CN107711514A - A kind of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method - Google Patents
A kind of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method Download PDFInfo
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- CN107711514A CN107711514A CN201711192038.3A CN201711192038A CN107711514A CN 107711514 A CN107711514 A CN 107711514A CN 201711192038 A CN201711192038 A CN 201711192038A CN 107711514 A CN107711514 A CN 107711514A
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- sharon
- rose
- nonirrigated farmland
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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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- 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 invention discloses a kind of nonirrigated farmland rose of Sharon Hibiscus aridicola Anthony purple flowers system fine individual plant tissue culture and rapid propagation method.Using nonirrigated farmland rose of Sharon purple flower system fine individual plant as explant, sterilized, induced, broken up, breed and taken root a series of incubation steps by explant, efficiently solve nonirrigated farmland rose of Sharon tissue-culturing rapid propagation, introduction and acclimatization and its commercial utilization bottleneck problem, natural plant amount caused by effectively avoiding wild resource excessive use simultaneously is reduced to wreck with natural vegetation area, particularly has positive role in terms of the stability of fine individual plant special shape is kept.Exploitation prospect is fabulous.This method induces differentiation rate 80%, 30 days breeding cycles, growth coefficient 4; rooting rate is 95%; transplanting survival rate is 92%, drastically increases the reproductive number and growth rate of the nonirrigated farmland rose of Sharon, and technical support has been supplied for the introduction and acclimatization of the species, preservation and large-scale production.
Description
Technical field
The present invention relates to plant tissue culture quick-breeding method in biotechnology, relates in particular to excellent to nonirrigated farmland rose of Sharon purple flower system
Good individual plant tissue culture and rapid propagation method.
Background technology
Nonirrigated farmland rose of Sharon Hibiscus aridicola Anthony are under the jurisdiction of Malvaceae Malvaceae Hibiscuses, beautiful
More flowering shrubs.The nonirrigated farmland rose of Sharon is the xeothermic dry-warm valley endemic species in Jinsha jiang River, is only distributed in Lijiang, yunnan and Yanbian, Sichuan Province area, sea
In the xeothermic dry-warm valley clump for lifting 600-2500 rice.Pattern is relatively conventional with white, and purple flower and pink colour flower are extremely rare.Together
When due to NATURAL DISTRIBUTION point be less than 5, population continuous downturn, be put within 2004《Chinese species Red List》Species in imminent danger
[EN B2ab(ii)].In recent years, because construction of reservoir dam, habitat are progressively lost, frequently influenceed plus by mankind's activity, drought
The living environment of the ground rose of Sharon is by more serious threat.
At present, the sterile quick breeding in biotechnology has turned into the kind such as Chinese medicine, flowers, endangered species, economic fruit
The important means of seedling production.
So far, prior art is not on nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation and related biology
The report of technical elements.
The content of the invention
It is an object of the invention to provide the method for nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation, the nonirrigated farmland rose of Sharon is filled up
Blank in biotechnology, while the problem of solve the nonirrigated farmland rose of Sharon distributed areas are narrow in the wild, and introduction and acclimatization difficulty is big.This hair
It is bright that for the nonirrigated farmland rose of Sharon, comprehensively seedling breeding basis has been established in exploitation and sustainable utilization, the preservation of purple flower genotype and expansion.
In order to realize the purpose of the present invention, the invention provides following technical scheme：
A kind of method of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation, this method include explant and selected with disappearing
Poison, induction and differentiation culture, propagation and squamous subculture, strong sprout and culture of rootage, bottle seedling transplant step：
The explant selection and sterilization are to take the tender terminal bud of nonirrigated farmland rose of Sharon purple flower system fine individual plant and lateral bud, are used
1% suds are soaked 10 minutes, and explant sterilization is carried out after flowing water is rinsed well；
Described induce with differentiation culture is that the explant after sterilization is being induced and cultivated in differential medium, is induced with dividing
Change culture medium as MS culture medium+1mg/L 6-BA+0.5mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8, cultivation cycle 20
My god；
The propagation is that induction is being bred and cultivated in subculture medium with changing the material after cultivating with squamous subculture, is bred
It is MS culture medium+0.8mg/L 6-BA+0.5mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8 with subculture medium, cultivates
30 days cycles；
The strong sprout is by breeding with the seedling of squamous subculture in strong sprout with being cultivated in root media, strengthening with culture of rootage
Seedling is MS culture medium+0.3mg/LNAA+0.3mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8 with root media, is cultivated
20 days cycles；
The bottle seedling is transplanted：Bottle seedling of taking root after taking root carries the last week and is placed in greenhouse hardening, prepares matrix, matrix
For the perlite of volume ratio: fertile soil: raw laterite=1: 2: 3, spray mixing soil with 800 times of carbendazim, use plastic film seal
Sterilization 7 days, pH to 5.8 is adjusted, open seedling taking, gently clean base portion culture medium, transplanted, in time water spray and plastic covering film, greatly
Canopy temperature is 20~30 DEG C, air humidity 50~60%, soil humidity 75~85%.
