CN103329807A - Method for rapidly breeding hybrid orchid by root, as well as culture medium - Google Patents
Method for rapidly breeding hybrid orchid by root, as well as culture medium Download PDFInfo
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- CN103329807A CN103329807A CN2013102995058A CN201310299505A CN103329807A CN 103329807 A CN103329807 A CN 103329807A CN 2013102995058 A CN2013102995058 A CN 2013102995058A CN 201310299505 A CN201310299505 A CN 201310299505A CN 103329807 A CN103329807 A CN 103329807A
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Abstract
The invention discloses a method for rapidly breeding a hybrid orchid by a root, as well as a culture medium, and belongs to the technical field of plant tissue culture. The method comprises steps as follows: a root segment of the hybrid orchid is selected and inoculated on an induction culture medium for dark culture, and a protocorm is induced; after a large amount of the protocorms are proliferated, the protocorms are transferred for light culture and develop into tufted seedlings; and the tufted seedlings are cut into single seedlings and inoculated to a strong seedling rooting culture medium, and a rooting plant is cultured and obtained. According to the method, MS (murashige and skoog) is taken as a basic culture medium, 6-BA (6-benzyladenine), PIC (4- amino-3,5,6-trichloropyridine-2-acid) and casein hydrolysate are required to be added to the induction culture medium, and the strong seedling rooting culture medium is the MS culture medium without hormone. According to the method and the culture medium, the breeding efficiency is high, the operation is simple and convenient, and all of the protocorm inductivity, the plant rooting rate and the transplanting survival rate of root explants can reach 100%.
Description
Technical field
The invention belongs to field of plant tissue culture technique, be specifically related to a kind of root that utilizes and breed hybridization blue method and medium fast.
Background technology
The hybridization orchid be the novel orchid that come out by China blue and hybrid cymbidium selection cross [Lu Jiliang. blue production and marketing present situation [J] is hybridized in pre-test Yunnan. flowers and trees bonsai (flower gardening), 2013, (2): 36-37; Liang Fang, Cui Bo, Ma Jie, Ye Yongzhong. hybridize blue protocorm propagation and differentiation research [J]. Anhui agricultural science, 2008,36 (13): 5309-5310,5353].Crossbred orchid is medium-sized, look gorgeous, delicate fragrance, grace, remedied the shortcoming of hybrid cymbidium British plain spirits, spending of their collection hybrid cymbidiums is big, look gorgeous, the florescence is long and the delicate fragrance of China blue, elegance and charm are one, have very high ornamental value and vast market prospect [Zhang Xianyun, Yuan Xiuyun, the horse outstanding person. the blue tissue of hybridization is cultivated and quick propagating technology research [J]. Henan science, 2009,27 (4): 419-421; Lu Ran. Yunnan hybridization blue fast-selling [J]. Chinese flower gardening 2012, (1): 18].It is strong to hybridize blue adaptability, some early flower variety can bloom for 1 year twice, remedied the deficiency that the hybrid cymbidium florescence concentrates.Because above-mentioned advantage, the hybridization orchid is welcome by grower and consumer deeply, the situation that is always in recent years that both production and marketing thrive, development swift and violent [Lu Jiliang. blue production and marketing present situation [J] is hybridized in pre-test Yunnan. flowers and trees bonsai (flower gardening), 2013, (2): 36-37; Lu Ran. Yunnan hybridization blue fast-selling [J]. Chinese flower gardening 2012, (1): 18].
Traditional division propagation cycle is long, and speed is slow, and reproduction coefficient is low, far can not satisfy the blue industrialization production requirement of hybridization; Therefore, application organizes is cultivated breeding hybridization orchid fast, and is significant for the industrialization development that hybridization is blue.
The blue hereditary factor that carries the state orchid of hybridization, compare tissue cultivation difficulty with cattleyas such as hybrid cymbidium, Moth orchides bigger, at present only find 3 pieces about hybridizing the classical paper [Liang Fang of blue tissue-culturing quick-propagation, Cui Bo, Ma Jie, Ye Yongzhong. hybridize blue protocorm propagation and differentiation research [J]. Anhui agricultural science, 2008,36 (13): 5309-5310,5353; Zhang Xianyun, Yuan Xiuyun, horse outstanding person. the blue tissue of hybridization is cultivated and quick propagating technology research [J]. Henan science, 2009,27 (4): 419-421; Shen Hanguo, Deng Ying, Yang Zhenming, Lan Weiquan, Luo Lixia. hybridization blue ' 18 sound of laughing ' Study on tissue culture. Zhang Qixiang. Chinese ornamental horticulture progress [M]. Beijing: China Forest publishing house, 2012:311-313].
