantly improves, and pass through nature temperature control light-control technology energy-saving branch, it can effectively solve the problems, such as Gao Chengben, low transplanting survival rate in existing China fir tissue-cultured seedling production process, the foundation for facilitating excellent Chinese fir clone nursery base, provides high quality seedling for production of forestry.
Description
Technical field
The present invention relates to agricultural technology fields, more particularly to the method for a kind of China fir tissue-cultured seedling culture of rootage and hardening.
Background technique
China fir (Cunninghamia Lanceolata) is Taxodiaceae (Taxodiaceae), Cunninghamia aiphyllium tree species,
Growth is fast, material is good, is important one of the fast-growing commodity commerical tree species of south China.Plant Tissue Breeding is quick breeding plant
The important method of new varieties can largely breed high quality seedling for life while holding kind excellent hereditary capacity in a short time
Application is produced, the improved variety process of Industry plantation has been greatly facilitated.China Fir Study on tissue culture last decade is quickly grown,
China fir tissue culture production line is established on Fujian, Guangdong and other places, but the generally existing high production cost of the prior art, nursery effect are low etc. asks
Topic makees support there has been no the method for tissue culture of high-efficiency and economic and establishes high quality seedling production base, in addition existing breeding garden yield
Deficiency, leading to Cunninghamia Lanceolata Plantations breeding nursery stock, supply falls short of demand, and production is very different with kind of hereditary quality, how economical, quickly numerous
It is extremely urgent to educate breeding of new generation.The seedling-wood breeding period is carried out by the existing tissue culture seedling rooting of China fir and hardening technology merely
Length, transplanting survival rate are low, have no relevant shortening China fir tissue culture seedling rooting and the method for hardening time report.
Summary of the invention
Based on this, the present invention provides the method for a kind of China fir tissue-cultured seedling culture of rootage and hardening, this method can shorten
The time of China fir tissue culture seedling rooting and hardening shortens the seedling-wood breeding period, and the transplanting survival rate of nursery stock can be improved.
Specific technical solution is as follows.
A kind of method of China fir tissue-cultured seedling culture of rootage and hardening, comprising the following steps:
(1) select Tube plantlets: selection length is more than the tissue culture bud of 3cm in the China fir Tube plantlets for carrying out Multiplying culture
Material of the seedling as culture of rootage；
(2) take root inoculation: the part of the top tip of Tube plantlets described in clip long 2.5cm~3.0cm down, insertion is equipped with life
In the culture bottle of root culture medium, seal；
(3) culture of rootage: inoculated Tube plantlets are subjected to culture of rootage, cultivation temperature is 25 ± 5 DEG C, relative humidity
Be 50%~60%, carried out again after dark culture 3~5 days optical culture 15~20 days, the intensity of illumination of optical culture be 1500Lx~
3000Lx, light application time are daily 12h~16h；
(4) take root seal cap hardening: the nursery stock being spread in a single layer after step (3) culture of rootage has every bottle seedling wood
Natural light irradiation carries out capping hardening 10~15 days, and nursery stock is made to hardening on one side take root while, and the temperature for covering hardening is 20~35
℃；
(5) it uncaps hardening: when capping hardening makes the root system of nursery stock grow to length 0.5cm~1.0cm, opening culture bottle
Lid, it is more than media surface 0.5cm~1.0cm that water surface elevation is filled into culture bottle, uncap hardening 2~5 days, opens
The temperature of lid hardening is 20~35 DEG C, and illumination condition is natural lighting.
In wherein some embodiments, the time of step (2) inoculation of taking root is August~next year 2 months.
In wherein some embodiments, the root media is to contain 0.5~0.8mg/L methyl α-naphthyl acetate, 0.1~0.2mg/
The 1/2MS culture medium of L indolebutyric acid, 24~26g/L sucrose and 6~8g/L agar.
In wherein some embodiments, the volume of the culture bottle is 220~260mL, a height of 85~95mm, bore 60
~64mm is inoculated with 12~15 plants of Tube plantlets in each culture bottle.
In wherein some embodiments, the culture of rootage carries out in Sterile culture room, daily with ultraviolet lamp to sterile
Culturing room sterilizes 2~3 times, 25~35min every time.
In wherein some embodiments, the capping hardening and uncap hardening in vinyl house or in glass room into
Row.