According to described nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, purple flower system fine individual plant children
As explant, the illumination condition of induction and differentiation, propagation and subculture, strong sprout and culture of rootage is people for tender terminal bud and lateral bud
The auxiliary smooth 1500LUX of work, temperature are 23~30 DEG C.
According to described nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, the propagation is entered in a manner of nodal bud
Row breeding, growth is rapid, and growing height reaches 5.6cm within 30 days, average breeding rate 4.
According to the method for described nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation, induction closes two with differential medium
For one, propagation is combined into one with subculture medium, and strong sprout is combined into one with root media, cultural method simplification.
According to nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, bottle seedling of taking root transplanting is preposition to refine in greenhouse
Transplanting is in ready matrix after seedling adapts to one week.
According to nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, transplanting medium is：The pearl of volume ratio
Rock: fertile soil: raw laterite=1: 2: 3, spray mixing soil with 800 times of carbendazim, sterilized 7 days with plastic film seal, adjust pH
To 5.8, seedling taking is opened, gently cleans base portion culture medium, transplanting, water spray and plastic covering film, greenhouse temperature are 20~30 in time
DEG C, shading rate 75~85%, air humidity 50~60%, soil humidity survival rate 92% after 75~85%, 30 days.
The young tender terminal bud and lateral bud for taking nonirrigated farmland rose of Sharon purple flower system fine individual plant are explant, soak 10 with 1% suds
Minute, after flowing water is rinsed well, take on super-clean bench, defoliation, the small stem section of belt segment of 1cm sizes is cut into, with 75% alcohol
Sterilize 10s, after aseptic water washing 3 times, then with 0.1% mercuric chloride solution sterilize 8min, sterile water wash 3~5 times, be inoculated in preparation
In good inducing culture, every bottle is inserted 1 bud.
The preparation of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation culture medium：Induction is cultivated with differential medium for MS
Base+0.8mg/L 6-BA (6- benzyl purines)+0.5mg/L NAA (naa)+sucrose 30g/L+ agar 5g/L, pH5.8, illumination
Intensity is 1000LUX, daily illumination 10 hours, temperature is 23~30 DEG C.After inducing one week, terminal bud or lateral bud start to sprout.
Propagation with subculture medium be MS culture medium+0.8mg/L 6-BA (6- benzyl purines)+0.5mg/L NAA (how second
Acid)+sucrose 30g/L+ agar 5g/L, pH5.8, cultivation cycle 30 days.Luminous intensity is 1500LUX, and temperature is 23~30 DEG C.
Strong sprout and root media be MS culture mediums+0.5mg/LNAA (naa)+0.5mg/L IAA (heteroauxin)+
Sugared 30g/L+ agar 5g/L, pH5.8, luminous intensity 1500LUX, temperature are 23~30 DEG C, cultivation cycle 20 days.
Bottle seedling is transplanted：Bottle seedling of taking root after cultivating 20 days, it is 75%~85% greenhouse hardening one that need to move to shading rate in advance
Week.Matrix is perlite：Fertile soil：Raw laterite (volume ratio)=1：2：3, spray mixing soil with 800 times of carbendazim, use plastics
Film sealing sterilization 7 days, adjusts pH to 5.8.Seedling is gently taken out in corkage, cleans the culture medium of root, transplanting to the base sterilized
In matter, water spray and plastic covering film moisturizing in time.Greenhouse temperature is 20~30 DEG C, and obscurity is 75%~85%, air humidity
50~60%, soil humidity 75~85%.Survival rate 92% after 30 days.