Explant is based on stem apex, lateral bud, and is big to the plant injury; Be prone to brownization after the explant inoculation, the initial incubation difficulty is bigger; The incubation complexity, reproduction coefficient is low; Be the subject matter of the blue tissue-culturing quick-propagation existence of hybridization at present.
Organize the explant of cultivating with the root work, have big, the easy acquisition of the amount of drawing materials, plant is injured little unique advantage, but in the blue Study on tissue culture of present hybridization, do not see report and the patent of invention of making explant with root.
Summary of the invention
The objective of the invention is to overcome above-mentioned the deficiencies in the prior art, provide a kind of root that utilizes to breed hybridization blue method and medium fast.Being specially with the root is explant, induces, breeds protocorm at inducing culture and make protocorm develop into the seedling of growing thickly, and breeding hybridization is blue fast; Required defined medium and the culture technique of each cultivation stage proposed simultaneously.The blue root of hybridization provided by the present invention is cultivated the method for regeneration plant, and that can finish protocorm a step induces, breeds and become the seedling process; The protocorm inductivity of explant reaches 100%, plant rooting rate 100%, transplanting survival rate 100%; Have reproduction coefficient height, planting percent height, transplanting survival rate height and the low characteristics of cost.
The objective of the invention is to be achieved through the following technical solutions:
First aspect the present invention relates to a kind of root that utilizes and breeds the blue method of hybridization fast, comprises the steps:
Step 1 is chosen the blue root segment of hybridization, is seeded on the inducing culture and secretly cultivates, and induces, breeds protocorm;
Step 2 changes the protocorm on the described inducing culture over to light and cultivates, and protocorm develops into the seedling of growing thickly;
Step 3 cuts into single seedling with the described seedling of growing thickly and is transferred on the strengthening seedling and rooting medium, cultivates to obtain the plant that takes root.
Preferably, it is with a kind of inducing culture that inducing culture in the step 1 and step 2 transfer light inlet are cultivated used medium, and described inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.Described cultivation is dark the cultivation, and the temperature of cultivation is 25 ± 2 ℃.
Preferably, in the step 2, described inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.Described cultivation is that light is cultivated, and the temperature of cultivation is 25 ± 2 ℃.
Preferably, in the step 3, described strengthening seedling and rooting medium is MS+AC100mg/L+ caseinhydrolysate 500mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.Described cultivation is that light is cultivated, and the temperature of cultivation is 25 ± 2 ℃.
The invention still further relates to a kind of inducing culture, this medium can be induced, be bred protocorm and make protocorm develop into the seedling of growing thickly, and described inducing culture is for adding 6-BA and PIC simultaneously in the MS minimal medium.
Preferably, described inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.
MS component in the medium used herein is seen document " Murashige T, Skoog F.A revised mediumfor rapid growth and bioassays with tobacco tissue cultures[J] .Physiol.Plant.1962,15,473-497 ", this belongs to the common practise of this area.
The present invention's employed PIC chemical name in medium is 4-amino-3,5,6-trichloropyridine-2-acid, and molecular formula is C
6H
3Cl
3N
2O
2, the general picloram by name of Chinese or picloram, English name is Picloram.
Compared with prior art, the present invention has following beneficial effect:
1. be that explant obtains regeneration plant to hybridize blue root first, enlarged the explant source that the blue tissue of hybridization is cultivated.Root obtains to compare with seed easily as explant,, compares with other vegetative tissue for the outer planting physical efficiency keeps the maternal plant proterties with the root, and the amount of drawing materials is bigger, and is little to the plant injury, and the present invention is significant for hybridization blue seedling production and industry development.
2. reproductive efficiency height, robust plant, transplanting survival rate height.Explant protocorm inductivity of the present invention reaches 100%, and brownization, vitrification phenomenon are seen in whole incubation end, have accelerated the cultivation process, have improved seedling quality.The regeneration plant well developed root system that obtains just can produce many sturdy roots, plant rooting rate 100% without root-growing agent; The plant strain growth stalwartness, transplanting survival rate 100%.
3. protocorm is induced with becoming seedling and is only needed two step incubation, and incubation and used medium are simple.First step protocorm is induced only to cultivate and need be added 6-BA, and PIC and caseinhydrolysate need not add other complicated organic matters such as banana puree, Coconut Juice, and the light requirement photograph has not been saved the energy.Second step only needed former medium is forwarded under the light, and protocorm just can develop into the seedling of growing thickly.The 3rd step strengthening seedling and rooting is cultivated, and does not need any growth regulator, and the whole plant that can directly be taken root is with low cost.
4. first picloram (PIC) is applied to hybridize the tissue cultivation of blue root, protocorm is induced, differentiation effect is good.
