CN104920225B - A kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L - Google Patents
A kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L Download PDFInfo
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- CN104920225B CN104920225B CN201510411373.2A CN201510411373A CN104920225B CN 104920225 B CN104920225 B CN 104920225B CN 201510411373 A CN201510411373 A CN 201510411373A CN 104920225 B CN104920225 B CN 104920225B
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Abstract
The invention discloses a kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L, comprise the following steps：In the newborn spray that 5cm or so is gathered after spring resting bud stripping, the MS culture mediums that additional 0.5 1.0mg/L 6 benzylaminopurines are inoculated in after surface sterilization, axillary bud sprouting is induced；By the tender stem of acquisition, the section of 2 3cm length is cut into, is inoculated in MS culture mediums, carry out Multiplying culture；The simple bud of robust growth, long 3 4cm is inoculated in 1/2MS culture mediums, culture of rootage is carried out；When root length is to 3 6cm, bottle cap is opened, hardening is transplanted after 3 days.Adjustment and the control of condition of culture that the present invention passes through nutrient media components, gold leaf Acer negundo. L body material axillary bud sprouting can comparatively fast be started, induction test tube seedling stem section directly produces the more adventitious bud of number without callus, fibrous root quantity is more, transplanting survival rate provides an effective way up to 85% for gold leaf Acer negundo. L industrial seedling rearing.
Description
Technical field
The present invention relates to method for plant tissue culture, and in particular to a kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L.
Background technology
Gold leaf Acer negundo. L (Acer negundo ' Aurea ') belongs to Aceraceae (Aceraceae) Acer (Acer) seeds, is multiple
The variety of leaf maple, deciduous tree is the excellent ornamental tree species that China introduced from Europe in recent years.Gold leaf Acer negundo. L growth potential
Vigorous, sprig is smooth, golden yellow；Imparipinnate leaf is to life, and leaf is larger, and leaf color is soft, and spring is golden yellow, fades to yellowish green
Color, with high ornamental value.Gold leaf Acer negundo. L Bioclimatic analysis is wide, and resistance, drought-resistant, cold resistant (is resistant to -40-45 DEG C
Low temperature), resistance to slight alkaline land, resistance to flue dust, root sprout tillers is strong.Because of its stronger suitable natural disposition and higher ornamental value, China of China
North, northeast, northwest and East China, which have, makees Landscape Trees cultivation, and supply falls short of demand for existing market.
The main modes of reproduction of gold leaf Acer negundo. L is propagation by grafiting in production.Propagation by grafiting survival rate is higher, but stock is selected
Select it is improper later stage tree vigo(u)r can be influenceed to grow, and breeding coefficient is relatively low, input cost is larger.Tissue culture technique is that breeding is fast numerous
One of main path, breeding coefficient is high, makes a variation small.Report on gold leaf Acer negundo. L tissue cultures is to its tissue culture seldom
The pre-test of performance, it is the same with the tissue cultures of other Aceraceae seeds, there is sprouting and start that slow, growth coefficient is low, performance of taking root
The low defect of poor, survival rate.
The content of the invention
The invention provides a kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L, overcome in the tissue culture culture of gold leaf Acer negundo. L
Start that slow, growth coefficient is low, the deficiency that poor performance of taking root, survival rate are low.
To achieve the above object, the technical scheme taken of the present invention is：
A kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L, comprises the following steps：
S1, the newborn spray in collection 5cm or so after spring resting bud stripping, are inoculated in additional 0.5- after surface sterilization
In the MS culture mediums of 1.0mg/L 6- benzylaminopurines (6-BA), in 20 ± 2 DEG C -25 ± 2 DEG C, illumination in 14 hours, illumination is strong
Spend culture under the conditions of 2000lx, the tender stem of induction axillary bud sprouting to 6-8cm length；
S2, the tender stem for taking step S1 to obtain, are cut into the section of 2 petioles of 2-3cm bands, are inoculated in additional 0.5mg/L 6- benzyls
In base adenine phosphate (6-BA), 0.01-0.05mg/L Thidiazurons (TDZ), the MS culture mediums of 0.05mg/L methyl α-naphthyl acetates (NAA), in
20 ± 2 DEG C -25 ± 2 DEG C, illumination in 14 hours under the conditions of intensity of illumination 2000lx, carries out Multiplying culture；
Robust growth, long 3-4cm simple bud obtained by S3, selecting step S2 are inoculated in the fourth of indoles containing 0.01-0.05mg/L
In the 1/2MS culture mediums of sour (IBA), in 20 ± 2 DEG C -25 ± 2 DEG C, illumination in 14 hours under the conditions of intensity of illumination 2000lx, is carried out
Culture of rootage.
