CN104604735A - Tissue culture and rapid propagation method for American lagerstroemia indica pink velour - Google Patents
Tissue culture and rapid propagation method for American lagerstroemia indica pink velour Download PDFInfo
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- CN104604735A CN104604735A CN201510106620.8A CN201510106620A CN104604735A CN 104604735 A CN104604735 A CN 104604735A CN 201510106620 A CN201510106620 A CN 201510106620A CN 104604735 A CN104604735 A CN 104604735A
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Abstract
The invention discloses a tissue culture and rapid propagation method for American lagerstroemia indica pink velour and belongs to the technical field of tissue culture of plants. The method carries out in vitro culture for semi-lignified branches of American lagerstroemia indica pink velour and comprises five steps: selection and disinfection of explants, primary culture, multiplication culture, rooting culture and acclimatization and transplant. The method has the advantages of being high in propagation coefficient and short in culture period and is not limited by seasons, and the induction rate is 88 percent. The multiplication coefficient is 5-6, the multiplied buds is fast in elongated growth, and the buds are 3-4cm after being cultured for 25d. The rooting rate is 100 percent, and the average root number is 4.54 pieces/strain. The tissue culture rooting seedlings are healthy and strong, and the survival rate of transplanting is more than 95 percent. In the actual production application, the method can carry out annual large-scale rapid seedling cultivation, produces healthy, strong and uniform American lagerstroemia indica pink velour seedlings, and has a wide application prospect.
Description
Technical field
The invention belongs to the technical field of tissue culture of plant, particularly the tissue culture and rapid propagation method of a kind of U.S. red autumnal leaves crape myrtle.
Background technology
Crape myrtle is Lythraceae (Lythraceae) Lagerstroemia defoliation small arbor or shrub, and have another name called crape myrtle, itch tree, all round victory etc., tree-like grace, bloomed in the hot summer, and pattern is gorgeous, and the florescence is long, is the excellent ornamental plantation seeds of China.U.S. red autumnal leaves crape myrtle (Lagerstroemia indica ' Pink Velour ') is the new quality product kind of crape myrtle introduced in 2003 from the U.S. by forest-science academy of Hunan Province, and its young leaves and the tender tip are claret, have good sight leaf effect; Flower rediance, inflorescence length can reach 70cm, dense and graceful, has high ornamental value.The first tenday period of a month June at florescence, to by the end of October, reach 5 months.Its strong adaptability, drought-enduring, can the low temperature of resistance to-23 DEG C.Be used in the lace in afforestation, flowering shrubs and other plant, also can make stub or potted landscape, be the excellent ornamental plant of afforesting and beautifying environment and floriculture at home, application prospect is huge.
Crape myrtle routine can adopt seminal propagation, but the differentiation of seedling pattern is large, unstable.Current U.S. red autumnal leaves crape myrtle produces upper main by cottage propagation, and cuttings degree of lignification is very large on the impact of red autumnal leaves crape myrtle cuttage root-taking, and adopt the cuttings that degree of lignification is higher, its cuttage survival rate is high.Because U.S. red autumnal leaves crape myrtle is deciduous species, after shoot sprouts, better cutting time is 7 ~ August, and the cuttage production time is limited, and reproductive speed is slow, causes its nursery stock to hold at high price, and constrains the application of its Rapid Popularization.U.S.'s red autumnal leaves crape myrtle group culturation rapid propagating technology system that this research is set up provides effective way for its scale whole year production, has important Theory and applications and is worth.Have not yet to see the tissue-culturing rapid propagation report of U.S. red autumnal leaves crape myrtle.
