CN102349438B - Tissue culture and rapid propagation breeding method for golden elf sundust - Google Patents
Tissue culture and rapid propagation breeding method for golden elf sundust Download PDFInfo
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- CN102349438B CN102349438B CN2011102575480A CN201110257548A CN102349438B CN 102349438 B CN102349438 B CN 102349438B CN 2011102575480 A CN2011102575480 A CN 2011102575480A CN 201110257548 A CN201110257548 A CN 201110257548A CN 102349438 B CN102349438 B CN 102349438B
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Abstract
The invention discloses a tissue culture and rapid propagation breeding method for golden elf sundust. The tissue culture and rapid propagation breeding method comprises the following steps of: with lateral bud and stem tip as primary culture materials, soaking the primary culture materials with a sanmate solution; soaking the mixed solution with a vitamin C distilled water solution; and conventionally sterilizing with 0.1 percent liter by percent of mercury, wherein the culture conditions for primary induction are realized by 1/2MS+sucrose+agar+activated carbon+sodium sulfite+TDZ (Thidiazuron)+NAA(Naphthaleneacetic Acid); the culture conditions for subculture multiplication are realized by 1/2MS+agar+sucrose+activated carbon+sodium sulfite+TDZ+NAA; culture conditions for rooting are realized by 1/2MS+agar+sucrose+activated carbon+sodium sulfite+6-BA(Butyl Acrylate)+NAA; and the transplanting conditions for test-tube plantlets are realized by sterilizing seedlings with aqueous solution of potassium permanganate and transplanting the sterilized seedlings. The invention provides a tissue culture method for the golden elf sundust which can be industrially cultured on a large scale.
Description
Technical field
The present invention relates to a kind of sword-leaved cymbidium hybrid new breed gold prodigy's tissue-culturing rapid propagation seedling-cultivating method.
Background technology
The gold prodigy (
GoldenElf ' Sundust ') be the blue and Cymbidium hookerianum hybridization and getting with the four seasons before more than 10 year of China Taiwan Province, Flowering Phenology and proterties be closer to the growth characteristics of sword-leaved cymbidium, so generally it is classified as at present the sword-leaved cymbidium kind.It has pattern golden yellow plain, the fragrance of a flower is just pure, the characteristics such as leaf appearance unrestrainedness, deeply be subjected to consumer's favor since entering the market because of its " inexpensive " always, the general trend of market development is fabulous, existing 2 swords/basin is bloomed the plant price about 160 yuan/basin, less to the research of ' gold prodigy ' at present, the research of tissue culture technical elements there is not yet report.
Sword-leaved cymbidium (
Cymbidium ensifolium) delicate fragrance attacks the people, pattern is plain, cultivation history is long, and is various in style, mainly is distributed in all provinces in China western part and the southeast, especially distribute with Fujian and cultivation the widest, be one of China's export state orchid the earliest.China's sword-leaved cymbidium tissue culture technical research starting early; the history of existing three more than ten years; but because the sword-leaved cymbidium genetic character is complicated; tradition is viewed and admired take strange flower, different leaf as main; thereby in producing mainly take seed hybrid breeding and division propagation as main because reproduction coefficient is low, new varieties first listing stage often sky-high price propagandize; thereafter again fast drop brings bad impact to the technical research of sword-leaved cymbidium biological control, production.Along with the progress of sword-leaved cymbidium crossbreeding technology and the transformation of consumption concept, color leaf, fragrant flower, the sword-leaved cymbidium kind of the multiple characters combination such as spending more progressively become the new lover in market, such as gold prodigy, line skill sword-leaved cymbidium, color heart sword-leaved cymbidium etc., make these kinds enter into common people house, only have the tissue culture technology to be only the unique channel that guarantees its merit, also report is very few but the group of at present relevant sword-leaved cymbidium training is studied.
Summary of the invention
The purpose of this invention is to provide a kind of energy and implement the gold prodigy's of large-scale industrialized cultivation method for tissue culture.