In wherein some embodiments, when the hardening time is 9~November, 70%~75% shading net is covered, and beat
Door window makes cross-ventilation, by the control of hardening temperature between 30 ± 5 DEG C；When the hardening time is December~next year 2 months, removal
Shading net, and close the doors and windows, by the control of hardening temperature between 25 ± 5 DEG C.
In wherein some embodiments, in the experienced seedling step of uncapping, the time for opening culture bottle cover is 4:00 in afternoon
After.
In wherein some embodiments, the method for the China fir tissue-cultured seedling culture of rootage and hardening further includes transplantation of seedlings out
Step: the nursery stock that takes root after hardening of uncapping is taken out from culture bottle, clean up after the culture medium of nursery stock base portion in the same day into
Row transplanting；The matrix of transplanting is yellow soil.
In wherein some embodiments, after the time of fine day transplanting is 4:00 in afternoon, the time of transplanting rainy days is complete
It.
The method of China fir tissue-cultured seedling culture of rootage and hardening of the invention has the following advantages and beneficial effects:
The present inventor has determined the side of a kind of China fir tissue-cultured seedling culture of rootage and hardening by lot of experiments
Method, this method operation is fairly simple, combines taking root with hardening, and first carrying out short-term culture of rootage, to wait for that nursery stock base portion has been formed swollen
When the root system of swollen callus or small particles sample of just emerging, that is, seal cap hardening culture of taking root is carried out, makes tissue-cultured seedling on one side
It takes root one side hardening；Dark culture is combined with optical culture when culture of rootage, and capping hardening is combined with hardening of uncapping when hardening；And
Each parameter in culture of rootage and hardening step is further rationally controlled, this method is allow significantly to shorten China fir tissue culture seedling rooting
It can effectively be solved existing with the time of hardening so that growing-seedling period shortening more significant than conventional method, nursery effect be made to significantly improve
China fir tissue-cultured seedling production process in Gao Chengben, low transplanting survival rate the problem of, facilitate excellent Chinese fir clone nursery base
Foundation, provide high quality seedling for production of forestry.
In addition, this method uses nature temperature control control light measure to hardening environment, that is, natural lighting and air humidity are utilized,
Be spread in a single layer nursery stock, and every bottle seedling wood is made to have uniform natural light irradiation, and 75% shading net is covered on vinyl house top, and beats
Door window keeps cross-ventilation to control temperature, more energy saving for more existing use air-conditioning and light temperature control control light
Section props up, is environmentally protective.
Specific embodiment
It is carried out below in conjunction with method of the specific embodiment to China fir tissue-cultured seedling culture of rootage and hardening of the invention further
Detailed description.
Embodiment 1
6 Clones of Cunninghamia Lanceolata are cultivated in Guangdong Academy of Forestry, the nursery stock that takes root amounts to 6,000 plants, transplants after hardening
5,000 plants of nursery stock.The method of China fir tissue-cultured seedling culture of rootage and hardening that the present embodiment uses is specific as follows:
(1) selection is used for the Tube plantlets of culture of rootage: on September 15th, 2015, in the China fir tissue culture for carrying out Multiplying culture
In sprout, robust growth is selected, the sprout that leaf color is dark green, length is more than 3cm carries out culture of rootage；
(2) take root inoculation: on superclean bench, top tip of the clip for the Tube plantlets of culture of rootage is grown down
The part of 2.5cm~3.0cm, insertion has been got ready in the glass culture bottle of root media in time, every bottle of inoculation nursery stock 12~15
Strain, covered plastic sealing cover, the volume of culture bottle is 240mL, a height of 90mm, bore 62mm.The group of root media becomes
1/2MS+0.5~0.8mg/L methyl α-naphthyl acetate (NAA)+0.1~0.2mg/L indolebutyric acid (IBA)+25g/L sucrose+7g/L agar；
(3) culture of rootage: on September 17th, 2015, the culture bottle for being inoculated with Tube plantlets be put into Sterile culture room into
Row culture of rootage, for the control of culture room temperature at 25 ± 2 DEG C, relative humidity control keeps indoor environment cleaning 50%~60%,
And After Hours opening ultraviolet lamp is sterile to culturing room disinfection 30min respectively in daily noon and evening staff；First to group
The dark culture that sprout carries out 5 days is trained, then on September 22nd, 2015 optical cultures for starting to carry out 15 days by a definite date, the light of optical culture