S4, when root length is to 3-6cm, bottle cap is opened, hardening gently presss from both sides out seedling after 3 days with tweezers, it is light in flowing water
It is light to rinse root culture medium, transplant into the disposal plastic cup equipped with matrix, by plastic cup, every 10 are placed in plastics valve bag
In, room temperature hardening grows after 30 days by potted plant in rooted seedling in the matrix containing perlite, vermiculite and rural area soil.
Wherein, 6g/L agar powders, 30g/L sucrose are also included in the culture medium in the step S1.
Wherein, proliferated culture medium also includes 6g/L agar powders, 30g/L sucrose, 0.25g/L 2- morpholines in the step S2
Ethyl sulfonic acid (MES), 0.2g/L l-glutamines.
Wherein, root media also includes 7g/L agar powders, 15g/L sucrose, 0.25g/L 2- morpholines in the step S3
Ethyl sulfonic acid (MES).
Wherein, in the step S4 matrix of disposal plastic cup by perlite and peat soil in 1: 1 ratio mixing
Into.
Wherein, perlite, vermiculite, the ratio of rural area soil are 3: 2: 5 in the step S4.
The invention has the advantages that：
Solve sprouting during Aceraceae seeds tissue culture and start that slow, growth coefficient is low, taking root property is poor, transplanting survival rate is low
The problem of.By the adjustment and the control of condition of culture of nutrient media components, it can comparatively fast start gold leaf Acer negundo. L body material armpit
Bud is sprouted, and induction test tube seedling stem section directly produces the more adventitious bud of number without callus, average each explant propagation
Bud subnumber is up to 5.8, and adventitious bud rooting rate is up to 100%, and main root is flourishing, and fibrous root quantity is more, and transplanting survival rate, up to 85%, is gold
Leaf Acer negundo. L industrial seedling rearing provides an effective way.
Embodiment
In order that objects and advantages of the present invention are more clearly understood, the present invention is carried out with reference to embodiments further
Describe in detail.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, it is not used to limit this hair
It is bright.
Embodiment 1
In the newborn spray that 5cm or so is gathered after spring resting bud stripping, it is inoculated in after surface sterilization containing various concentrations ladder
In the 6-BA (0,0.5,1.0mg/L) of degree, 6g/L agar powders, the MS culture mediums of 30g/L sucrose, in 25 ± 2 DEG C, 14 small time
According to culture under the conditions of intensity of illumination 2000lx induces axillary bud sprouting.Count the axillary bud sprouting time and sprout stem section state.As a result
Show, in the culture medium containing 0mg/L, 0.5mg/L, 1.0mg/L 6-BA the axillary bud sprouting time of explant be respectively 25 days, 15
My god, 12 days, addition 6-BA can shift to an earlier date axillary bud and start the time, and the tender stem state sprouted in the culture mediums of 6-BA containing 0.5-1.0mg/L is just
Often, blade is open and flat, sprouts tender stem elongation speed.
