CN101796923A - Tissue culture rapid breeding method of ctenanthe oppenheimiana - Google Patents
Tissue culture rapid breeding method of ctenanthe oppenheimiana Download PDFInfo
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- 238000009395 breeding Methods 0.000 title abstract description 10
- 241000358872 Ctenanthe oppenheimiana Species 0.000 title abstract 8
- 241000196324 Embryophyta Species 0.000 claims abstract description 21
- 230000001954 sterilising effect Effects 0.000 claims abstract description 15
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 15
- 238000012258 culturing Methods 0.000 claims abstract description 11
- 239000011159 matrix material Substances 0.000 claims abstract description 8
- 239000002609 medium Substances 0.000 claims description 59
- 235000010804 Maranta arundinacea Nutrition 0.000 claims description 27
- 235000012419 Thalia geniculata Nutrition 0.000 claims description 27
- NWBJYWHLCVSVIJ-UHFFFAOYSA-N N-benzyladenine Chemical compound N=1C=NC=2NC=NC=2C=1NCC1=CC=CC=C1 NWBJYWHLCVSVIJ-UHFFFAOYSA-N 0.000 claims description 24
- 238000005286 illumination Methods 0.000 claims description 21
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 21
- 229920001817 Agar Polymers 0.000 claims description 17
- 239000008272 agar Substances 0.000 claims description 17
- 239000001963 growth medium Substances 0.000 claims description 13
- 238000000034 method Methods 0.000 claims description 12
- 230000035755 proliferation Effects 0.000 claims description 10
- TWFZGCMQGLPBSX-UHFFFAOYSA-N Carbendazim Natural products C1=CC=C2NC(NC(=O)OC)=NC2=C1 TWFZGCMQGLPBSX-UHFFFAOYSA-N 0.000 claims description 7
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 7
- JNPZQRQPIHJYNM-UHFFFAOYSA-N carbendazim Chemical compound C1=C[CH]C2=NC(NC(=O)OC)=NC2=C1 JNPZQRQPIHJYNM-UHFFFAOYSA-N 0.000 claims description 7
- 239000006013 carbendazim Substances 0.000 claims description 7
- 229910052799 carbon Inorganic materials 0.000 claims description 7
- 230000000249 desinfective effect Effects 0.000 claims description 7
- 238000011010 flushing procedure Methods 0.000 claims description 7
- 229960002523 mercuric chloride Drugs 0.000 claims description 7
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims description 7
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 7
- 230000003020 moisturizing effect Effects 0.000 claims description 7
- 239000003415 peat Substances 0.000 claims description 7
- 239000002689 soil Substances 0.000 claims description 7
- 235000014347 soups Nutrition 0.000 claims description 7
- 238000005507 spraying Methods 0.000 claims description 7
- 229910052902 vermiculite Inorganic materials 0.000 claims description 7
- 235000019354 vermiculite Nutrition 0.000 claims description 7
- 239000010455 vermiculite Substances 0.000 claims description 7
- 238000005406 washing Methods 0.000 claims description 7
- 239000005972 6-Benzyladenine Substances 0.000 claims description 5
- 235000021552 granulated sugar Nutrition 0.000 claims description 2
- 244000145580 Thalia geniculata Species 0.000 claims 7
- 230000001488 breeding effect Effects 0.000 abstract description 6
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- 238000011031 large-scale manufacturing process Methods 0.000 abstract 1
- 238000004321 preservation Methods 0.000 abstract 1
- 244000151018 Maranta arundinacea Species 0.000 description 20
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 15
- 229930006000 Sucrose Natural products 0.000 description 15
- 239000005720 sucrose Substances 0.000 description 15
- 244000299452 Gouania lupuloides Species 0.000 description 5
- 235000000292 Gouania lupuloides Nutrition 0.000 description 5
- 241000283986 Lepus Species 0.000 description 5
- 230000004069 differentiation Effects 0.000 description 5
- 241000358866 Ctenanthe burle-marxii Species 0.000 description 2
- 241000234676 Marantaceae Species 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 241000238631 Hexapoda Species 0.000 description 1
- 241000757078 Maranta Species 0.000 description 1
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a tissue culture rapid breeding method of ctenanthe oppenheimiana, which comprises the following steps: taking newly-emerging underground lateral buds of the ctenanthe oppenheimiana as explants; carrying out sterilization on the explants, bud culture starting, bud propagation culture and root striking culture to form complete plantlets; and domesticating and transplanting the complete plantlets into a seedling culture matrix and growing into a normal ctenanthe oppenheimiana plant. For the issue culture rapid breeding method of the ctenanthe oppenheimiana, new bud sprout can be started after underground lateral buds are cultured for 13 days; each underground lateral bud can be differentiated into 2.8 buds; and after culturing the underground lateral buds for 35days, the propagation coefficient reaches 3.2, the rooting rate reaches 92 percent and the survival rate after transplanting reaches 95 percent. By using a tissue culture rapid breeding technology, the invention realizes the preservation of single plants of the ctenanthe oppenheimiana, solves the problems that the ctenanthe oppenheimiana has excessively-long long seedling culture period and low breeding coefficient and can be used for factory large-scale production of seedlings so as to meet the requirement of market on the ctenanthe oppenheimiana.
