CN104719134A - Dendrobenthemia japonica tissue culture method - Google Patents
Dendrobenthemia japonica tissue culture method Download PDFInfo
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Abstract
The invention belongs to the technical field of plant tissue culture and discloses a dendrobenthemia japonica tissue culture method. The dendrobenthemia japonica tissue culture method comprises the steps of pretreating, performing primary culture, enrichment culture and strong seedling culture, planting in a rooting medium and transplanting, wherein in the culturing process, the external conditions are as follows: pH is 5.8-6.0, the culture temperature is 23-25 DEG C, the illumination period is 8-10h/d, and the illumination intensity is 1800-2000lux. The culture method has the benefits that 1. with the adoption of the culture method, a growth coefficient is high, a rooting rate is high, and a cultured plant has excellent growth characteristic; and 2. large scale production can be achieved by the culture method, and the reproduction efficiency is high.
Description
Technical field
The invention belongs to field of plant tissue culture technique, be specifically related to a kind of Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture.
Background technology
Hong Kong Dendronenthamia japonica var.chinensis (Dendrobenthemia japonica), has another name called mountain lichee, rascal tree, for Cornaceae Dendronenthamia japonica var.chinensis belongs to aiphyllium.Sprout and spire are dredged by brown fine, soft fur.Because there being 2 pairs of large bracts of yellow-white petal-shaped outside inflorescence; The ripe redness of fruit or aubergine shape as strawberry and lichee, and obtain famous mountain lichee.Collective fruits is spherical red.Drupe gathers for spherical aggregate fruit meat, and 9 ~ October is ripe, becomes aubergine after maturation, the sweet edible and wine brewing use of fruit taste.
The tree-like rounding of Hong Kong Dendronenthamia japonica var.chinensis, in umbrella shape, blade light, tree performance is graceful.Inflorescence bract is pure white, and infructescence is spherical red gorgeous, is sight appearance, sees leaf, sees flower, sees the excellent ornamental plants really had both at the same time.Large and the pure white covering of petal-shaped bract is completely set, and is lining in bright greenery clump, through wind dynamic dance as group butterfly very unique; Ripening fruits aubergine is quite bright-coloured.When blade is tender, redness becomes brown green autumn in summer, especially gazes in greenery clump.Cold-resistant, drought resisting, there is no damage by disease and insect and impoverishment tolerant, resistance to transplanting.Integrate the excellent landscape planting material seen leaf, see flower, see fruit, especially winter and early spring entirely setting aubergine extremely grand, is the Colored-leaf Plants of one's native land having DEVELOPMENT PROSPECT, is the new select tree kind of landscape planting.
In the Anhui agronomy circular of publication in 2009, Wu Fangxing, Qu Yanchang etc. disclose utilization and the afforestation technique of a kind of Hong Kong Dendronenthamia japonica var.chinensis, first carry out nursery lot selection, about 20d prior to seeding, with the liquor potassic permanganate seed soaking 2h of 0.5%, pull out, rinse 2 times with clear water, then use the water seed soaking of about 50e, after water temperature cools naturally, soak 36h.Pull out and be placed in room temperature 25e vernalization.Want diligent in pregermination procedure and observe, duty stirs, sprays water on a small quantity, keep seed moisture content, about 20d can sow.Control line-spacing and the degree of depth during sowing, water in time, note weeding with a hoe, every 7d sprays the ferrous sulfate 1 time of 012% potassium permanganate and 1% simultaneously, and prevention seedling damping-off, the seedling growth phase can spray carbendazim or leaf spot prevented and treated by zineb.
Equally, in the periodical experience exchangement that the 8th phase in 2006 publishes, Zhang Jianfang also discloses the Cultivating techniques of the beautiful Dendronenthamia japonica var.chinensis in a kind of Hong Kong, by collecting seed with seed treatment, selecting to manage to garden, as after planting will often observed, within continuous 7 days, become a fine day, should water or irrigate once.Emerging when reaching 40% progressively to select the cloudy day to take off grass.Too close if emerged, should timely thinning in 1 month after emerging, keep every per mu yield seedling 2.2-2.5 ten thousand strain.Within emerging latter 1 month, if heat, shade with the sunshade net of 40% light transmittance.After emerging 20 days to September, execute three fertilizer respectively, execute urea 5% the 1st time; 2nd, 3 sealing fertilizers 8% and 10%.In addition, summer plum rain season to note draining and irrigation.The beautiful Dendronenthamia japonica var.chinensis strong stress resistance in Hong Kong, damage by disease and insect is less, and if there is root rot, root is watered in available agricultural streptomycin 500 times of liquid and Tuzet 800 times of liquid mixing.Leaf spot can be sprayed with carbendazim 700 times of liquid, continuous 7 days.