Embodiment 2
In the newborn spray that 5cm or so is gathered after spring resting bud stripping, 6- containing 0.5mg/L is inoculated in after surface sterilization
In BA, 6g/L agar powder, the MS culture mediums of 30g/L sucrose, in 25 ± 2 DEG C, illumination in 14 hours, under the conditions of intensity of illumination 2000lx
Culture, induces axillary bud sprouting.The tender stem that axillary bud sprouting is obtained, is cut into the section of 2 petioles of 2-3cm bands, is seeded in and contains
0.5mg/L 6BA, 0.05mg/L NAA, various concentrations gradient TDZ (0,0.01,0.05,0.1mg/L), 6g/L agar powders,
In 30g/L sucrose, the MS culture mediums of 0.25g/L MES, 0.2g/L l-glutamines, in 25 ± 2 DEG C, illumination in 14 hours, illumination
Culture carries out Multiplying culture under the conditions of intensity 2000lx.Culture counts proliferation times after 50 days.As a result show, containing 0mg/L,
In 0.01mg/L, 0.05mg/L, 0.1mg/L TDZ culture medium the bud subnumber of average each stem section propagation be respectively 2,4.8
It is individual, 5.8,2.6.The bud growth bred under first three concentration is normal, the bud tubbiness bred under latter concentration, turns round
Song, explant incision callusization is serious.
Embodiment 3
In the newborn spray that 5cm or so is gathered after spring resting bud stripping, 6- containing 0.5mg/L is inoculated in after surface sterilization
In BA, 6g/L agar powder, the MS culture mediums of 30g/L sucrose, in 25 ± 2 DEG C, illumination in 14 hours, under the conditions of intensity of illumination 2000lx
Culture, induces axillary bud sprouting.The tender stem that axillary bud sprouting is obtained, is cut into the section of 2 petioles of 2-3cm bands, is seeded in and contains
0.5mg/L 6BA, 0.05mg/L NAA, 0.05mg/L TDZ, 6g/L agar powder, 30g/L sucrose, 0.25g/L MES, 0.2g/L
In the MS culture mediums of l-glutamine, in 25 ± 2 DEG C, illumination in 14 hours is cultivated under the conditions of intensity of illumination 2000lx and bred
Culture.The robust growth obtained will be bred, long 3-4cm simple bud is seeded in the IBA of 1/2MS annex various concentrations gradients respectively
In (0.01,0.05,0.1mg/L) culture medium, and 1/2MS annex various concentrations gradients NAA (0.01,0.05,0.1mg/L)
Carry out also including 7g/L agar powders, 15g/L sucrose, 0.25g/L MES in culture of rootage, various culture mediums in culture medium.Culture
25 ± 2 DEG C of condition, illumination in 14 hours, intensity of illumination 2000lx.Count rootage duration, the rooting rate after 50 days, observed and recorded life
Root situation.As a result show, the adventitious bud rooting cultivated in the root media containing NAA is slow, root is short, thin and delicate, and main root is not obvious,
Earliest rootage duration is 21 days, maximum rooting rate 62%, the long 1.54cm of average root.Given birth to containing 0.01,0.05,0.1mg/L IBA
The earliest rootage duration of adventitious bud cultivated in root culture medium is respectively 15,11,11 days, rooting rate is up to 100%, average root length point
Wei not 4.2cm, 4.8cm, 5.1cm；The adventitious bud root growth cultivated in the culture medium of the IBA containing 0.01-0.05mg/L is fast, root
Sturdy, main root is obvious, fibrous root is more, and the adventitious bud lower end Gen Yujing junctions cultivated in the culture mediums of IBA containing 0.1mg/L are cured
Injured tissue block.