Description
Technical field
The present invention relates to tissue culture, specifically, relate to the quick breeding method for tissue culture of a kind of comb flower arrowroot (Ctenanthe burle-marxii).
Background technology
Comb flower arrowroot (Ctenanthe burle-marxii) is the colored Maranta plant of Marantaceae comb, originates in Brazil, Costa Rica and the Peru of South America, introduces China in recent years, in North China, there is a small amount of cultivation in areas such as East China and south China.Comb flower arrowroot is the perennial evergreen ornamental foliage plant, plant height 50～80cm, it is rectangular avette that blade is, the blade face light green color, sickle-shaped bottle green speckle is arranged along the vein both sides, the blade back aubergine, the stem branch is strong, cluster, whole strain attitude grace, lucuriant in design is that extremely good indoor ornamental foliage plant and garden beautifies plant.Comb flower arrowroot mainly adopts division propagation, but growing-seedling period is oversize, and reproduction coefficient is low, is difficult to meet the need of market, and spends arrowroot seedling price too high from the external comb of directly buying, and therefore presses for the method that can breed comb flower arrowroot in a large number fast that works out.
Tissue culture is an important channel of breeding the good plant kind fast, existing many genus of the plant of Marantaceae and kind have been carried out the relevant report of tissue culture, but the tissue culture of comb flower arrowroot does not but have relevant report, therefore need the tissue culture of research comb flower arrowroot, and form the special quick propagating technology system of a cover.
Summary of the invention
The objective of the invention is to overcome the deficiencies in the prior art, a kind of tissue culture quick propagation culturing method of comb flower arrowroot is provided.
A kind of tissue culture quick propagation culturing method of comb flower arrowroot comprises the steps:
1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.0～1.5cm, water flushing 20～30min with lateral bud, under aseptic condition with 70% alcohol disinfecting, 20～30s, again with 0.1% mercuric chloride sterilization, 12～16min, aseptic water washing 3～5 times, it is standby to put into bottle;
2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, cultivation cycle is 28～35d, cultivation temperature is 23～27 ℃, and light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), be divided into individual plant and be transferred to and carry out enrichment culture in the bud proliferated culture medium, the proliferation and subculture cycle is 28～35d, and cultivation temperature is 23～27 ℃, light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, cultivation cycle is 30～35d, and cultivation temperature is 23～27 ℃, and light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind 10～15d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 20～30s, transplanting in volume ratio then is in the mixed-matrix of 5: 1 peat soil and vermiculite, spraying and moisturizing 80%～90% shades 75%～85%, is incubated 20～25 ℃ of cultivations.
Described step 2) it is MS+6-BA 3.0～5.0mg/l+NAA 0.1～0.3mg/l that the young shoot in starts medium, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and 6-BA is a 6-benzyladenine, and NAA is a methyl.
Bud proliferated culture medium in the described step 3) is MS+6-BA 2.0～3.5mg/l+NAA 0.2～0.5mg/l, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and 6-BA is a 6-benzyladenine, and NAA is a methyl.
Root media in the described step 4) is 1/2MS+NAA 0.1～0.3mg/l+ active carbon 200～600mg/l, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and the macroelement concentration in this routine minimal medium is reduced by half obtains 1/2MS, and NAA is a methyl.
All add the agar of 6.0～8.0g/l and the white granulated sugar of 20～30g/l in described young shoot startup medium, bud proliferated culture medium, the root media.