In modern agriculture science and technology disclosed in the 19th phase in 2011, Wu Weihua, Zhong Yaping etc. disclose a kind of excellent greening seedling of one's native land beautiful Hong Kong Dendronenthamia japonica var.chinensis feature and seedling growing process, with above disclosed two technology types seemingly.
Visible, Dendronenthamia japonica var.chinensis existing propagation technique in Hong Kong mainly adopts planting seed nursery, also adopts cuttage, tillering propagation, mostly there is following shortcoming:
Prior art seminal propagation nursery stock can produce variation, can not keep merit and nursery stock irregular, affect seedling quality;
Although prior art cuttage, tillering propagation belong to vegetative propagation can keep merit, reproductive efficiency is low, speed slow, is unfavorable for amount reproduction and the popularization of improved seeds.
Summary of the invention
The object of the invention is to provide that a kind of growth coefficient is high, rooting rate is high, can Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture of a large amount of large-scale production in a short time.
In order to realize object of the present invention, the technical scheme of employing is:
A kind of Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture, is characterized in that, comprise the following steps:
1. pre-treatment: choose Hong Kong Dendronenthamia japonica var.chinensis stem section that growth is more neat, after clean with aseptic water washing, is cut into the segment that 0.7 ~ 0.8cm is long,
2. Initial culture: segment Initial culture base obtained above is carried out Fiber differentiation, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, the Multiplying culture cycle is 40 ~ 45 days, chooses the bud that growth conditions is good, color is dark green and enters next stage;
Wherein Initial culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 3 ~ 4mg/L+NAA 0.1 ~ 0.2mg/L;
3. Multiplying culture: the healthy budlet derived is proceeded in subculture multiplication medium and carries out Multiplying culture, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, and the Multiplying culture cycle is 30 ~ 35 days, when sprouting highly grows to 3.5 ~ 4.0cm, get colors normal, the bud of robust growth starts next stage; The bud highly not reaching 3.5cm takes out separately, again carries out Multiplying culture,
Wherein subculture multiplication medium comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 2 ~ 3mg/L+NAA 0.06 ~ 0.12mg/L,
4. strong seedling culture: the bud after cultivating in proliferated culture medium is carried out cultivation 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C in strong seedling culture base, and periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux,
Wherein strong seedling culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 1 ~ 2mg/L+NAA 0.1 ~ 0.3mg/L,
5. culture of rootage: continue to be inoculated in root media and carry out culture of rootage, cultivate 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux, when root grows to 2.0 ~ 2.5cm, start next stage, rooting rate reaches more than 80%
6. transplant: transplant in being in the nutrition cup of 1:1 containing rice chaff ash and yellow soil volume ratio after cleaning growing tall to 3 ~ 5cm, the cultivation shoot root portion of having taken root, in placement booth, control cultivation temperature 25 ~ 28 DEG C, keep relative moisture 85% ~ 95%, early stage sunshade net shelter from heat or light process 20d, after remove sunshade net gradually, survive rear full exposure.
Wherein root media comprises: female medium+sucrose 20000mg/L+ carragheen 6000 ~ 7000mg/L+ IBA 0.2 ~ 1mg/L+NAA 0.1 ~ 0.3mg/L.