By literature search, any report about U.S.'s red autumnal leaves crape myrtle tissue-culturing rapid propagation is had not yet to see.Rarely seen a small amount of correlative study to crape myrtle (Lagerstroemia indica) cultured in vitro.If carry out tissue-culturing rapid propagation with reference to existing crape myrtle group training medium to U.S. red autumnal leaves crape myrtle, there will be the jaundice of group training bud and fall leaf, bud is little and weak, and elongation growth is slow, finally occur withered and yellow phenomenon, the situations such as effective bud number is few, cause growth coefficient little, seedling is of poor quality, and reproduction speed is slow, and production cost is high.
Summary of the invention
The object of the invention is to overcome the shortcoming that exists in prior art with not enough, the tissue culture and rapid propagation method of a kind of U.S. red autumnal leaves crape myrtle is provided.The method has that adventitious bud inducing is fast, propagation multiplying power is high, growth is fast, rooting rate is high, the features such as seedling well developed root system is healthy and strong, transplanting survival rate is high, production cost is low of taking root.
Object of the present invention is achieved through the following technical solutions: the tissue culture and rapid propagation method of a kind of U.S. red autumnal leaves crape myrtle, comprises the steps:
(1) explant collection: choose plant type grace, fast growth, without the fine individual plant of damage by disease and insect for maternal, clip trunk semi-lignified branch, cleans up with pure water after defoliation, for subsequent use;
(2) explant sterilization and axillary bud deriving: by branch sterile water wash one time, then use alcoholic solution, mercuric chloride solution soaking disinfection successively, be inoculated into inducing culture with after aseptic water washing, is put into culturing room and cultivates, obtain axillalry bud;
(3) Multiplying culture: the axillalry bud of step (2) is cut and is transferred on proliferated culture medium and cultivates, obtains breeding seedling;
(4) culture of rootage: cut bud and be forwarded to root media and cultivate from the propagation seedling that step (3) obtains, obtains group training and to take root seedling;
(5) hardening and transplanting: step (4) is organized training seedling of taking root and move on to greenhouse and carry out acclimatization and transplants, obtain container seedling.
In step (1), described clip butt is sprouted branch and is adopted semi-lignified branch; Before gathering explant, replace sprinkling sprout branch 4 ~ 5 times every 10d carbendazim, tpn; Branch disleaf after collection, clean up with pure water, low-temperature moisture preservation is for subsequent use.
In step (2):
Described inducing culture is ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/L IBA+30g/L sucrose+8g/L carragheen; PH is 5.8-6.0;
Described ZW medium is containing, for example lower composition: 1200mg/L NH
4nO
3+ 400mg/L KNO
3+ 450mg/L K
2sO
4+ 440mg/L Ca (NO
3)
22H
2o+76mg/L CaCl
24H
2o+260mg/L MgSO
47H
2o+170mg/L KH
2pO
4+ 8.6mg/L ZnSO
47H
2o+6.2mg/L H
3bO
3+ 0.83mg/L KI+22.3mg/L MnSO
44H
2o+0.025mg/L CoCl
2+ 27.8mg/L FeSO
47H
2o+37.3mg/LNa
2eDTAH
2o+100mg/L inositol+4.0mg/L glycine+1.2mg/L thiamine hydrochloride+2.5mg/L nicotinic acid+1.2mg/L puridoxine hydrochloride+2.4mg/L D-VB5 calcium+2.0mg/L ascorbic acid, pH 5.8,121 DEG C of sterilizings 20 minutes.
Described adopting with alcoholic solution, mercuric chloride solution soaking disinfection is carried out with the following method: be 20 ~ 30s by concentration 70 ~ 75% alcoholic solution soak time, to outwell after alcoholic solution aseptic water washing 1 ~ 2 time, then 0.1% mercuric chloride solution is added, soak 3 ~ 10min, constantly shake therebetween, after outwelling mercuric chloride solution, use aseptic water washing 5 ~ 6 times.