The pH value that described bud starts medium, bud proliferated culture medium and root media is 5.6～5.8.
The present invention compared with prior art, the beneficial effect that has: (1) has set up the tissue culture quick propagation culturing method of comb flower arrowroot, compares traditional division propagation method, has greatly improved reproduction coefficient, for the research and production cultivation provides technical support; (2) seedling that obtains by quick breeding by group culture, it is strong to grow, and the leaf look dark green, and uniformity is strong, becomes seedling easy, and adaptability is strong, and damage by disease and insect is few, is easy to management; (3) utilize quick breeding by group culture comb flower arrowroot, removed the restriction of season and weather, thereby shortened growing-seedling period, increased breeding amount; (4) very important meaning is arranged aspect the good mutant or the good first generation of hybrid preserving and breed, but a strain has the comb flower arrowroot mutant of clear superiority or offspring's large tracts of land after tissue culture of a tool clear superiority is promoted.
Embodiment
Following examples are used to illustrate the present invention, but are not used for limiting the scope of the invention.
Described in the present invention MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and 6-BA is a 6-benzyladenine, and NAA is a methyl.
Embodiment 1
(1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.0cm, water flushing 20min with lateral bud, under aseptic condition with 70% alcohol disinfecting 20s, again with 0.1% mercuric chloride sterilization 12min, aseptic water washing 3 times, it is standby to put into bottle;
(2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, this medium is MS+6-BA 3.0mg/l+NAA 0.1mg/l, the agar that adds 6.0g/l in the medium, the sucrose of 20g/l, pH value are 5.6, and cultivation cycle is 28d, cultivation temperature is 23 ℃, light application time is 12h/d, and intensity of illumination is 2000lux, and lateral bud begins to sprout behind the cultivation 16d, induce young shoot at the lateral bud base portion, each explant starts 1.8 young shoots of differentiation.
(3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), being divided into individual plant is transferred to and carries out enrichment culture in the bud proliferated culture medium, this medium is: MS+6-BA 2.0mg/l+NAA 0.2mg/l, the agar that adds 6.0g/l in the medium, the sucrose of 20g/l, the pH value is 5.6, the proliferation and subculture cycle is 28d, cultivation temperature is 23 ℃, and light application time is 12h/d, and intensity of illumination is 2000lux, differentiate the bud of growing thickly behind the enrichment culture 20d, the adventitious buds proliferation coefficient reaches 2.0 times, and average bud height is 2.2cm, and bud seedling robust growth, mounted blade are better.
(4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, this medium is: 1/2MS+NAA 0.1mg/l+ active carbon 200mg/l, the agar that adds 6.0g/l in the medium, the sucrose of 20g/l, pH value are 5.6, and cultivation cycle is 30d, cultivation temperature is 23 ℃, light application time is 12h/d, and intensity of illumination is 2000lux, and bastem portion begins to occur white root original hase projection behind the 12d, grow new root gradually, the seedling base portion grows several complete root systems behind the 23d, forms complete seedling, and rooting rate is 82%, the number of on average taking root is 2.5, and average root is long to be 2.9cm.
(5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind the 10d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 20s, transplanting then in volume ratio is that spraying and moisturizing 80% shades 75% in the mixed-matrix of 5: 1 peat soil and vermiculite, be incubated 20 ℃ of cultivations, survival rate reaches 86%.
Embodiment 2
(1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.2cm, water flushing 23min with lateral bud, under aseptic condition with 70% alcohol disinfecting 24s, again with 0.1% mercuric chloride sterilization 13min, aseptic water washing 3 times, it is standby to put into bottle;
(2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, this medium is MS+6-BA 3.5mg/l+NAA 0.1mg/l, the agar that adds 6.5g/l in the medium, the sucrose of 23g/l, pH value are 5.6, and cultivation cycle is 30d, cultivation temperature is 24 ℃, light application time is 13h/d, and intensity of illumination is 2200lux, and lateral bud begins to sprout behind the cultivation 14d, induce young shoot at the lateral bud base portion, each explant starts 2.6 young shoots of differentiation.