As preferred technical scheme: described female medium composition comprises macroelement, trace element, molysite, Organic additives, and described female medium presses densimeter, comprising:
Macroelement: potassium nitrate 1600 ~ 4000mg/L, ammonium nitrate 1400 ~ 3200mg/L, epsom salt 300 ~ 340mg/L, potassium dihydrogen sulfate 150 ~ 180mg/L, calcium chloride dihydrate 200 ~ 480mg/L;
Trace element: potassium iodide 0.8 ~ 1.0mg/L, four water manganese sulphate 20 ~ 25mg/L, white vitriol, 8 ~ 10mg/L, boric acid 6 ~ 7mg/L, cupric sulfate pentahydrate 0.02 ~ 0.03mg/L, cobalt chloride 0.02 ~ 0.03mg/L, Sodium Molybdate Dihydrate 0.2 ~ 0.3mg/L;
Molysite: ferrous sulfate heptahydrate 25 ~ 30mg/L, disodium ethylene diamine tetraacetate 35 ~ 40mg/L;
Organic principle: thiamine hydrochloride 0.1 ~ 0.3mg/L, pyridoxine hydrochloride 0.4 ~ 0.8mg/L, nicotinic acid 0.4 ~ 0.8mg/L, glycine 1 ~ 3mg/L, inositol 100 ~ 130mg/L.
Wherein, the consisting of of macroelement in described Initial culture medium: potassium nitrate 3500 ~ 4000mg/L, ammonium nitrate 2800 ~ 3200mg/L, epsom salt 300 ~ 340mg/L, potassium dihydrogen sulfate 150 ~ 180mg/L, calcium chloride dihydrate 400 ~ 480mg/L.
The consisting of of macroelement in described subculture multiplication medium and strong seedling culture base: potassium nitrate 1600 ~ 2000mg/L, ammonium nitrate 1400 ~ 2000mg/L, epsom salt 300 ~ 340mg/L, potassium dihydrogen sulfate 150 ~ 180mg/L, calcium chloride dihydrate 400 ~ 480mg/L.
The consisting of of macroelement in described root media: potassium nitrate 1600 ~ 2000mg/L, ammonium nitrate 1400 ~ 2000mg/L, epsom salt 300 ~ 340mg/L, potassium dihydrogen sulfate 150 ~ 180mg/L, calcium chloride dihydrate 200 ~ 280mg/L.
In fact, identical with Initial culture medium of subculture multiplication medium, strong seedling culture base and the trace element in root media, molysite and organic principle, just macroelement content is different, and therefore their minimal medium is all similar.Initial culture medium Main Function is the growth of induced bud, and subculture multiplication medium Main Function is the growth rate stimulating seedling, and strong seedling culture base can make the more healthy and stronger of seedling length, and root media is conducive to the growth of root.
6-BA, chemical name: 6-benzyl aminoadenine, IBA cries again " indolebutyric acid ", and NAA is methyl α-naphthyl acetate.Female medium is similar with MS medium, and the composition contained by both is substantially identical, and difference is the content difference of each composition.And all with Initial culture medium, subculture multiplication medium, strong seedling culture base and root media that female medium is minimal medium, ingredient is also substantially identical, just component content is different.
The beneficial effect that the present invention has:
The incubation of Hong Kong Dendronenthamia japonica var.chinensis tissue is divided into four-stage, and the difference of each growth period characteristic is organized according to Hong Kong Dendronenthamia japonica var.chinensis, correspondence is furnished with a medium, and controls the external condition in incubation well, makes Hong Kong Dendronenthamia japonica var.chinensis show good growth characteristics:
1, adopt this cultural method to cultivate, growth coefficient is high, rooting rate is high, and the plant of cultivation has excellent growth characteristics;
2, cultural method of the present invention can a large amount of large-scale production in a short time, and reproductive efficiency is high.
Embodiment
Below in conjunction with embodiment, the invention will be further described, but protection scope of the present invention is not only confined to embodiment.
Embodiment 1
Wherein Initial culture base comprises: female medium+sucrose 20000mg/L+ carragheen 6500mg/L+6-BA 3.2mg/L+NAA 0.15mg/L;
Wherein subculture multiplication medium comprises: female medium+sucrose 20000mg/L+ carragheen 6500mg/L+6-BA 2.5mg/L+NAA 0.061mg/L,
Wherein strong seedling culture base comprises: female medium+sucrose 20000mg/L+ carragheen 6500mg/L+6-BA 1.5mg/L+NAA 0.2mg/L,
Wherein root media comprises: female medium+sucrose 20000mg/L+ carragheen 6500mg/L+ IBA 0.5mg/L+NAA 0.2mg/L.