Embodiment 4
In the newborn spray that 5cm or so is gathered after spring resting bud stripping, 6- containing 0.5mg/L is inoculated in after surface sterilization
In BA, 6g/L agar powder, the MS culture mediums of 30g/L sucrose, in 25 ± 2 DEG C, illumination in 14 hours, under the conditions of intensity of illumination 2000lx
Culture, induces axillary bud sprouting.The tender stem that axillary bud sprouting is obtained, is cut into the section of 2 petioles of 2-3cm bands, is seeded in and contains
0.5mg/L 6BA, 0.05mg/L NAA, 0.05mg/L TDZ, 6g/L agar powder, 30g/L sucrose, 0.25g/L MES, 0.2g/L
In the MS culture mediums of l-glutamine, in 25 ± 2 DEG C, illumination in 14 hours is cultivated under the conditions of intensity of illumination 2000l x and bred
Culture.The robust growth obtained will be bred, long 3-4cm simple bud is seeded in IBA containing 0.05mg/L, 7g/L agar powders, 15g/L
In sucrose, 0.25g/L MES 1/2MS culture mediums, in 25 ± 2 DEG C, illumination in 14 hours is taken root under the conditions of intensity of illumination 2000lx
Culture.When root length is to 3-6cm, bottle cap is opened, hardening 3 days.Seedling is gently first pressed from both sides out with tweezers, is gently rinsed in flowing water
Root culture medium, is transplanted to equipped with matrix (perlite: peat soil=1: in disposal plastic cup 1), by every 10 of plastic cup
It is placed in plastics valve bag, room temperature hardening investigates survival rate after 30 days, will be potted plant in perlite+vermiculite+rural area soil in rooted seedling
Grown in the matrix of (ratio 3: 2: 5).As a result show, the tissue-culturing rapid propagation of gold leaf Acer negundo. L is carried out by this program, field collection
After explant inoculation, axillary bud, which starts, to be started after cultivating 15 days；Start the sterilizable material proliferation times average out to 5.8 obtained,
Reach as high as 6.2；Adventitious bud rooting rate is up to 100%, the sturdy prosperity of root system, and transplanting survival rate is up to 78%.
Embodiment 5
In the newborn spray that 5cm or so is gathered after spring resting bud stripping, 6- containing 0.5mg/L is inoculated in after surface sterilization
In BA, 6g/L agar powder, the MS culture mediums of 30g/L sucrose, in 20 ± 2 DEG C, illumination in 14 hours, intensity of illumination 2000l x conditions
Lower culture, induces axillary bud sprouting.The tender stem that axillary bud sprouting is obtained, is cut into the section of 2 petioles of 2-3cm bands, is seeded in and contains
0.5mg/L 6BA, 0.05mg/L NAA, 0.05mg/L TDZ, 6g/L agar powder, 30g/L sucrose, 0.25g/L MES, 0.2g/L
In the MS culture mediums of l-glutamine, in 20 ± 2 DEG C, illumination in 14 hours is cultivated under the conditions of intensity of illumination 2000lx and bred
Culture.The robust growth obtained will be bred, long 3-4cm simple bud is seeded in IBA containing 0.05mg/L, 7g/L agar powders, 15g/L
In sucrose, 0.25g/L MES 1/2MS culture mediums, in 20 ± 2 DEG C, illumination in 14 hours is raw under the conditions of intensity of illumination 2000l x
Root culture.When root length is to 3-6cm, bottle cap is opened, hardening 3 days.Seedling is gently first pressed from both sides out with tweezers, is gently rushed in flowing water
Root culture medium is washed, is transplanted to equipped with matrix (perlite: peat soil=1: in disposal plastic cup 1), by plastic cup every 10
Individual to be placed in plastics valve bag, room temperature hardening investigates survival rate after 30 days, will be potted plant in perlite+vermiculite+rural area in rooted seedling
Grown in the matrix of native (ratio 3: 2: 5).The present embodiment is with embodiment 4 only difference is that cultivation temperature is different.As a result show,
The test tube seedling cultivated under the conditions of 25 ± 2 DEG C, slight mistake occurs in lower plate blade as changed culture medium not in time the phase after incubation
Green albinism, influences transplanting survival rate；Cultivation temperature is down to after 20 ± 2 DEG C, can effectively prevent test tube seedling lower blade chlorosis
Albinism, improves transplanting survival rate, and transplanting survival rate at this temperature is 85%.Two kinds of temperature start time, propagation in bud
Multiple, aspect of performance of taking root influence be not notable.
In summary, by adjusting the ratio of hormone in culture medium, condition of culture is controlled, can significantly shift to an earlier date gold leaf compound leaf
The bud of maple tissue cultures starts the time, improves test tube seedling growth coefficient, increases performance of taking root, and promotes transplanting survival rate, the present invention
Cultural method averagely each explant propagation bud subnumber up to 5.8, adventitious bud rooting rate is up to 100%, and main root is flourishing, fibrous root
Quantity is more, and transplanting survival rate provides an effective way up to 85% for gold leaf Acer negundo. L industrial seedling rearing.