(3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), being divided into individual plant is transferred to and carries out enrichment culture in the bud proliferated culture medium, this medium is: MS+6-BA 2.5mg/l+NAA 0.3mg/l, the agar that adds 6.5g/l in the medium, the sucrose of 23g/l, the pH value is 5.6, the proliferation and subculture cycle is 30d, cultivation temperature is 24 ℃, and light application time is 13h/d, and intensity of illumination is 2200lux, differentiate the bud of growing thickly behind the enrichment culture 17d, the adventitious buds proliferation coefficient reaches 2.5 times, and average bud height is 2.7cm, and bud seedling robust growth, mounted blade are better.
(4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, this medium is: 1/2MS+NAA 0.2mg/l+ active carbon 300mg/l, the agar that adds 6.5g/l in the medium, the sucrose of 23g/l, pH value are 5.6, and cultivation cycle is 32d, cultivation temperature is 24 ℃, light application time is 13h/d, and intensity of illumination is 2200lux, and bastem portion begins to occur white root original hase projection behind the 10d, grow new root gradually, the seedling base portion grows several complete root systems behind the 19d, forms complete seedling, and rooting rate is 85%, the number of on average taking root is 2.8, and average root is long to be 3.5cm.
(5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind the 10d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 24s, transplanting then in volume ratio is that spraying and moisturizing 85% shades 80% in the mixed-matrix of 5: 1 peat soil and vermiculite, be incubated 22 ℃ of cultivations, survival rate reaches 90%.
Embodiment 3
(1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.4cm, water flushing 26min with lateral bud, under aseptic condition with 70% alcohol disinfecting 25s, again with 0.1% mercuric chloride sterilization 14min, aseptic water washing 4 times, it is standby to put into bottle;
(2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, this medium is MS+6-BA 4.0mg/l+NAA 0.2mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 25g/l, pH value are 5.6, and cultivation cycle is 33d, cultivation temperature is 25 ℃, light application time is 15h/d, and intensity of illumination is 2500lux, and lateral bud begins to sprout behind the cultivation 15d, induce young shoot at the lateral bud base portion, each explant starts 2.7 young shoots of differentiation.
(3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), being divided into individual plant is transferred to and carries out enrichment culture in the bud proliferated culture medium, this medium is: MS+6-BA 2.8mg/l+NAA 0.4mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 25g/l, the pH value is 5.6, cultivation cycle is 33d, cultivation temperature is 25 ℃, and light application time is 15h/d, and intensity of illumination is 2500lux, differentiate the bud of growing thickly behind the enrichment culture 15d, the adventitious buds proliferation coefficient reaches 3.0 times, and average bud height is 2.5cm, and bud seedling robust growth, mounted blade are better.
(4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, this medium is: 1/2MS+NAA 0.25mg/l+ active carbon 400mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 25g/l, pH value are 5.6, and cultivation cycle is 33d, cultivation temperature is 25 ℃, light application time is 15h/d, and intensity of illumination is 2500lux, and bastem portion begins to occur white root original hase projection behind the 12d, grow new root gradually, the seedling base portion grows several complete root systems behind the 23d, forms complete seedling, and rooting rate is 92%, the number of on average taking root is 2.3, and average root is long to be 3.7cm.
(5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind the 14d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 26s, transplanting then in volume ratio is that spraying and moisturizing 86% shades 80% in the mixed-matrix of 5: 1 peat soil and vermiculite, be incubated 23 ℃ of cultivations, survival rate reaches 95%.
Embodiment 4
(1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.4cm, water flushing 28min with lateral bud, under aseptic condition with 70% alcohol disinfecting 28s, again with 0.1% mercuric chloride sterilization 15min, aseptic water washing 4 times, it is standby to put into bottle;
(2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, this medium is MS+6-BA 4.5mg/l+NAA 0.2mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 28g/l, pH value are 5.7, and cultivation cycle is 32d, cultivation temperature is 26 ℃, light application time is 15h/d, and intensity of illumination is 2800lux, and lateral bud begins to sprout behind the cultivation 13d, induce young shoot at the lateral bud base portion, each explant starts 2.8 young shoots of differentiation.
(3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), being divided into individual plant is transferred to and carries out enrichment culture in the bud proliferated culture medium, this medium is: MS+6-BA 2.5mg/l+NAA 0.3mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 28g/l, the pH value is 5.7, the proliferation and subculture cycle is 32d, cultivation temperature is 26 ℃, and light application time is 15h/d, and intensity of illumination is 2800lux, differentiate the bud of growing thickly behind the enrichment culture 15d, the adventitious buds proliferation coefficient reaches 3.2 times, and average bud height is 3.0cm, and bud seedling robust growth, mounted blade are better.