The it is proposed of technical solution of the present invention is based on following Research foundations：
Nonirrigated farmland rose of Sharon distributed areas are narrow, are the xeothermic dry-warm valley endemic species in Jinsha jiang River, population continuous downturn, manually draw
Kind domestication difficulty is big.Its purple flower genotype is even more rare.It is few to solve its natural resources, reach in the short time amount reproduction,
Preservation and sustainable utilization, fill up research blank of the nonirrigated farmland rose of Sharon in biotechnology.So nonirrigated farmland rose of Sharon purple flower system is invented
Fine individual plant group culturation rapid propagating technology, for the nonirrigated farmland rose of Sharon, comprehensively exploitation and sustainable use are laid a good foundation.
Compared with prior art, beneficial effects of the present invention are：
1. the present invention establishes effective nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, solves its distribution
It is narrow, introduction and acclimatization is difficult, purple flower genotype is rare, filled up research and development blank of the nonirrigated farmland rose of Sharon in biotechnology.
2. the present invention is by nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method, seedling is easy, propagation steps letter
Change, effective proliferative speed is high, significant to preserve and expanding nonirrigated farmland rose of Sharon population.
3. the present invention highly maintains nonirrigated farmland aniline violet by nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method
The genetic stability and uniformity of dyeing defect system fine individual plant, for the nonirrigated farmland rose of Sharon, comprehensively exploitation and sustainable utilization are laid a good foundation.
4. the nonirrigated farmland rose of Sharon that the nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture and rapid propagation method of the present invention is bred, induce within 20 days
Differentiation rate 87%, it is 4 in 30 days internal breeding coefficients, rooting rate 95%, transplanting survival rate 92%, drastically increases nonirrigated farmland
The breeding coefficient of the rose of Sharon, it is non-using providing for the performance of the introduction and acclimatization of the species, preservation, gardens gardening and its commercial value
Normal effective propagation method.
Brief description of the drawings
Fig. 1 is nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue culture propagation situation of the present invention.
Fig. 2 is that nonirrigated farmland rose of Sharon purple flower system of the present invention fine individual plant tissue culture is taken root situation.
Embodiment
Below in conjunction with the accompanying drawings, the technical characterstic of the present invention and substance is expanded on further with the specific embodiment of the present invention
Content, but the present invention is not limited with this.
Embodiment 1
1. culture medium and Screening matrix：
The young tender terminal bud and lateral bud for taking nonirrigated farmland rose of Sharon purple flower system fine individual plant are explant, soak 10 with 1% suds
Minute, after flowing water is rinsed well, take on super-clean bench, be cut into the stem section with single section, with 75% alcohol disinfecting 10s, nothing
After bacterium water rinses 3 times, surface sterilization 5min is carried out with 0.1% mercuric chloride solution, sterile water wash 3~5 times, is inoculated in ready
In the rose of Sharon inducing culture of nonirrigated farmland, every bottle of 1 section.
Stem section is seeded in MS+0.1mg/l 6-BA+0.5mg/L NAA, MS+0.5mg/l 6-BA+0.5mg/L respectively
NAA, MS+0.8mg/l 6-BA+0.5mg/LNAA, MS+1mg/l 6-BA+0.5mg/LNAA, MS+2mg/l 6-BA+0.5mg/L
On NAA culture mediums, sucrose 30g/L, agar 5g/L, pH5.8.Luminous intensity is 1000LUX, and temperature is 23~30 DEG C, light application time
10 hours.Count inductivity situation such as following table：
The fine individual plant stem section inducing culture screening of 1 nonirrigated farmland rose of Sharon purple flower system of table
Nonirrigated farmland rose of Sharon stem section inducing culture is screened, the results showed that, in MS+0.8mg/l 6-BA+0.5mg/
On LNAA culture mediums, inductivity reaches 86%.
The subculture medium hormone of table 2 screens
Using MS culture mediums as propagation and the minimal medium of subculture hormone screening, the basic element of cell division is 6-BA, concentration model
Enclose for (0.1mg/L, 0.5mg/L, 1mg/L, 2mg/L, 4 concentration gradients), auxin concentration be respectively (0mg/L, 0.1mg/L,
0.5mg/L, 1mg/L, 4 concentration gradients), carried out using uniform design, it is as a result as follows：
MS culture medium+0.5mg/L 6-BA (6- benzyl purines)+0.5mg/L NAA (naa)+sucrose 30g/L+ agar
5g/L, pH5.8, cultivation cycle 30 days, luminous intensity 1500LUX, temperature are 23~30 DEG C.Bud joint number amount (including terminal bud) 4~
5, appreciation rate 4~5, plant leaf stretching, extension, the speed of growth is most fast, 5~6cm of plant height, the thick 2mm-3mm of stem.
