CN108207629A - A kind of Rhododendron jinggangshanicum Tam method for tissue culture - Google Patents
A kind of Rhododendron jinggangshanicum Tam method for tissue culture Download PDFInfo
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- CN108207629A CN108207629A CN201810014953.1A CN201810014953A CN108207629A CN 108207629 A CN108207629 A CN 108207629A CN 201810014953 A CN201810014953 A CN 201810014953A CN 108207629 A CN108207629 A CN 108207629A
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- rhododendron
- jinggangshanicum
- tam
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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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- Developmental Biology & Embryology (AREA)
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a kind of Rhododendron jinggangshanicum Tam method for tissue culture, it is characterized in that, Rhododendron jinggangshanicum Tam terminal bud and lateral bud is induced to obtain primary sterile test tube sprout using tissue cultures, after subculture and culture of rootage, aseptic seedling is tamed and obtains Rhododendron jinggangshanicum Tam tissue-cultured seedling.The Rhododendron jinggangshanicum Tam method for tissue culture of the present invention can obtain a large amount of rooted seedling in a short time, and production is not subject to seasonal restrictions, and this method provides a kind of new approach for the breeding and protection of China's endemic species Rhododendron jinggangshanicum Tam, has great importance.
Description
Technical field
The present invention relates to a kind of method for propagation, relate generally to a kind of Rhododendron jinggangshanicum Tam method for tissue culture.
Background technology
Rhododendron jinggangshanicum Tam (Rhododendron jinggangshanicum Tam) is the evergreen wood of Ericaceae Rhododendron
This plant is China endemic species, is distributed mainly on the Guidong in Shangyou County of Luoxiao mountain range, Suichuan, Jinggang Mountain and Hunan, and branches and leaves are numerous
Cyclopentadienyl, tree-like grace, pattern is gorgeous, and there is higher ornamental plantation to be worth.Rhododendron jinggangshanicum Tam is planted mainly by seminal propagation
Sub- germination rate is low, and the breeding time is long, it is bred and is protected so it is still necessary to a kind of effective quick-breeding methods.Tissue training
Foster technology is a kind of ways for training quickly bred risen in the present age, not only simply, rapidly, reproduction speed it is fast, more conducively plant
Popularization, large-scale production and the protection of new varieties.
Invention content
It is breeding and the guarantor of Rhododendron jinggangshanicum Tam it is an object of the invention to establish a kind of Rhododendron jinggangshanicum Tam method for tissue culture
Shield provides technical foundation.
The Rhododendron jinggangshanicum Tam method for tissue culture of the present invention, includes the following steps：
Rhododendron jinggangshanicum Tam terminal bud or lateral bud are taken, is cleaned with tap water, its outermost layer is peelled off on superclean bench, with 70% alcoholometer
After face sterilizes 30~60 s, 0.1% mercuric chloride sterilizes 8 minutes, it is inoculated in 0.5 mgL of adventitious bud induction culture base Read+ NAA-1
2.0 mgL of+6-BA-120 gL of+sucrose-17 gL of+agar-1, on pH 5.6,25 ± 2 DEG C of cultures, 12 h/d, light
Strong 2000 lx；Every 35 days in 1 mgL of subculture medium Read+6-BA-1+ ZT0.5mgL-120 gL of+sucrose-1+
7 gL of agar-1, subculture is primary in pH 5.6；Healthy and strong Rhododendron jinggangshanicum Tam clump bud seedling of the subculture sprout height more than 2cm is cut into list
Bud is seeded in 1 mgL of Read+IBA-1+ BA0.1mgL-1 + sucrose 30gL-1 7 gL of+agar-1, pH
5.6, culture of rootage 40d；Practice seedling 3d in domestication room, Rhododendron jinggangshanicum Tam domestication matrix presses following preparation：+ 30% leech of 70% turfy soil
Stone uses loading hole tray after a week with 20% carbendazim, and Rhododendron jinggangshanicum Tam tissue culture rooted seedling is planted after cleaning in soil matrix.
0.1% carbendazim solution is poured after transplanting immediately and determines root, takes Small plastic shed plastic covering thin film and sunshade net, relative humidity is kept in canopy
80%~90%, temperature is controlled at 20~28 DEG C.1 promoter and 1/2MS nutrient solutions are sprayed weekly, and culture is gradually taken off after 4 weeks
Remove sunshade net and plastic film.
The Rhododendron jinggangshanicum Tam method for tissue culture of the present invention can obtain a large amount of rooted seedling in a short time, and production is not
It is subject to seasonal restrictions, this method provides a kind of new approach for the breeding and protection of China's endemic species Rhododendron jinggangshanicum Tam, has weight
The meaning wanted.