Described cultivation is adopted and is carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d; Fiber differentiation 20d, cuts the axillalry bud of length>=1.5cm and is transferred to proliferated culture medium cultivation; Continuous subculture 2 ~ 3 times, axillalry bud is differentiated to form sprouting clump, and the stem section high for bud>=2.0cm edible tender branch being cut into 1.0cm is transferred and to be cultivated into proliferated culture medium;
Described axillalry bud be bud high be the axillalry bud of 1 ~ 2cm.
In step (3),
Described proliferated culture medium is ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/L IBA+30g/L sucrose+8g/L carragheen; PH is 5.8-6.0;
Described cultivation is adopted and is carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d; Continuous subculture 2 ~ 3 times, axillalry bud is differentiated to form bud clump, is cut by the tender stem of high for bud>=1.5cm, access root media.
In step (4),
Described root media is ZW medium+0.1 ~ 0.5mg/L NAA+15g/L sucrose+8g/L carragheen; PH is 5.8-6.0;
Described cultivation is adopted and is carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d;
Described bud is 2cm bud.
In step (5),
Described group trains bottle seedling of taking root that seedling of taking root is culture of rootage 15d.
Described cultivation is adopted and is carried out with the following method: after culture of rootage 15d, a bottle seedling of taking root moves on in greenhouse and adapts to external environment 5 ~ 7d, then by tissue culture bottle lid from partly reaching standard-sized sheet hardening 1d, plantlet in vitro is taken out, clean the medium sticked on root, be transplanted to peat soil: on perlite=3:1 mixed-matrix, before after transplanting, 15d adopts epiphragma moisturizing, then opens film seedling management routinely.
The present invention has following advantage and effect relative to prior art:
(1) the present invention with fast growth, plant type is graceful, pattern is gorgeous, the adult fine individual plant of the U.S. red autumnal leaves crape myrtle of strong stress resistance is maternal, to sprout branch for explant, by the optimization of minimal medium design, hormone kind, concentration and proportioning thereof, set up its efficiently in vitro plant regeneration technique, reach that propagation bud stalwartness is extended, growth is fast, the rate of increase is high, rooting rate is high, seedling stalwartness of taking root, object that transplanting survival rate is high.The present invention has that adventitious bud inducing is fast, bud is sturdy, elongation growth is fast, effectively bud is many; propagation multiplying power is high, rooting rate is high, take root seedling well developed root system stalwartness, transplanting survival rate high; whole technical system is stablized; production cost is low; pass through scale breeding; produce the red autumnal leaves crape myrtle seedling of stalwartness, neat and consistent, for realizing red autumnal leaves crape myrtle Rapid Popularization, meet the demand in market technical support and breeding guarantee are provided, have a extensive future.
(2) U.S. of the present invention red autumnal leaves crape myrtle tissue culture and rapid propagation method, the axillary bud deriving speed of explant is fast; Inductivity is high, reach 88%; Growth coefficient is large, reach 5 ~ 6, and the elongation growth of propagation bud is fast, cultivates 25d bud up to 3 ~ 4cm; Rooting rate can reach 100%, and mean elements is 4.54/strain; Group training takes root seedling stalwartness, transplanting survival rate up to more than 95%.
Accompanying drawing explanation
Fig. 1 is the axillary bud deriving result figure of the sterilizable material of embodiment 1;
Fig. 2 is the propagation clump bud figure of embodiment 1;
Fig. 3 is the propagation bottle seedling figure of embodiment 1;
Fig. 4 is the seedling figure of taking root in the root media of embodiment 1;
Fig. 5 is the shoot root system figure of taking root of embodiment 1;
Fig. 6 is the transplanted seedling growing state figure of embodiment 1.
Embodiment
Below in conjunction with embodiment, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.