The root media hormone the selection result of table 3
MS culture mediums+0.5mg/LNAA (naa)+0.5mg/L IAA (heteroauxin)+sucrose 30g/L+ agar 5g/L,
PH5.8, cultivation cycle 20 days, rooting rate is up to 95%.
The screening experiment result of the cultivation matrix of table 4
Three kinds of matrix coordinate in experiment 4., add matrix gas porosity and root gas permeability, are also provided enough for root
Nutrition, while laterite is soil layer deep soil, content of molds is less, is advantageous to the growth of aseptic seedling.Transplanting survival rate reaches after 30 days
To 95%.
Embodiment form above is explained in detail to the above of the present invention, but this should not be interpreted as to the present invention
The scope of above-mentioned theme is only limitted to the embodiment of the above, and all technologies realized based on the above of the present invention belong to the present invention
Protection domain.
Claims (7)
1. the method for a kind of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation, it is characterised in that this method is selected including explant
Select and sterilize, induce with breaking up culture, propagation and squamous subculture, strong sprout and culture of rootage, bottle seedling transplant step,
The explant selection and sterilization are to take the tender terminal bud of nonirrigated farmland rose of Sharon purple flower system fine individual plant and lateral bud, with 1% fertilizer
Soap water is soaked 10 minutes, and explant sterilization is carried out after flowing water is rinsed well；
The induction with differentiation culture is induced and trained with differentiation with being cultivated in differential medium in induction by the explant after sterilization
Base is supported as MS culture medium+1mg/L 6-BA+0.5mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8, cultivation cycle 20 days；
It is described propagation with squamous subculture be induction with differentiation culture after material breed and cultivated in subculture medium, breed and
Subculture medium is MS culture medium+0.8mg/L 6-BA+0.5mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8, and culture is all
30 days phases；
The strong sprout and culture of rootage be by breeding with the seedling of squamous subculture in strong sprout with being cultivated in root media, strong sprout and
Root media is MS culture medium+0.3mg/LNAA+0.3mg/L IAA+ sucrose 30g/L+ agar 5g/L, pH5.8, cultivation cycle
20 days；
The bottle seedling is transplanted：Bottle seedling of taking root after taking root carries the last week and is placed in greenhouse hardening, prepares matrix, matrix is body
The perlite of product ratio: fertile soil: raw laterite=1: 2: 3, spray mixing soil with 800 times of carbendazim, 7 are sterilized with plastic film seal
My god, pH to 5.8 is adjusted, opens seedling taking, cleans base portion culture medium, transplanting, timely water spray and plastic covering film, greenhouse temperature 20
~30 DEG C, air humidity 50~60%, soil humidity 75~85%.
2. nonirrigated farmland rose of Sharon purple flower system according to claim 1 fine individual plant tissue culture and rapid propagation method, it is characterised in that described
Induction and differentiation, propagation and the illumination condition of subculture, strong sprout and culture of rootage are artificial auxiliary smooth 1500LUX, and temperature is 23~
30℃。
3. nonirrigated farmland rose of Sharon purple flower system according to claim 1 fine individual plant tissue culture and rapid propagation method, it is characterised in that described
Propagation is bred in a manner of nodal bud, and growth is rapid, and growing height reaches 5.6cm within 30 days, and average breeding rate is 4.
4. the method for nonirrigated farmland rose of Sharon purple flower system according to claim 1 fine individual plant tissue-culturing rapid propagation, it is characterised in that institute
Induction to be stated to be combined into one with differential medium, propagation is combined into one with subculture medium, and strong sprout is combined into one with root media,
Cultural method simplifies.
5. nonirrigated farmland rose of Sharon purple flower system according to claim 1 fine individual plant tissue culture and rapid propagation method, it is characterised in that take root
The preposition transplanting after greenhouse hardening adapts to one week of bottle seedling transplanting is in ready matrix.