Specific embodiment
The adventitious bud is derived from horseradish terminal bud or lateral bud, after being cleaned with tap water, aseptically peels off its outermost layer,
Again with 70% alcohol surface sterilization, 30~60 s, 0.1% mercuric chloride after ten minutes, is inoculated in adventitious bud induction culture base（Read+ NAA
0.5 mg·L-12.0 mgL of+6-BA-120 gL of+sucrose-17 gL of+agar-1,pH 5.6）, 25 ± 2 DEG C of cultures 40
After it, it is inoculated in subculture medium within every 35 days（Read + 6-BA 1 mg·L-1+ ZT0.5mgL-120 gL of+sucrose-1+
7 gL of agar-1, pH5.6.Sprout is cut into simple bud when growing to 2cm or so and is inoculated into root media（Read + IBA 1
mg·L-1+ BA0.1mgL-130 gL of+sucrose-17 gL of+agar-1, pH 5.6）Culture bottle bottle is unclamped in culture 40 days
Lid practices seedling 3d, then tissue-cultured seedling is taken out out of bottle in domestication room, remaining culture medium is cleaned up and carries out domestication culture.Jing Gang
The culture substrate of ghent azalea presses following preparation：+ 30% vermiculite of 70% turfy soil is used be packed into hole tray after a week with 20% carbendazim.It moves
0.1% carbendazim solution is poured after cultivation immediately and determines root, takes Small plastic shed plastic covering thin film and sunshade net, relative humidity is protected in canopy
It holds 80%~90%, temperature is controlled at 20~28 DEG C.1 promoter and 1/2MS nutrient solutions are sprayed weekly, after cultivating 4 weeks gradually
Throw off sunshade net and plastic film.Later, strengthen ventilation, normal fertilizer and water management, after 4 weeks during Rhododendron jinggangshanicum Tam seedling length to 5 leaf
Crop field can be transplanted.
1st, the influence that hormone concentration induces Rhododendron jinggangshanicum Tam primary
Primary inducing culture is Read+BA 0.5mgL-1+ NAA 0.1mgL-1；Read+BA 1.0mg·L-1+ NAA
0.1mg·L-1；Read+ BA 2.0mg·L-1+ NAA 0.5mgL-1Three kinds of culture mediums.It is added in 3 groups of culture mediums equal
The agar and sucrose of quality, the amount of agar is 7.5mgL-1, the amount of sucrose is 20gL-1.Primary is counted after 40d to lure
Conductance.The experimental results showed that Rhododendron jinggangshanicum Tam bud primary induction optimal medium is Read+ BA 2.0mgL-1+ NAA
0.5mg·L-1, primary inductivity is up to 87% under this concentration（It is shown in Table 1）
The influence that 1 hormon concentration of table induces Rhododendron jinggangshanicum Tam adventitious bud primary
| Number | Culture medium | Averagely it is inoculated with bud number（It is a） | Adventitious bud induction frequency |
| 1 | Read+BA 0.5mg·L-1+ NAA 0.1mgL-1 | 15 | 27% |
| 2 | Read+BA 1.0mg·L-1+ NAA 0.1mgL-1 | 15 | 33% |
| 3 | Read+BA 2.0mg·L-1+ NAA 0.5mgL-1 | 15 | 87% |
2nd, the influence of hormon sterile seedling proliferation to Rhododendron jinggangshanicum Tam
The aseptic seedling of induction is inoculated into respectively in the read culture mediums containing different hormone combinations（Table 2）, 9 processing altogether,
Each processing repeats 5 times, adds 2% sucrose, 0.7% agar, adjusts pH value to 5.6.In (25+2) DEG C, 12 h/d, light intensity 2000
35d is cultivated under the conditions of lx, counts bud proliferative conditions.The experimental results showed that Rhododendron jinggangshanicum Tam bud proliferation optimal medium is Read
+ 6-BA 1 mg·L-1+ 0.5mgL-120 gL of ZT+ sucrose-17 gL of+agar-1, proliferation times highest under this concentration,
Up to 3.76 times or more
2 orthogonal test L of table8（27）
3 Rhododendron jinggangshanicum Tam adventitious bud proliferation orthogonal experiments of table
Influence of 3 hormons to Rhododendron jinggangshanicum Tam adventitious bud rooting
The adventitious bud of induction is inoculated into respectively in the read culture mediums containing different hormone combinations（Table 4）, 4 processing altogether, often
A processing repeats 5 times, adds 2% sucrose, 0.7% agar, adjusts pH value to 5.6.In (25+2) DEG C, 12 h/d, 2000 lx of light intensity
40d is cultivated under illumination condition, the rear growth conditions for observing seedling and statistical average are taken root item number and average root long.Experimental result table
It is bright, Read+IBA0.5mgL-1+ BA 0.1mgL-1In the green of health, average root is grown to be reached culture medium plant leaf blade
2.44cm, mean elements 4.2（It is shown in Table 4）.