(4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, this medium is: 1/2MS+NAA 0.2mg/l+ active carbon 500mg/l, the agar that adds 7.0g/l in the medium, the sucrose of 28g/l, pH value are 5.7, and cultivation cycle is 32d, cultivation temperature is 26 ℃, light application time is 15h/d, and intensity of illumination is 2800lux, and bastem portion begins to occur white root original hase projection behind the 12d, grow new root gradually, the seedling base portion grows several complete root systems behind the 23d, forms complete seedling, and rooting rate is 87%, the number of on average taking root is 2.8, and average root is long to be 3.2cm.
(5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind the 10d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 20s, transplanting then in volume ratio is that spraying and moisturizing 87% shades 80% in the mixed-matrix of 5: 1 peat soil and vermiculite, be incubated 23 ℃ of cultivations, survival rate reaches 93%.
Embodiment 5
(1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.5cm, water flushing 30min with lateral bud, under aseptic condition with 70% alcohol disinfecting 30s, again with 0.1% mercuric chloride sterilization 16min, aseptic water washing 5 times, it is standby to put into bottle;
(2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, this medium is MS+6-BA 5.0mg/l+NAA 0.3mg/l, the agar that adds 8.0g/l in the medium, the sucrose of 30g/l, pH value are 5.8, and cultivation cycle is 35d, cultivation temperature is 27 ℃, light application time is 16h/d, and intensity of illumination is 3000lux, and lateral bud begins to sprout behind the cultivation 18d, induce young shoot at the lateral bud base portion, each explant starts 2.5 young shoots of differentiation.
(3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), being divided into individual plant is transferred to and carries out enrichment culture in the bud proliferated culture medium, this medium is: MS+6-BA 3.5mg/l+NAA 0.5mg/l, the agar that adds 8.0g/l in the medium, the sucrose of 30g/l, the pH value is 5.8, cultivation cycle is 35d, cultivation temperature is 27 ℃, and light application time is 16h/d, and intensity of illumination is 3000lux, differentiate the bud of growing thickly behind the enrichment culture 18d, the adventitious buds proliferation coefficient reaches 2.8 times, and average bud height is 2.8cm, and bud seedling robust growth, mounted blade are better.
(4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, this medium is: 1/2MS+NAA 0.3mg/l+ active carbon 600mg/l, the agar that adds 8.0g/l in the medium, the sucrose of 30g/l, the pH value is 5.8, cultivation cycle is 35d, cultivation temperature is 27 ℃, and light application time is 16h/d, and intensity of illumination is 3000lux, bastem portion begins to occur white root original hase projection behind the 12d, grow new root gradually, the seedling base portion grows several complete root systems behind the 25d, forms complete seedling, rooting rate is 92%, and the number of on average taking root is 2.6, and average root is long to be 3.0cm.
(5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind the 15d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 30s, transplanting then in volume ratio is that spraying and moisturizing 90% shades 85% in the mixed-matrix of 5: 1 peat soil and vermiculite, be incubated 25 ℃ of cultivations, survival rate reaches 88%.