In Initial culture medium, macroelement consists of: potassium nitrate 3600mg/L, ammonium nitrate 3000mg/L, epsom salt 320mg/L, potassium dihydrogen sulfate 160mg/L, calcium chloride dihydrate 440mg/L.
The consisting of of macroelement in subculture multiplication medium and strong seedling culture base: potassium nitrate 1800mg/L, ammonium nitrate 1500mg/L, epsom salt 320mg/L, potassium dihydrogen sulfate 160mg/L, calcium chloride dihydrate 440mg/L.
The consisting of of macroelement in root media: potassium nitrate 1800mg/L, ammonium nitrate 1500mg/L, epsom salt 320mg/L, potassium dihydrogen sulfate 160mg/L, calcium chloride dihydrate 240mg/L.
Identical with Initial culture base of subculture multiplication medium, strong seedling culture base and the trace element in root media, molysite and organic principle.Trace element: potassium iodide 0.8mg/L, four water manganese sulphate 22.5mg/L, white vitriol 8.6mg/L, boric acid 6mg/L, cupric sulfate pentahydrate 0.025mg/L, cobalt chloride 0.025mg/L, Sodium Molybdate Dihydrate 0.25mg/L;
Molysite: ferrous sulfate heptahydrate 27.5mg/L, disodium ethylene diamine tetraacetate 37.5mg/L;
Organic principle: thiamine hydrochloride 0.1mg/L, pyridoxine hydrochloride 0.5mg/L, nicotinic acid 0.5mg/L, glycine 2mg/L, inositol 100mg/L.
Embodiment 2
Wherein Initial culture base comprises: female medium+sucrose 25000mg/L+ carragheen 6000mg/L+6-BA 3mg/L+NAA 0.1mg/L;
Wherein subculture multiplication medium comprises: female medium+sucrose 25000mg/L+ carragheen 6000mg/L+6-BA 2mg/L+NAA 0.06mg/L,
Wherein strong seedling culture base comprises: female medium+sucrose 25000mg/L+ carragheen 6000mg/L+6-BA 1mg/L+NAA 0.1mg/L,
Wherein root media comprises: female medium+sucrose 25000mg/L+ carragheen 6000mg/L+IBA 0.2mg/L+NAA 0.1mg/L.
In Initial culture medium, macroelement consists of: potassium nitrate 1600mg/L, ammonium nitrate 1400mg/L, epsom salt 300mg/L, potassium dihydrogen sulfate 150mg/L, calcium chloride dihydrate 200mg/L;
The consisting of of macroelement in subculture multiplication medium and strong seedling culture base: potassium nitrate 2000mg/L, ammonium nitrate 1400mg/L, epsom salt 340mg/L, potassium dihydrogen sulfate 180mg/L, calcium chloride dihydrate 480mg/L.
The consisting of of macroelement in root media: potassium nitrate 2000mg/L, ammonium nitrate 2000mg/L, epsom salt 340mg/L, potassium dihydrogen sulfate 180mg/L, calcium chloride dihydrate 200mg/L.
Identical with Initial culture base of subculture multiplication medium, strong seedling culture base and the trace element in root media, molysite and organic principle.Trace element: potassium iodide 0.8mg/L, four water manganese sulphate 20mg/L, white vitriol 8mg/L, boric acid 6mg/L, cupric sulfate pentahydrate 0.02mg/L, cobalt chloride 0.02mg/L, Sodium Molybdate Dihydrate 0.2mg/L;
Molysite: ferrous sulfate heptahydrate 25mg/L, disodium ethylene diamine tetraacetate 35mg/L;
Organic principle: thiamine hydrochloride 0.1mg/L, pyridoxine hydrochloride 0.4mg/L, nicotinic acid 0.4mg/L, glycine 1mg/L, inositol 100mg/L.