Influence of 4 hormon of table to Rhododendron jinggangshanicum Tam adventitious bud rooting
Claims (2)
1. a kind of Rhododendron jinggangshanicum Tam method for tissue culture, it is characterised in that：It includes the following steps：
（1）, take Rhododendron jinggangshanicum Tam terminal bud or lateral bud, clean, its outermost layer is peelled off on superclean bench, is disappeared with 70% alcohol surface
After 30~60 s of poison, 0.1% mercuric chloride sterilize 8 minutes, it is inoculated in 0.5 mg L-1+6- of adventitious bud induction culture base Read+ NAA
On 2.0 mg L-1+ sucrose of BA, 20 g L-1+ agar, 7 g L-1, pH 5.6,25 ± 2 DEG C of cultures, 12 h/d, light intensity 2000
lx；
（2）, every 35 days in 1 mg L-1+ZT0.5mg L-1+ sucrose of subculture medium Read+6-BA, 20 g L-1+ agar
Subculture is primary in 7 g L-1, pH 5.6；
（3）, subculture sprout height be cut into simple bud more than the healthy and strong Rhododendron jinggangshanicum Tam clump bud seedling of 2cm, be seeded in Read+IBA 1
Mg L-1+BA0.1mg L-1+sucrose 30g L-1+agar 7 g L-1, pH 5.6, culture of rootage 40d；
（4）, in domestication room practice seedling 3d, domestication matrix is by following preparation ,+30% vermiculite of 70% turfy soil, with 20% carbendazim, one
Zhou Houyong is packed into hole tray, and Rhododendron jinggangshanicum Tam tissue culture rooted seedling is planted after cleaning in soil matrix；
（5）, pour 0.1% carbendazim solution immediately after transplanting and determine root, take Small plastic shed plastic covering thin film and sunshade net, it is opposite in canopy
Humidity is maintained at 80%~90%, and temperature is controlled at 20~28 DEG C, and culture throws off sunshade net and plastic film after 4 weeks.
2. Rhododendron jinggangshanicum Tam method for tissue culture according to claim 1, it is characterised in that：The step（5）In, weekly
Spray 1 promoter and 1/2MS nutrient solutions.
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Citations (6)
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|---|---|---|---|---|
| CN1732759A (en) * | 2005-08-17 | 2006-02-15 | 中国林业科学研究院林业研究所 | Tissue culturing, rapid propagating and transplanting method of Rhododendron mucronulatum Turcz. |
| CN101044840A (en) * | 2006-03-31 | 2007-10-03 | 贵州科学院 | Method for fast breeding western azalea and culture medium therefor |
| CN101292628A (en) * | 2008-06-11 | 2008-10-29 | 国家林业局泡桐研究开发中心 | Cultivation method for north America rhododendron |
| CN102870680A (en) * | 2012-10-23 | 2013-01-16 | 刘如石 | Efficient rapid propagation technique appropriate for detoxified rabbiteye blueberries |
| WO2014077779A1 (en) * | 2012-11-16 | 2014-05-22 | Temasek Life Sciences Laboratory Limited | Direct and indirect organogenesis of jatropha |
| KR101439618B1 (en) * | 2012-08-21 | 2014-09-11 | 경상대학교산학협력단 | A Method for Mass Propagation of Rhododendron Keiskei var. hypoglaucum by Plant Tissue Culture |
-
2018
- 2018-01-08 CN CN201810014953.1A patent/CN108207629A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1732759A (en) * | 2005-08-17 | 2006-02-15 | 中国林业科学研究院林业研究所 | Tissue culturing, rapid propagating and transplanting method of Rhododendron mucronulatum Turcz. |
| CN101044840A (en) * | 2006-03-31 | 2007-10-03 | 贵州科学院 | Method for fast breeding western azalea and culture medium therefor |
| CN101292628A (en) * | 2008-06-11 | 2008-10-29 | 国家林业局泡桐研究开发中心 | Cultivation method for north America rhododendron |
| KR101439618B1 (en) * | 2012-08-21 | 2014-09-11 | 경상대학교산학협력단 | A Method for Mass Propagation of Rhododendron Keiskei var. hypoglaucum by Plant Tissue Culture |
| CN102870680A (en) * | 2012-10-23 | 2013-01-16 | 刘如石 | Efficient rapid propagation technique appropriate for detoxified rabbiteye blueberries |
| WO2014077779A1 (en) * | 2012-11-16 | 2014-05-22 | Temasek Life Sciences Laboratory Limited | Direct and indirect organogenesis of jatropha |
Non-Patent Citations (3)
| Title |
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| YULIANNA G. ZAYTSEVA等: "Effects of thidiazuron on in vitro morphogenic response", 《IN VITRO CELLULAR & DEVELOPMENTAL BIOLOGY-PLANT》 * |
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Application publication date: 20180629 |