It is 3000Lx according to intensity, light application time is daily 16h, when nursery stock base portion has formed the callus of swelling or just emerges little Bai
When the root system of point sample, seal cap hardening culture of taking root can be carried out；
(4) it takes root seal cap hardening: on October 7th, 2015 nursery stock after culture of rootage being moved to modeling from Sterile culture room
Seal cap hardening treatment of taking root is carried out in material greenhouse, using natural lighting and air humidity, be spread in a single layer nursery stock, makes every bottle seedling wood
There is natural light irradiation；To prevent temperature of shed excessively high, 75% shading net is covered on vinyl house top, and opens door and window guarantor
Cross-ventilation is held, temperature is controlled in (30 ± 5) DEG C；
(5) it uncaps hardening: when the long 0.5cm~1.0cm of seedling root, after 4:00 in afternoon, opening on October 27th, 2015
The lid of culture bottle injects tap water, and the water surface is more than media surface 0.5cm~1.0cm, carries out hardening of uncapping, hardening of uncapping
Temperature and the same step of humid control (4)；
(6) go out transplantation of seedlings: on October 30th, 2015, at the cloudy day, the nursery stock that takes root after hardening of uncapping taken out from culture bottle,
It is transplanted after cleaning up the culture medium of nursery stock base portion, transplanting medium is yellow soil, and average transplanting survival rate is 82%.This reality
Applying take root hardening situation and its effect of example, see Table 1 for details.
Comparative example 1
By taking clone C7 as an example, this comparative example use tradition take root with hardening off method the following steps are included:
(1) in the Clones of Cunninghamia Lanceolata Tube plantlets for carrying out Multiplying culture, selection robust growth, leaf color is dark green, length is super
The sprout for crossing 3cm carries out culture of rootage；
(2) with the step (2) in embodiment 1；
(3) culture bottle for being inoculated with Tube plantlets is put into Sterile culture room and carries out culture of rootage, cultivate room temperature control
At 25 ± 2 DEG C, relative humidity is controlled 50%~60% system, is carried out 5 days dark cultures to rooted seedling first, is then scheduled to last
30 days optical cultures, intensity of illumination 3000Lx, the daily 16h of light application time can be carried out as the long 0.5cm or more of seedling root
Hardening；
(4) nursery stock to have taken root is moved to vinyl house inner cover hardening 15 days from Sterile culture room, seedling root is long
More than 1.5cm；
(5) after Yu Qingtian 4:00 in afternoon or cloudy, the nursery stock that takes root after hardening is taken out from culture bottle, cleans up seedling
It is transplanted after the culture medium of the wooden base portion, transplanting medium is yellow soil, average transplanting survival rate 50%.The refining of taking root of this comparative example
See Table 1 for details for growth of cereal crop seedlings condition.
The implementation result of the method for 1 China fir tissue-cultured seedling culture of rootage of table and hardening
From 2010, inventor carried out extensive tissue-cultured seedlings of the cunninghamia lanceolata-clone experimental study, and total breeding in 6 years is asexual
It is 10, nearly 1,500,000 plants of tissue-culture container seedling, takes root about 100,000 plants of nursery stock, average rooting rate 89%, transplant survival nursery stock is less than 50,000
Strain, for average transplanting survival rate less than 50%, nursery stock rootage duration is 25~45 days, and the hardening time is 15~20 days, takes root and adds refining
Seedling process time-consuming amounts to 40~65 days, breeds about 2.0 yuan/plant of cost.Finally develop the tissue-cultured seedling culture of rootage of the present embodiment
With the method for hardening, nearly 30,000 plants of China fir tissue culture rooted seedling are cultivated, average rooting rate 92%, transplanting survival rate is improved to 82%, raw
Root adds hardening process to foreshorten to 30~45 days, and seedling cost is down to about 1.5 yuan/plant.This method is in addition to shortening cultivation period, to refining
Seedling environment uses nature temperature control control light measure, that is, utilizes natural lighting and air humidity, and be spread in a single layer nursery stock, makes every bottle seedling wood
Have uniform natural light irradiation, on vinyl house top cover 75% shading net, and open door and window keep cross-ventilation to
Temperature is controlled, compared to more existing using for air-conditioning and light temperature control control light, more energy-saving branch, environmentally protective.By this method
Applied in excellent Chinese fir clonal tissue culture production line, good base has been established for the production and popularization and application of China fir high quality seedling
Plinth.