6. nonirrigated farmland rose of Sharon purple flower system according to claim 1 fine individual plant tissue culture and rapid propagation method, it is characterised in that transplanting
Matrix is：The perlite of volume ratio: fertile soil: raw laterite=1: 2: 3, spray mixing soil with 800 times of carbendazim, use plastic foil
Sealing sterilization 7 days, adjusts pH to 5.8, opens seedling taking, gently clean base portion culture medium, transplanting, in time water spray and plastic covering
Film, greenhouse temperature are 20~30 DEG C, and shading rate 75~85%, air humidity 50~60%, soil humidity is after 75~85%, 30 days
Survival rate 92%.
7. the method for a kind of nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation, it is characterised in that this method includes：Take nonirrigated farmland
The young tender terminal bud and lateral bud of rose of Sharon purple flower system fine individual plant are explant, are soaked 10 minutes with 1% suds, are rinsed through flowing water
After clean, take on super-clean bench, defoliation, be cut into the small stem section of belt segment of 1cm sizes, with 75% alcohol disinfecting 10s, sterilized water punching
After washing 3 times, then with 0.1% mercuric chloride solution sterilize 8min, sterile water wash 3~5 times, be inoculated in ready inducing culture
In, every bottle is inserted 1 bud；
Nonirrigated farmland rose of Sharon purple flower system fine individual plant tissue-culturing rapid propagation culture medium is prepared, induced and broken up, bred and subculture, strong sprout
With taking root, bottle seedling transplanting：
Induction and differential medium are MS culture medium+0.8mg/L 6-BA+0.5mg/L NAA+ sucrose 30g/L+ agar 5g/L,
PH5.8, intensity of illumination 1000LUX, daily illumination 10 hours, temperature is 23~30 DEG C, after inducing one week, terminal bud or lateral bud
Start to sprout；
Propagation and subculture medium are MS culture medium+0.8mg/L 6-BA+0.5mg/L NAA+ sucrose 30g/L+ agar 5g/L,
PH5.8, cultivation cycle 30 days, luminous intensity 1500LUX, temperature are 23~30 DEG C；
Strong sprout and root media are MS culture medium+0.5mg/LNAA+0.5mg/L IAA+ sugar 30g/L+ agar 5g/L, pH5.8,
Luminous intensity is 1500LUX, and temperature is 23~30 DEG C, cultivation cycle 20 days；
Bottle seedling is transplanted：Bottle seedling of taking root after cultivating 20 days, shading rate need to be moved in advance and is 75%~85% greenhouse hardening one week, base
Matter is the perlite of volume ratio: fertile soil: raw laterite=1: 2: 3, spray mixing soil with 800 times of carbendazim, and it is close with plastic foil
Envelope sterilization 7 days, pH to 5.8 being adjusted, seedling is gently taken out in corkage, cleans the culture medium of root, transplants into the matrix sterilized,
Water spray and plastic covering film moisturizing in time, greenhouse temperature are 20~30 DEG C, obscurity 75%~85%, air humidity 50~
60%, soil humidity survival rate 92% after 75~85%, 30 days.
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| CN201711192038.3A CN107711514B (en) | 2017-11-24 | 2017-11-24 | Tissue culture and rapid propagation method for excellent individual plant of hibiscus purpureus in dry land |
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| CN201711192038.3A CN107711514B (en) | 2017-11-24 | 2017-11-24 | Tissue culture and rapid propagation method for excellent individual plant of hibiscus purpureus in dry land |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110663552A (en) * | 2019-11-08 | 2020-01-10 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method of Yunnan tung tree |
| CN112243861A (en) * | 2020-10-27 | 2021-01-22 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method for Huagaimu |
| CN112243860A (en) * | 2020-10-27 | 2021-01-22 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method for Chinese parasol trees |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110663552A (en) * | 2019-11-08 | 2020-01-10 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method of Yunnan tung tree |
| CN112243861A (en) * | 2020-10-27 | 2021-01-22 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method for Huagaimu |
| CN112243860A (en) * | 2020-10-27 | 2021-01-22 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method for Chinese parasol trees |
| CN112243861B (en) * | 2020-10-27 | 2021-04-09 | 中国科学院昆明植物研究所 | Tissue culture and rapid propagation method for Huagaimu |
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| Publication number | Publication date |
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| CN107711514B (en) | 2020-02-21 |
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