5. can obtain more root explant by the root regeneration plant, and then induce more protocorm in a large number, constantly, realize circulation propagation, be conducive to batch production production.
Description of drawings
By reading the detailed description of non-limiting example being done with reference to the following drawings, it is more obvious that other features, objects and advantages of the present invention will become:
Fig. 1 is hybridized blue root segment and is induced the schematic diagram that forms protocorm;
Fig. 2 protocorm is bred the schematic diagram with growth under no optical condition;
Fig. 3 protocorm changes the back schematic diagram that turns green and be divided into the seedling of growing thickly under the light over to;
Fig. 4 changes the schematic diagram of the plant that takes root that forms behind the strengthening seedling and rooting medium over to;
The schematic diagram of Fig. 5 hybridization orchid of transplant survival in the greenhouse.
Embodiment
The present invention is described in detail below in conjunction with specific embodiment and accompanying drawing.Following examples will help those skilled in the art further to understand the present invention, but not limit the present invention in any form.Should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make some distortion and improvement.These all belong to protection scope of the present invention.
Employed term except as otherwise noted, generally has the implication of those of ordinary skills' common sense in the present invention.In following embodiment, the various processes of Xiang Ximiaoshuing and method are not conventional methods as known in the art, and agents useful for same and be necessary to list its composition person is all indicated when occurring first, thereafter used identical reagent if no special instructions, all identical with the content of indicating first.
Embodiment 1
Be material with the root of hybridizing blue test-tube plantlet, inoculation after cutting into chunks, the dark cultivation, about 25 ℃ of temperature (25 ± 2 ℃), induce protocorm, the protocorm inducing culture is MS (Murashige and Skoog, 1962)+6-BA (6-benzyl aminoadenine) 0.5mg/L+PIC (4-amino-3,5,6-trichloropyridine-2-acid) 0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.Root segment expands after 12 days, begins to produce protocorm, and as shown in Figure 1, protocorm is bred in a large number subsequently, and breaks up the bud seedling gradually, as shown in Figure 2, and protocorm inductivity 100%.
To induce, breed the protocorm inducing culture that a large amount of protocorms, bud seedling to be transferred under the light, about 25 ℃ of temperature (25 ± 2 ℃), protocorm and bud seedling begin to turn green after 3 days, and develop into the seedling of growing thickly gradually, protocorm also continues propagation simultaneously, as shown in Figure 3.
The seedling of growing thickly is cut into single seedling to be transferred on the strengthening seedling and rooting medium of MS+AC (active carbon) 100mg/L+ caseinhydrolysate 500mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L, illumination every day 16 hours, about 25 ℃ of temperature, the unrooted seedling will develop into normal plant gradually, take root simultaneously, as shown in Figure 4, the plant rooting rate 100%.Through cultivation in about 65 days, each root explant can induce 60～80 plant that take root.
To be generated taking root in after strain grows 5～6 leaves opened bottle cap, and at room temperature hardening is 7～10 days, plant in the greenhouse behind the flush away agar, and robust growth, survival rate 100%, as shown in Figure 5.
Embodiment 2
The root of the test-tube plantlet that obtains with embodiment 1 is material, inoculation after cutting into chunks, the dark cultivation, about 25 ℃ of temperature (25 ± 2 ℃), induce protocorm, the protocorm inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L, and the protocorm inductivity is 100%.Cultivate after 30 days, the bud seedling of selecting differentiation moves to and carries out the light cultivation on the new protocorm inducing culture MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L, simultaneously undifferentiated protocorm is stayed and continued on the former medium to change the light cultivation again over to after propagation, the differentiation, adopt later on the incubation step identical with embodiment 1, through cultivation in about 80 days, but more than average root induction plant 100 strains of each root explant.
More than specific embodiments of the invention are described.It will be appreciated that the present invention is not limited to above-mentioned specific implementations, those skilled in the art can make various distortion or modification within the scope of the claims, and this does not influence flesh and blood of the present invention.
Claims (8)
1. one kind is utilized root to breed the blue method of hybridization fast, it is characterized in that, comprises the steps:
Step 1 is chosen the blue root segment of hybridization, is seeded on the inducing culture and secretly cultivates, and induces, breeds protocorm;
Step 2 changes the protocorm on the described inducing culture over to light and cultivates, and protocorm develops into the seedling of growing thickly;
Step 3 cuts into single seedling with the described seedling of growing thickly and is transferred on the strengthening seedling and rooting medium, cultivates to obtain the plant that takes root.
2. the root that utilizes as claimed in claim 1 is bred the blue method of hybridization fast, it is characterized in that, it is with a kind of inducing culture that inducing culture in the step 1 and step 2 transfer light inlet are cultivated used medium, and described inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.