Each technical characteristic of embodiment described above can be combined arbitrarily, for simplicity of description, not to above-mentioned reality
It applies all possible combination of each technical characteristic in example to be all described, as long as however, the combination of these technical characteristics is not deposited
In contradiction, all should be considered as described in this specification.
The embodiments described above only express several embodiments of the present invention, and the description thereof is more specific and detailed, but simultaneously
It cannot therefore be construed as limiting the scope of the patent.It should be pointed out that coming for those of ordinary skill in the art
It says, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to protection of the invention
Range.Therefore, the scope of protection of the patent of the invention shall be subject to the appended claims.
Claims (6)
1. a kind of method of China fir tissue-cultured seedling culture of rootage and hardening, which comprises the following steps:
(1) select Tube plantlets: selection length is more than that the Tube plantlets of 3cm are made in the China fir Tube plantlets for carrying out Multiplying culture
For the material of culture of rootage；
(2) take root inoculation: the part of the top tip of Tube plantlets described in clip long 2.5cm~3.0cm down, insertion is equipped with training of taking root
In the culture bottle for supporting base, seal；
(3) culture of rootage: inoculated Tube plantlets are subjected to culture of rootage, cultivation temperature is 25 ± 5 DEG C, and relative humidity is
Carried out again after 50%~60%, dark culture 3~5 days optical culture 15~20 days, the intensity of illumination of optical culture be 1500Lx~
3000Lx, light application time are daily 12h~16h；
(4) take root seal cap hardening: the nursery stock being spread in a single layer after step (3) culture of rootage makes every bottle seedling wood have nature
Light irradiation, carries out capping hardening 10~15 days, and nursery stock is made to hardening on one side take root while, and the temperature for covering hardening is 20~35 DEG C；
(5) it uncaps hardening: when capping hardening makes the root system of nursery stock grow to length 0.5cm~1.0cm, opening the lid of culture bottle
Son, it is more than media surface 0.5cm~1.0cm that water surface elevation is filled into culture bottle, uncap hardening 2~5 days, uncaps
The temperature of hardening is 20~35 DEG C, and illumination condition is natural lighting；
The root media is to contain 0.5~0.8mg/L methyl α-naphthyl acetate, 0.1~0.2mg/L indolebutyric acid, 24~26g/L sucrose
With the 1/2MS culture medium of 6~8g/L agar；
The capping hardening and hardening of uncapping carry out in vinyl house or in glass room；
When the hardening time is 9~November, 70%~75% shading net is covered, and open door and window to make cross-ventilation, by hardening
Temperature controls between 30 ± 5 DEG C；When the hardening time is December~next year 2 months, shading net is removed, and close the doors and windows, will refined
Seedling temperature controls between 25 ± 5 DEG C；
Further include the steps that transplantation of seedlings out: the nursery stock that takes root after hardening of uncapping being taken out from culture bottle, cleans up nursery stock base
It was transplanted after the culture medium in portion in the same day；The matrix of transplanting is yellow soil.
2. the method for China fir tissue-cultured seedling culture of rootage and hardening according to claim 1, which is characterized in that step (2) institute
The time for stating inoculation of taking root is August~next year 2 months.
3. the method for China fir tissue-cultured seedling culture of rootage and hardening according to claim 1 or 2, which is characterized in that the training
The volume for supporting bottle is 220~260mL, a height of 85~95mm, bore are 60~64mm, and Tube plantlets 12 are inoculated in each culture bottle
~15 plants.
4. the method for China fir tissue-cultured seedling culture of rootage and hardening according to claim 1 or 2, which is characterized in that the life
Root culture carries out in Sterile culture room, is sterilized 2~3 times with ultraviolet lamp to Sterile culture room daily, 25~35min every time.
5. the method for China fir tissue-cultured seedling culture of rootage and hardening according to claim 1 or 2, which is characterized in that described to open
It covers and practices in seedling step, after the time for opening culture bottle cover is 4:00 in afternoon.
6. the method for China fir tissue-cultured seedling culture of rootage and hardening according to claim 1, which is characterized in that fine day transplanting
After time is 4:00 in afternoon, the time of transplanting rainy days is whole day.
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| CN109348953A (en) * | 2018-10-18 | 2019-02-19 | 南阳师范学院 | A kind of Kiwi berry culture of rootage and acclimatization and transplants method |
| CN113826547A (en) * | 2021-02-01 | 2021-12-24 | 广东省林业科学研究院 | Method for recycling fir tissue culture polluted adventitious buds |
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