Embodiment 1
A tissue culture and rapid propagation method for U.S. red autumnal leaves crape myrtle, comprises the steps:
(1) explant collection: choose plant type grace, fast growth, without the fine individual plant of damage by disease and insect for maternal, the select tree age of tree more than 15 years, about in fine the morning 10 time, clip trunk semi-lignified branch, clean up with pure water after defoliation, for subsequent use;
(2) explant sterilization and axillary bud deriving: on superclean bench, by branch sterile water wash one time, be 20 ~ 30s by concentration 70 ~ 75% alcoholic solution soak time again, to outwell after alcoholic solution aseptic water washing 1 ~ 2 time, then 0.1% mercuric chloride solution is added, soak 3 ~ 10min, constantly shake therebetween, aseptic water washing 5 ~ 6 times are used after outwelling mercuric chloride solution, after aseptic water washing, aseptic filter paper blots sterilizable material surface water drops, stem section being cut into 2cm is with axil cut and be inoculated into inducing culture (inducing culture: ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/L IBA+30g/L sucrose+8g/L carragheen, pH is 5.8-6.0 (hormone is purchased from Sigma-Aldrich company)), be put into culturing room, in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d, Fiber differentiation 15d, axillalry bud length being greater than 2cm cuts and is transferred to proliferated culture medium cultivation, cultivate 30d, axillalry bud is differentiated to form sprouting clump, and the stem section high for bud>=3cm edible tender branch being cut into 1.0 ~ 1.5cm is transferred and to be cultivated into proliferated culture medium, every bottle graft kind explant, after 5d, petiole starts loose or dislocation, and axillalry bud is sprouted, and after 15d, axillalry bud can grow to about 2cm, and blade is carried out, leaf color jade green (Fig. 1), and inductivity is 88%.
(3) Multiplying culture: the axillalry bud induced of step (2) be cut into the stem section of 1.0 ~ 1.5cm and be transferred to proliferated culture medium (ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/L IBA+30g/L sucrose+8g/L carragheen; PH is 5.8-6.0) on cultivate, temperature controls at 25 ± 2 DEG C, illumination (60 μm of ol.m
-2.s
-1) time 12h/d; After subculture 3 ~ 4 times, simple bud forms bud clump, and bud elongation growth is fast, and growth coefficient and effective bud (bud high>=2cm) are respectively up to 5.21 and 6.2/clump (Fig. 2, Fig. 3).
(4) culture of rootage: cut 2.0cm bud and be forwarded to root media (ZW medium+0.1 ~ 0.5mg/L NAA+15g/L sucrose+8g/L carragheen from the propagation seedling that step (3) obtains; PH is 5.8-6.0) on cultivate, temperature controls at 25 ± 2 DEG C, illumination (60 μm of ol.m
-2.s
-1) time 12h/d, after 15d, the inductivity of adventive root is 100% (Fig. 4).Robust plant, leaf color jade green, root system is sturdy, mean elements 4.54/strain (Fig. 5).
(5) hardening and transplanting: step (4) is organized training seedling of taking root and move on to greenhouse and carry out acclimatization and transplants, obtain container seedling.A bottle seedling of taking root of culture of rootage 15d is moved on in greenhouse and adapts to external environment 5 ~ 7d, then by tissue culture bottle lid from partly reaching standard-sized sheet hardening 1d, plantlet in vitro is carefully taken out, clean the medium sticked on root, be transplanted to peat soil: on perlite=3:1 mixed-matrix, before after transplanting, 15d adopts epiphragma moisturizing, and then open film seedling management routinely, this transplanting survival rate is higher than 95% (Fig. 6).
In step (1), described clip butt is sprouted branch and is adopted semi-lignified branch; Before gathering explant, replace sprinkling sprout branch 4 ~ 5 times every 10d carbendazim, tpn; Branch disleaf after collection, clean up with pure water, low-temperature moisture preservation is for subsequent use.