3. the root that utilizes as claimed in claim 1 is bred the blue method of hybridization fast, it is characterized in that in the step 3, described strengthening seedling and rooting medium is MS+AC100mg/L+ caseinhydrolysate 500mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.
4. the root that utilizes as claimed in claim 1 is bred the blue method of hybridization fast, it is characterized in that in the step 1, the temperature of described dark cultivation is 25 ± 2 ℃.
5. the root that utilizes as claimed in claim 1 is bred the blue method of hybridization fast, it is characterized in that in the step 2, the temperature that described light is cultivated is 25 ± 2 ℃.
6. the root that utilizes as claimed in claim 1 is bred the blue method of hybridization fast, it is characterized in that, in the step 3, described cultivation is that light is cultivated, and the temperature of cultivation is 25 ± 2 ℃.
7. one kind is used for utilizing root to breed the inducing culture of the blue method of hybridization fast, it is characterized in that, described inducing culture can be induced, be bred protocorm and make protocorm develop into the seedling of growing thickly, and described inducing culture is in the MS minimal medium, adds 6-BA and PIC simultaneously.
8. inducing culture as claimed in claim 7 is characterized in that, described inducing culture is MS+6-BA0.5mg/L+PIC0.6mg/L+ caseinhydrolysate 200mg/L+ sucrose 30000mg/L+ agar powder 5000mg/L.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103843567A (en) * | 2014-03-10 | 2014-06-11 | 上海市农业科学院 | Method for rapidly propagating Chinese cymbidium by using root induced protocorm and culture medium thereof |
| CN104255492A (en) * | 2014-09-19 | 2015-01-07 | 东莞市生物技术研究所 | Seedling culturing method for preventing hybrid orchid virus |
| CN108990799A (en) * | 2018-07-25 | 2018-12-14 | 杭州市农业科学研究院 | A method of hybrid orchid sterilizable material is efficiently obtained using newborn lateral bud |
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| KR20040050684A (en) * | 2002-12-10 | 2004-06-16 | 제주도(농업기술원) | New Variety 'Eoseungsaeng' -Interspecific hybrid between Cymbidium 'Daemyeongran' variety and Cymbidium 'Heidi' |
| KR20050041817A (en) * | 2003-10-31 | 2005-05-04 | 제주도(농업기술원) | New variety 'daewon' - interspecific hybrid between cymbidium goeringii and cymbidium sinecense |
| CN101455178A (en) * | 2007-12-11 | 2009-06-17 | 姜红颖 | Culture method of new species hybrid cymbidium |
| CN102754599A (en) * | 2012-07-16 | 2012-10-31 | 上海市农业科学院 | Method for quickly breeding cymbidium hybridium by use of root inducing protocorm |
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2013
- 2013-07-16 CN CN201310299505.8A patent/CN103329807B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20040050684A (en) * | 2002-12-10 | 2004-06-16 | 제주도(농업기술원) | New Variety 'Eoseungsaeng' -Interspecific hybrid between Cymbidium 'Daemyeongran' variety and Cymbidium 'Heidi' |
| KR20050041817A (en) * | 2003-10-31 | 2005-05-04 | 제주도(농업기술원) | New variety 'daewon' - interspecific hybrid between cymbidium goeringii and cymbidium sinecense |
| CN101455178A (en) * | 2007-12-11 | 2009-06-17 | 姜红颖 | Culture method of new species hybrid cymbidium |
| CN102754599A (en) * | 2012-07-16 | 2012-10-31 | 上海市农业科学院 | Method for quickly breeding cymbidium hybridium by use of root inducing protocorm |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103843567A (en) * | 2014-03-10 | 2014-06-11 | 上海市农业科学院 | Method for rapidly propagating Chinese cymbidium by using root induced protocorm and culture medium thereof |
| CN104255492A (en) * | 2014-09-19 | 2015-01-07 | 东莞市生物技术研究所 | Seedling culturing method for preventing hybrid orchid virus |
| CN104255492B (en) * | 2014-09-19 | 2016-11-30 | 东莞市生物技术研究所 | A kind of method for culturing seedlings preventing hybrid orchid to visit wire drawing virus |
| CN108990799A (en) * | 2018-07-25 | 2018-12-14 | 杭州市农业科学研究院 | A method of hybrid orchid sterilizable material is efficiently obtained using newborn lateral bud |
| CN108990799B (en) * | 2018-07-25 | 2021-11-12 | 杭州市农业科学研究院 | Method for efficiently obtaining sterile material of hybrid orchid by utilizing new lateral buds |
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