Gold prodigy's of the present invention tissue-culturing rapid propagation seedling-cultivating method comprises the anti-brownization processing of outer plant corpus sterilization, just induces cultivation, shoot proliferation cultivation, culture of rootage and test-tube seedling transplanting, may further comprise the steps:
(1) the anti-brownization processing of explant sterilization: getting gold prodigy lateral bud stem apex is first culture material, older tissue prunes away after the collection, under running water, rinse well, with 0.1% carbendazim+0.2% detergent aqueous solution soaking 30 minutes, rinse well, then soaked 30 minutes with anti-brownization agent vitamin C distilled water solution (2%), with 0.1% mercuric chloride routine disinfection 10 minutes, separate the lateral bud stem apex at superclean bench at last;
(2) just induce condition of culture to be: 1/2MS+20-30g/L sucrose+8-10g/L agar+3.0mg/L TDZ+0.2mg/L NAA is medium; Medium pH 5.8; Illumination cultivation 14 hr/d; Intensity of illumination 1500-2000 Lux; Cultivation temperature is 25 ± 2 ℃;
(3) the shoot proliferation condition of culture is: 1/2MS+10g/L agar+30g/L sucrose+1-7mg/L TDZ+0.05-0.4mg/L NAA is medium; Medium pH 5.8; Illumination cultivation 14hr/d; Intensity of illumination 1500-2000Lux; Cultivation temperature is 25 ± 2 ℃;
(4) the culture of rootage condition is: 1/2MS+10g/L agar+35g/L sucrose+1g/L active carbon+1g/L sodium sulphite+0.1-0.9mg/L 6-BA+0.4-2.0mg/L NAA is medium, medium pH 5.8; Illumination cultivation 14hr/d; Intensity of illumination 1500-2000Lux; Cultivation temperature is 25 ± 2 ℃;
(5) the test-tube seedling transplanting condition is: culture of rootage is after 60 days, and hardening is 20 days again; Transplant after disinfecting seedling with 0.05% potassium permanganate solution; Room temperature is controlled at 15-30 ℃, and air humidity shelters from heat or light 60 ~ 70% at 80%-90%.
TDZ is 3-3.5mg/L in the gold prodigy's of the present invention tissue-culturing rapid propagation seedling-cultivating method shoot proliferation medium, and NAA is 0.1-0.2mg/L.
Gold prodigy's of the present invention tissue-culturing rapid propagation seedling-cultivating method is characterized in that: the 6-BA in the root media is 0.3-0.5mg/L, and NAA is 0.8-1.2mg/L.
Gold prodigy's of the present invention tissue-culturing rapid propagation seedling-cultivating method is characterized in that: can add 1g/L active carbon and 1g/L sodium sulphite in first inducing culture and the shoot proliferation medium.
Gold prodigy's of the present invention tissue-culturing rapid propagation seedling-cultivating method is characterized in that: the matrix of test-tube seedling transplanting is vermiculite or bark.
The beneficial effect that the present invention reaches:
(1) the lateral bud stem apex is used 0.1% mercuric chloride routine disinfection again with 2% vitamin C aqueous solution soaking after 30 minutes before sterilization, can effectively delay the Brown time and keep the explant vigor;
(2) after interpolation resists brownization agent 1g/L active carbon and 1g/L sodium sulphite simultaneously in the 1/2MS medium, explant, protocorm and Multiple Buds produce brownization in can reduction group training process, reached at present that explant is just induced, the brown rate of protocorm shoot proliferation is controlled at below 15%, the brown rate of Multiple Buds root induction is lower than below 5%;
(3) concentration is that the TDZ of 3.0-3.5mg/L and NAA that concentration is 0.1-0.2mg/L combination are conducive to explant and just induce and Protocorm Multiplication; Along with the increase of TDZ concentration, be conducive to the protocorm differentiation Multiple Buds, otherwise then be conducive to the protocorm differentiation protocorm, growth coefficient reaches 7.33, and shoot proliferation is cultivated 4-5 generation continuously; Concentration is the 6-BA of 0.3-0.5 mg/L and NAA that concentration is 0.8-1.2 mg/L combination, is conducive to the Multiple Buds root induction most, and inductivity reaches 100.0%;
(4) the test-tube seedling transplanting condition is: culture of rootage is after 60 days, and hardening is 20 days again; Transplant after disinfecting seedling with 0.05% potassium permanganate solution; Room temperature is controlled at 15-30 ℃, and air humidity is at 80%-90%, shelters from heat or light 70% to transplant, and adopting vermiculite or bark is transplanting medium, and survival rate reaches more than 95%;
(5) the first culture base of the present invention and shoot proliferation medium difference are less, grow seedlings than being easier to industrialized tissue culture.
Embodiment:
Getting gold prodigy 0.4cm lateral bud stem apex is first culture explant.With the older tissue that prunes away after the above-mentioned material collection, under running water, rinse well, with 0.1% carbendazim+0.2% detergent aqueous solution soaking 30 minutes, rinse well; Then soaked 30 minutes with anti-brownization agent vitamin C distilled water solution (2%), with 0.1% mercuric chloride routine disinfection 10 minutes, separate the lateral bud stem apex at superclean bench at last; Be seeded in just on the inducing culture first inducing culture prescription: 1/2MS+30g/L sucrose+10g/L agar+1g/L active carbon+1g/L sodium sulphite+3.5mg/L TDZ+0.2mg/L NAA with peeling off lateral bud stem apex after the sterilization; Light application time 14hr/d; Intensity of illumination 1800lux; Cultivation temperature (25 ± 2 ℃); Just culture is about 60 days, and the lateral bud stem apex is induced and differentiated 2-3 diameter 0.2cm protocorm, with its cutting and separating, inoculation; Take 1/2MS+10g/L agar+30g/L sucrose as minimal medium, add the 1g/L active carbon and add 1g/L active carbon+1g/L sodium sulphite; Shoot proliferation was cultivated after 60 days, and protocorm differentiates Multiple Buds and protocorm simultaneously, inoculated after Multiple Buds is cut into 2 protocorm simple buds of base portion band; So that 1/2MS+10g/L agar+30g/L sucrose+1g/L active carbon+1g/L sodium sulphite+3mg/L TDZ+0.2mg/L NAA is as the shoot proliferation medium, growth coefficient reaches 7.33, and shoot proliferation is cultivated 4-5 generation continuously; After being divided into the unrooted simple bud, the high Multiple Buds of 1-1.5cm that protocorm is induced inoculates; The root induction medium: 1/2MS+8g/L agar+25g/L sucrose+1g/L active carbon+0.5mg/L 6-BA+1.2mg/L NAA, root induction is after 60 days, 100% root induction, the 2-3 bar of taking root, the long 0.5-3.5cm of root; After the culture of rootage 60 days, hardening is 20 days again; With vermiculite or be transplanting medium, to transplant after disinfecting seedling with 0.05% potassium permanganate solution, room temperature is controlled at 25-30 ℃, and air humidity shelters from heat or light 70% at 80%-90%, and transplanting survival rate reaches 96%.