Described ZW medium is containing, for example lower composition: 1200mg/L NH
4nO
3+ 400mg/L KNO
3+ 450mg/L K
2sO
4+ 440mg/L Ca (NO
3)
22H
2o+76mg/L CaCl
24H
2o+260mg/L MgSO
47H
2o+170mg/L KH
2pO
4+ 8.6mg/L ZnSO
47H
2o+6.2mg/L H
3bO
3+ 0.83mg/L KI+22.3mg/L MnSO
44H
2o+0.025mg/L CoCl
2+ 27.8mg/L FeSO
47H
2o+37.3mg/LNa
2eDTAH
2o+100mg/L inositol+4.0mg/L glycine+1.2mg/L thiamine hydrochloride+2.5mg/L nicotinic acid+1.2mg/L puridoxine hydrochloride+2.4mg/L D-VB5 calcium+2.0mg/L ascorbic acid, pH 5.8,121 DEG C of sterilizings 20 minutes.
The present invention with fast growth, plant type is graceful, pattern is gorgeous, the adult fine individual plant of the U.S. red autumnal leaves crape myrtle of strong stress resistance is maternal, to sprout branch for explant, by the optimization of minimal medium design, hormone kind, concentration and proportioning thereof, set up its efficiently in vitro plant regeneration technique, reach that propagation bud stalwartness is extended, growth is fast, the rate of increase is high, rooting rate is high, seedling stalwartness of taking root, object that transplanting survival rate is high.The present invention has that adventitious bud inducing is fast, bud is sturdy, elongation growth is fast, effectively bud is many; propagation multiplying power is high, rooting rate is high, take root seedling well developed root system stalwartness, transplanting survival rate high; whole technical system is stablized; production cost is low; pass through scale breeding; produce the red autumnal leaves crape myrtle seedling of stalwartness, neat and consistent, for realizing red autumnal leaves crape myrtle Rapid Popularization, meet the demand in market technical support and breeding guarantee are provided, have a extensive future.
In this method, the axillary bud deriving speed of explant is fast; Inductivity is high, reach 88%; Growth coefficient is large, reach 5 ~ 6, and the elongation growth of propagation bud is fast, cultivates 25d bud up to 3 ~ 4cm; Rooting rate can reach 100%, and mean elements is 4.54/strain; Group training takes root seedling stalwartness, transplanting survival rate up to more than 95%.
Above-described embodiment is the present invention's preferably embodiment; but embodiments of the present invention are not restricted to the described embodiments; change, the modification done under other any does not deviate from Spirit Essence of the present invention and principle, substitute, combine, simplify; all should be the substitute mode of equivalence, be included within protection scope of the present invention.
Claims (10)
1. a tissue culture and rapid propagation method for U.S.'s red autumnal leaves crape myrtle, is characterized in that, comprises the steps:
(1) explant collection: choose plant type grace, fast growth, without the fine individual plant of damage by disease and insect for maternal, clip trunk semi-lignified branch, cleans up with pure water after defoliation, for subsequent use;
(2) explant sterilization and axillary bud deriving: by branch sterile water wash one time, then use alcoholic solution, mercuric chloride solution soaking disinfection successively, be inoculated into inducing culture with after aseptic water washing, is put into culturing room and cultivates, obtain axillalry bud;
(3) Multiplying culture: the axillalry bud of step (2) is cut and is transferred on proliferated culture medium and cultivates, obtains breeding seedling;
(4) culture of rootage: cut bud and be forwarded to root media and cultivate from the propagation seedling that step (3) obtains, obtains group training and to take root seedling;
(5) hardening and transplanting: step (4) is organized training seedling of taking root and move on to greenhouse and carry out acclimatization and transplants, obtain container seedling.
2. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, it is characterized in that, the inducing culture described in step (2) is ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/LIBA+30g/L sucrose+8g/L carragheen; PH is 5.8-6.0.