Embodiment 3
Wherein Initial culture base comprises: female medium+sucrose 35000mg/L+ carragheen 7000mg/L+6-BA 4mg/L+NAA 0.2mg/L;
Wherein subculture multiplication medium comprises: female medium+sucrose 35000mg/L+ carragheen 7000mg/L+6-BA 3mg/L+NAA 0.12mg/L,
Wherein strong seedling culture base comprises: female medium+sucrose 35000mg/L+ carragheen 7000mg/L+6-BA 2mg/L+NAA 0.3mg/L,
Wherein root media comprises: female medium+sucrose 35000mg/L+ carragheen 7000mg/L+IBA 1mg/L+NAA 0.3mg/L.
In Initial culture medium, macroelement consists of: potassium nitrate 4000mg/L, ammonium nitrate 3200mg/L, epsom salt 340mg/L, potassium dihydrogen sulfate 180mg/L, calcium chloride dihydrate 480mg/L;
The consisting of of macroelement in subculture multiplication medium and strong seedling culture base: potassium nitrate 3500mg/L, ammonium nitrate 3000mg/L, epsom salt 340mg/L, potassium dihydrogen sulfate 180mg/L, calcium chloride dihydrate 480mg/L.
The consisting of of macroelement in root media: potassium nitrate 3000mg/L, ammonium nitrate 2500mg/L, epsom salt 340mg/L, potassium dihydrogen sulfate 180mg/L, calcium chloride dihydrate 200mg/L.
Identical with Initial culture base of subculture multiplication medium, strong seedling culture base and the trace element in root media, molysite and organic principle.Trace element: potassium iodide 1mg/L, four water manganese sulphate 25mg/L, white vitriol 10mg/L, boric acid 7mg/L, cupric sulfate pentahydrate 0.03mg/L, cobalt chloride 0.03mg/L, Sodium Molybdate Dihydrate 0.3mg/L;
Molysite: ferrous sulfate heptahydrate 30mg/L, disodium ethylene diamine tetraacetate 40mg/L;
Organic principle: thiamine hydrochloride 0.3mg/L, pyridoxine hydrochloride 0.8mg/L, nicotinic acid 0.8mg/L, glycine 3mg/L, inositol 130mg/L.
Embodiment 4
Wherein Initial culture base comprises: female medium+sucrose 30000mg/L+ carragheen 6800mg/L+6-BA 3.4mg/L+NAA 0.25mg/L;
Wherein subculture multiplication medium comprises: female medium+sucrose 30000mg/L+ carragheen 6800mg/L+6-BA 2.5mg/L+NAA 0.25mg/L,
Wherein strong seedling culture base comprises: female medium+sucrose 30000mg/L+ carragheen 6800mg/L+6-BA 0.5mg/L+NAA 0.25mg/L,
Wherein root media comprises: female medium+sucrose 30000mg/L+ carragheen 6800mg/L+IBA 0.6mg/L+NAA 0.2mg/L.
In Initial culture medium, macroelement consists of: potassium nitrate 2800mg/L, ammonium nitrate 2300mg/L, epsom salt 330mg/L, potassium dihydrogen sulfate 170mg/L, calcium chloride dihydrate 460mg/L;
The consisting of of macroelement in subculture multiplication medium and strong seedling culture base: potassium nitrate 2500mg/L, ammonium nitrate 2000mg/L, epsom salt 320mg/L, potassium dihydrogen sulfate 170mg/L, calcium chloride dihydrate 440mg/L.
The consisting of of macroelement in root media: potassium nitrate 2500mg/L, ammonium nitrate 2000mg/L, epsom salt 320mg/L, potassium dihydrogen sulfate 170mg/L, calcium chloride dihydrate 220mg/L.
Identical with Initial culture base of subculture multiplication medium, strong seedling culture base and the trace element in root media, molysite and organic principle.Trace element: potassium iodide 0.9mg/L, four water manganese sulphate 24mg/L, white vitriol 9mg/L, boric acid 6.8mg/L, cupric sulfate pentahydrate 0.03mg/L, cobalt chloride 0.03mg/L, Sodium Molybdate Dihydrate 0.28mg/L;
Molysite: ferrous sulfate heptahydrate 28mg/L, disodium ethylene diamine tetraacetate 38mg/L;
Organic principle: thiamine hydrochloride 0.25mg/L, pyridoxine hydrochloride 0.87mg/L, nicotinic acid 0.7mg/L, glycine 2.5mg/L, inositol 120mg/L.