Described above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, under the premise without departing from the principles of the invention, some improvements and modifications can also be made, these improvements and modifications also should
It is considered as protection scope of the present invention.
Claims (6)
1. a kind of tissue culture and rapid propagation method of gold leaf Acer negundo. L, it is characterised in that comprise the following steps：
S1, the newborn spray in collection 5cm or so after spring resting bud stripping, are inoculated in after surface sterilization and with the addition of 0.5-
In the MS culture mediums of 1.0mg/L 6- benzylaminopurines, in 20 ± 2 DEG C -25 ± 2 DEG C, illumination in 14 hours, intensity of illumination
Cultivated under the conditions of 2000lx, the tender stem of induction axillary bud sprouting to 6-8cm length；
S2, the tender stem for taking step S1 to obtain, are cut into the section of 2 petioles of 2-3cm bands, are inoculated in and with the addition of 0.5mg/L 6- benzyls
In adenine phosphate, 0.01-0.05mg/L Thidiazurons, the MS culture mediums of 0.05mg/L methyl α-naphthyl acetates, in 20 ± 2 DEG C of -25 ± 2 DEG C, 14
Under the conditions of hour illumination, intensity of illumination 2000lx, Multiplying culture is carried out；
Robust growth, long 3-4cm simple bud obtained by S3, selecting step S2, which are inoculated in, with the addition of 0.01-0.05mg/L indoles fourths
In the 1/2MS culture mediums of acid, in 20 ± 2 DEG C -25 ± 2 DEG C, illumination in 14 hours under the conditions of intensity of illumination 2000lx, is taken root
Culture.
S4, when root length is to 3-6cm, bottle cap is opened, hardening is gently pressed from both sides out seedling with tweezers, gently rushed in flowing water after 3 days
Root culture medium is washed, is transplanted into the disposal plastic cup equipped with matrix, every 10 are placed in plastics valve bag by plastic cup, room
Warm hardening grows after 30 days by potted plant in rooted seedling in the matrix containing perlite, vermiculite and rural area soil.
2. gold leaf Acer negundo. L tissue culture and rapid propagation method according to claim 1, it is characterised in that the culture in the step S1
6g/L agar powders, 30g/L sucrose are also added with base.
3. gold leaf Acer negundo. L tissue culture and rapid propagation method according to claim 1, it is characterised in that breed training in the step S2
Foster base is also added with 6g/L agar powders, 30g/L sucrose, 0.25g/L MESs, 0.2g/L l-glutamines.
4. gold leaf Acer negundo. L tissue culture and rapid propagation method according to claim 1, it is characterised in that training of being taken root in the step S3
Foster base is also added with 7g/L agar powders, 15g/L sucrose, 0.25g/L MESs.
5. gold leaf Acer negundo. L tissue culture and rapid propagation method according to claim 1, it is characterised in that disposable in the step S4
The matrix of plastic cup is mixed by perlite and peat soil in 1: 1 ratio.
6. gold leaf Acer negundo. L tissue culture and rapid propagation method according to claim 1, it is characterised in that pearl in the step S4
Rock, vermiculite, the ratio of rural area soil are 3: 2: 5.
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| CN106305427B (en) * | 2016-08-30 | 2018-04-10 | 长江三峡生态园林有限公司 | A kind of method of Amur maple tissue-culturing quick-propagation |
| CN107455261B (en) * | 2017-09-15 | 2019-08-13 | 安徽农业大学 | A kind of method of Acer saccharum tissue culture quick breeding |
| CN114711143B (en) * | 2022-04-21 | 2023-08-08 | 东北林业大学 | Method for asexually and rapidly propagating Acer ginnala Maxim seedlings through tissue culture |
| CN116267604B (en) * | 2023-02-18 | 2024-07-23 | 西北农林科技大学 | Method for obtaining acer truncatum aseptic seedlings and directly inducing aseptic buds |
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