3. the tissue culture and rapid propagation method of the U.S. according to claim 2 red autumnal leaves crape myrtle, is characterized in that, described ZW medium is containing, for example lower composition: 1200mg/L NH
4nO
3+ 400mg/L KNO
3+ 450mg/LK
2sO
4+ 440mg/L Ca (NO
3)
22H
2o+76mg/L CaCl
24H
2o+260mg/L MgSO
47H
2o+170mg/L KH
2pO
4+ 8.6mg/L ZnSO
47H
2o+6.2mg/L H
3bO
3+ 0.83mg/L KI+22.3mg/L MnSO
44H
2o+0.025mg/L CoCl
2+ 27.8mg/L FeSO
47H
2o+37.3mg/LNa
2eDTAH
2o+100mg/L inositol+4.0mg/L glycine+1.2mg/L thiamine hydrochloride+2.5mg/L nicotinic acid+1.2mg/L puridoxine hydrochloride+2.4mg/L D-VB5 calcium+2.0mg/L ascorbic acid, pH 5.8,121 DEG C of sterilizings 20 minutes.
4. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, it is characterized in that, adopting with alcoholic solution, mercuric chloride solution soaking disinfection described in step (2) is carried out with the following method: be 20 ~ 30s by concentration 70 ~ 75% alcoholic solution soak time, to outwell after alcoholic solution aseptic water washing 1 ~ 2 time, then 0.1% mercuric chloride solution is added, soak 3 ~ 10min, constantly shake therebetween, after outwelling mercuric chloride solution, use aseptic water washing 5 ~ 6 times.
5. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, is characterized in that, the cultivation described in step (2) is adopted and carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d; Fiber differentiation 20d, cuts the axillalry bud of length>=1.5cm and is transferred to proliferated culture medium cultivation; Continuous subculture 2 ~ 3 times, axillalry bud is differentiated to form sprouting clump, and the stem section high for bud>=2.0cm edible tender branch being cut into 1.0cm is transferred and to be cultivated into proliferated culture medium.
6. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, is characterized in that, the axillalry bud described in step (2) be bud high be the axillalry bud of 1 ~ 2cm.
7. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, it is characterized in that, the proliferated culture medium described in step (3) is ZW medium+0.5 ~ 1.5mg/L 6-BA+0.05 ~ 0.2mg/LIBA+30g/L sucrose+8g/L carragheen; PH is 5.8-6.0; Described cultivation is adopted and is carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d; Continuous subculture 2 ~ 3 times, axillalry bud is differentiated to form bud clump, is cut by the tender stem of high for bud>=1.5cm, access root media.
8. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, is characterized in that, the root media described in step (4) is ZW medium+0.1 ~ 0.5mg/L NAA+15g/L sucrose+8g/L carragheen; PH is 5.8-6.0.
9. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, is characterized in that, the cultivation described in step (4) is adopted and carried out with the following method: in 25 ± 2 DEG C, 60 μm ol.m
-2.s
-1illumination 12h/d.
10. the tissue culture and rapid propagation method of the U.S. according to claim 1 red autumnal leaves crape myrtle, it is characterized in that, cultivation described in step (5) is adopted and is carried out with the following method: after culture of rootage 15d, a bottle seedling of taking root moves on in greenhouse and adapts to external environment 5 ~ 7d, then by tissue culture bottle lid from partly reaching standard-sized sheet hardening 1d, plantlet in vitro is taken out, clean the medium sticked on root, be transplanted to peat soil: on perlite=3:1 mixed-matrix, before after transplanting, 15d adopts epiphragma moisturizing, then opens film seedling management routinely.
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| CN109287483A (en) * | 2018-11-02 | 2019-02-01 | 杭州市园林绿化股份有限公司 | A kind of method of bigleaf hydrangea container seedling quickly breeding |
| CN109287483B (en) * | 2018-11-02 | 2022-03-25 | 杭州市园林绿化股份有限公司 | Method for quickly cultivating hydrangea macrophylla container seedlings |
| CN113892430A (en) * | 2021-11-15 | 2022-01-07 | 湖北格林凯尔农业科技有限公司 | Tissue culture and rapid propagation method of pinus koraiensis |
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