A kind of Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture, comprises the following steps:
1. pre-treatment: choose Hong Kong Dendronenthamia japonica var.chinensis stem section that growth is more neat, after clean with aseptic water washing, is cut into the segment that 0.7 ~ 0.8cm is long,
2. Initial culture: segment Initial culture base obtained above is carried out Fiber differentiation, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, the Multiplying culture cycle is 40 ~ 45 days, chooses the bud that growth conditions is good, color is dark green and enters next stage;
Wherein Initial culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 3 ~ 4mg/L+NAA 0.1 ~ 0.2mg/L;
Carry out 4 groups of tests simultaneously, select the medium of embodiment 1-embodiment 4 to cultivate respectively, often group test cultivation 20 bottles, 5 every bottle, the culture parameters such as light application time, intensity of illumination, temperature control is completely the same, observes and add up stem section germinating growth situation after 45 days, as shown in table 1 below:
The growing state of table 1 Hong Kong Dendronenthamia japonica var.chinensis bud
As shown in Table 1, by Initial culture medium, induced bud number is many, and the growth conditions of bud is good.
3. Multiplying culture: the healthy budlet derived is proceeded in subculture multiplication medium and carries out Multiplying culture, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, and the Multiplying culture cycle is 30 ~ 35 days, when sprouting highly grows to 3.5 ~ 4.0cm, get colors normal, the bud of robust growth starts next stage; The bud highly not reaching 3.5cm takes out separately, again carries out Multiplying culture,
Wherein subculture multiplication medium comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 2 ~ 3mg/L+NAA 0.06 ~ 0.12mg/L,
Add up after Multiplying culture and measure growth coefficient and growing height, effective seedling number/inoculation seedling number that growth coefficient=bottle seedling is formed in one-period, as shown in table 2 below:
The Multiplying culture situation of table 2 Hong Kong Dendronenthamia japonica var.chinensis bud
As shown in Table 2, after being cultivated by proliferated culture medium, Hong Kong Dendronenthamia japonica var.chinensis well-grown in breeding, growth coefficient very growing height value is all very high.
4. strong seedling culture: the bud after cultivating in proliferated culture medium is carried out cultivation 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C in strong seedling culture base, and periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux,
Wherein strong seedling culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 1 ~ 2mg/L+NAA 0.1 ~ 0.3mg/L,
5. culture of rootage: continue to be inoculated in root media and carry out culture of rootage, cultivate 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux, when root grows to 2.0 ~ 2.5cm, start next stage, rooting rate reaches more than 80%
Wherein root media comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+ IBA 0.2 ~ 1mg/L+NAA 0.1 ~ 0.3mg/L.
Check after culture of rootage and calculate rooting rate and average root is long, explant number × 100% of the explant number/inoculation of wherein rooting rate=take root, the total length/root of average root length=root total
Number; Result is as shown in table 3 below,
The culture of rootage situation of table 3 Hong Kong Dendronenthamia japonica var.chinensis bud
As shown in Table 3, by the cultivation of root media, the bud rooting rate of Hong Kong Dendronenthamia japonica var.chinensis is high, and root is long, has embodied good growth characteristics.
6. transplant: transplant in being in the nutrition cup of 1:1 containing rice chaff ash and yellow soil volume ratio after cleaning growing tall to 3 ~ 5cm, the cultivation shoot root portion of having taken root, in placement booth, control cultivation temperature 25 ~ 28 DEG C, keep relative moisture 85% ~ 95%, early stage sunshade net shelter from heat or light process 20d, after remove sunshade net gradually, survive rear full exposure.
In sum, can greatly improve growth coefficient and rooting rate by cultural method of the present invention, easily a large amount of large-scale production in a short time, reproductive efficiency is high.
Last it is noted that above embodiment only in order to illustrate the present invention and and unrestricted technical scheme described in the invention; Therefore, although this specification with reference to each above-mentioned embodiment to present invention has been detailed description, those of ordinary skill in the art should be appreciated that and still can modify to the present invention or equivalent to replace; And all do not depart from technical scheme and the improvement thereof of the spirit and scope of the present invention, it all should be encompassed in right of the present invention.