Though, above with a general description of the specific embodiments the present invention has been done detailed description, on basis of the present invention, can to some modifications of do and improvement, this will be apparent to those skilled in the art.Therefore, these modifications or improvements all belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Claims (6)
1. the tissue culture quick propagation culturing method of a comb flower arrowroot is characterized in that comprising the steps:
1) selection of explant and sterilization: getting comb flower arrowroot, neonatally to descend lateral bud be explant, peel off the blade and the leaf sheath of expansion, water is rinsed well, excision base portion lignified tissue is cut into the young shoot of 1.0～1.5cm, water flushing 20～30min with lateral bud, under aseptic condition with 70% alcohol disinfecting, 20～30s, again with 0.1% mercuric chloride sterilization, 12～16min, aseptic water washing 3～5 times, it is standby to put into bottle;
2) young shoot starts cultivation: on the sterile working platform, from bottle, take out the young shoot in the step 1), peel off with soup and contact and injured outermost layer bract, be inoculated on the young shoot startup medium and start cultivation, cultivation cycle is 28～35d, cultivation temperature is 23～27 ℃, and light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
3) young shoot of 1.0～1.5cm downcuts bud enrichment culture: with step 2), be divided into individual plant and be transferred to and carry out enrichment culture in the bud proliferated culture medium, the proliferation and subculture cycle is 28～35d, and cultivation temperature is 23～27 ℃, light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
4) culture of rootage: the bud of growing thickly of 2.5～3.5cm in the step 3) is downcut, be inoculated into and carry out culture of rootage on the root media, cultivation cycle is 30～35d, and cultivation temperature is 23～27 ℃, and light application time is 12～16h/d, and intensity of illumination is 2000～3000lux;
5) domestication of plant and transplanting: the complete seedling that obtains in the step 4) is connected bottle place the hardening of remaining silent under the natural lighting, take out behind 10～15d, clean the medium that adheres to, place 1000 times of carbendazim solutions to soak 20～30s, transplanting in volume ratio then is in the mixed-matrix of 5: 1 peat soil and vermiculite, spraying and moisturizing 80%～90% shades 75%～85%, is incubated 20～25 ℃ of cultivations.
2. a kind of comb flower arrowroot tissue culture quick propagation culturing method according to claim 1, it is characterized in that: it is MS+6-BA 3.0～5.0mg/l+NAA 0.1～0.3mg/l that the young shoot described step 2) starts medium, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and 6-BA is a 6-benzyladenine, and NAA is a methyl.
3. a kind of comb flower arrowroot tissue culture quick propagation culturing method according to claim 1, it is characterized in that: the bud proliferated culture medium in the described step 3) is MS+6-BA 2.0～3.5mg/l+NAA 0.2～0.5mg/l, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and 6-BA is a 6-benzyladenine, and NAA is a methyl.
4. a kind of comb flower arrowroot tissue culture quick propagation culturing method according to claim 1, it is characterized in that: the root media in the described step 4) is 1/2MS+NAA 0.1～0.3mg/l+ active carbon 200～600mg/l, and wherein: MS is conventional minimal medium Murashige﹠amp; Skoog 1962, and the macroelement concentration in this routine minimal medium is reduced by half obtains 1/2MS, and NAA is a methyl.
5. a kind of comb flower arrowroot tissue culture quick propagation culturing method according to claim 1 is characterized in that: all add the agar of 6.0～8.0g/l and the white granulated sugar of 20～30g/l in described young shoot startup medium, bud proliferated culture medium, the root media.
6. a kind of comb flower arrowroot tissue culture quick propagation culturing method according to claim 1, it is characterized in that: the pH value that described bud starts medium, bud proliferated culture medium and root media is 5.6～5.8.
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| CN108812322A (en) * | 2018-07-09 | 2018-11-16 | 苏州枫彩生态科技集团有限公司 | A kind of processing method of emerald green arrowroot explant |
| CN109156363A (en) * | 2018-11-15 | 2019-01-08 | 广州市名卉景观科技发展有限公司 | A kind of rapid propagation method of arrow feather arrowroot |
| CN110476812A (en) * | 2019-08-20 | 2019-11-22 | 广州市名卉景观科技发展有限公司 | A method of for Calathea roseopicta fast breeding |
| CN112352679A (en) * | 2020-11-19 | 2021-02-12 | 广州市卉通农业科技有限公司 | Culture medium for efficient production of double-line arrowroot tissue culture seedlings and preparation method thereof |
| CN113100067A (en) * | 2021-05-14 | 2021-07-13 | 中国科学院华南植物园 | A kind of fast propagation method of arrowroot tissue culture |
| CN114946656A (en) * | 2022-05-17 | 2022-08-30 | 高兰芹 | Method for cultivating high-quality arrowroot |
| CN114946656B (en) * | 2022-05-17 | 2023-11-24 | 高兰芹 | Method for cultivating high-quality arrowroot |
| CN116998406A (en) * | 2023-09-07 | 2023-11-07 | 广州市林业和园林科学研究院 | Rapid propagation seedling method for mosaic arrowroot |
| CN118476472A (en) * | 2024-05-31 | 2024-08-13 | 海南热作两院种业科技有限责任公司 | Tissue culture and rapid propagation method for bamboo potato by taking bamboo potato tubers as explants |
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