Claims (5)
1. gold prodigy's tissue-culturing rapid propagation seedling-cultivating method comprises the anti-brownization processing of explant sterilization, just induces cultivation, shoot proliferation cultivation, culture of rootage and test-tube seedling transplanting, it is characterized in that:
(1) the anti-brownization processing of explant sterilization: getting gold prodigy lateral bud stem apex is first culture material, older tissue prunes away after the collection, under running water, rinse well, with 0.1% carbendazim+0.2% detergent aqueous solution soaking 30 minutes, rinse well, then soaked 30 minutes with 2% anti-brownization agent vitamin C distilled water solution, with 0.1% mercuric chloride routine disinfection 10 minutes, separate the lateral bud stem apex at superclean bench at last;
(2) just induce cultivation, medium is: 1/2MS+20-30g/L sucrose+8-10g/L agar+3.0-3.5mg/L TDZ+0.1-0.2mg/L NAA, and pH 5.8, illumination cultivation 14 hr/d, intensity of illumination 1500-2000 Lux, cultivation temperature is 25 ± 2 ℃;
(3) shoot proliferation is cultivated, and medium is: 1/2MS+10g/L agar+30g/L sucrose+1-7mg/L TDZ+0.05-0.4mg/L NAA, and medium pH 5.8, illumination cultivation 14hr/d, intensity of illumination 1500-2000Lux, cultivation temperature is 25 ± 2 ℃;
(4) culture of rootage, medium is: 1/2MS+10g/L agar+35g/L sucrose+1g/L active carbon+1g/L sodium sulphite+0.1-0.9mg/L 6-BA+0.4-2.0mg/L NAA, and pH 5.8, illumination cultivation 14hr/d, intensity of illumination 1500-2000Lux, cultivation temperature is 25 ± 2 ℃;
(5) the test-tube seedling transplanting condition is: culture of rootage is after 60 days, and hardening is 20 days again, transplants after disinfecting seedling with 0.05% potassium permanganate solution, and room temperature 15-30 ℃, air humidity 80%-90% shelters from heat or light 60 ~ 70%.
2. gold prodigy's according to claim 1 tissue-culturing rapid propagation seedling-cultivating method, it is characterized in that: TDZ is 3-3.5mg/L in the shoot proliferation medium, NAA is 0.1-0.2mg/L.
3. gold prodigy's according to claim 1 tissue-culturing rapid propagation seedling-cultivating method, it is characterized in that: the 6-BA in the root media is 0.3-0.5mg/L, NAA is 0.8-1.2mg/L.
4. gold prodigy's according to claim 1 tissue-culturing rapid propagation seedling-cultivating method is characterized in that: just induce with the shoot proliferation medium in add 1g/L active carbon and 1g/L sodium sulphite.
5. gold prodigy's according to claim 1 tissue-culturing rapid propagation seedling-cultivating method, it is characterized in that: the matrix of test-tube seedling transplanting is vermiculite or bark.
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| CN104521751B (en) * | 2014-12-04 | 2017-03-22 | 中国科学院华南植物园 | Tissue culture efficient breeding and seedling growing method of hybrid orchid Huangjinxiaoshentong |
| CN106718885A (en) * | 2016-11-29 | 2017-05-31 | 广西南岜仔科技有限公司 | A kind of culture medium of efficient rapid induction plum blossom Callus formation |
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Application publication date: 20120215 Assignee: QIANXINAN GREEN ANIMAL AND PLANT SCIENCE AND TECHNOLOGY DEVELOPMENT CO.,LTD. Assignor: GUIZHOU INSTITUTE OF BIOLOGY Contract record no.: 2015520000010 Denomination of invention: Tissue culture and rapid propagation breeding method for golden elf sundust Granted publication date: 20130313 License type: Exclusive License Record date: 20150527 |
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