Claims (2)
1. Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture, is characterized in that, comprises the following steps:
1. pre-treatment: choose Hong Kong Dendronenthamia japonica var.chinensis stem section that growth is more neat, after clean with aseptic water washing, is cut into the segment that 0.7 ~ 0.8cm is long,
2. Initial culture: segment Initial culture base obtained above is carried out Fiber differentiation, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, the Multiplying culture cycle is 40 ~ 45 days, chooses the bud that growth conditions is good, color is dark green and enters next stage;
Wherein Initial culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 3 ~ 4mg/L+NAA 0.1 ~ 0.2mg/L;
3. Multiplying culture: the healthy budlet derived is proceeded in subculture multiplication medium and carries out Multiplying culture, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, intensity of illumination is 1800 ~ 2000lux, and the Multiplying culture cycle is 30 ~ 35 days, when sprouting highly grows to 3.5 ~ 4.0cm, get colors normal, the bud of robust growth starts next stage; The bud highly not reaching 3.5cm takes out separately, again carries out Multiplying culture;
Wherein subculture multiplication medium comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 2 ~ 3mg/L+NAA 0.06 ~ 0.12mg/L;
4. strong seedling culture: the bud after cultivating in proliferated culture medium is carried out cultivation 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C in strong seedling culture base, and periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux;
Wherein strong seedling culture base comprises: female medium+sucrose 20000 ~ 35000 mg/L+ carragheen 6000 ~ 7000mg/L+6-BA 1 ~ 2mg/L+NAA 0.1 ~ 0.3mg/L,
5. culture of rootage: continue to be inoculated in root media and carry out culture of rootage, cultivate 30 ~ 35 days, PH5.8 ~ 6.0, cultivation temperature 23 ~ 25 DEG C, periodicity of illumination is 8 ~ 10h/d, and intensity of illumination is 1800 ~ 2000lux, when root grows to 2.0 ~ 2.5cm, start next stage, rooting rate reaches more than 80%
Wherein root media comprises: female medium+sucrose 20000mg/L+ carragheen 6000 ~ 7000mg/L+ IBA 0.2 ~ 1mg/L+NAA 0.1 ~ 0.3mg/L,
6. transplant: transplant in being in the nutrition cup of 1:1 containing rice chaff ash and yellow soil volume ratio after cleaning growing tall to 3 ~ 5cm, the cultivation shoot root portion of having taken root, in placement booth, control cultivation temperature 25 ~ 28 DEG C, keep relative moisture 85% ~ 95%, early stage sunshade net shelter from heat or light process 20d, after remove sunshade net gradually, survive rear full exposure.
2. a kind of Hong Kong Dendronenthamia japonica var.chinensis method for tissue culture according to claim, is characterized in that: described female medium composition comprises macroelement, trace element, molysite, Organic additives, and described female medium presses densimeter, comprising:
Macroelement: potassium nitrate 1600 ~ 4000mg/L, ammonium nitrate 1400 ~ 3200mg/L, epsom salt 300 ~ 340mg/L, potassium dihydrogen sulfate 150 ~ 180mg/L, calcium chloride dihydrate 200 ~ 480mg/L;
Trace element: potassium iodide 0.8 ~ 1.0mg/L, four water manganese sulphate 20 ~ 25mg/L, white vitriol, 8 ~ 10mg/L, boric acid 6 ~ 7mg/L, cupric sulfate pentahydrate 0.02 ~ 0.03mg/L, cobalt chloride 0.02 ~ 0.03mg/L, Sodium Molybdate Dihydrate 0.2 ~ 0.3mg/L;
Molysite: ferrous sulfate heptahydrate 25 ~ 30mg/L, disodium ethylene diamine tetraacetate 35 ~ 40mg/L;
Organic principle: thiamine hydrochloride 0.1 ~ 0.3mg/L, pyridoxine hydrochloride 0.4 ~ 0.8mg/L, nicotinic acid 0.4 ~ 0.8mg/L, glycine 1 ~ 3mg/L, inositol 100 ~ 130mg/L.
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| CN104982334A (en) * | 2015-06-26 | 2015-10-21 | 江苏绿苑园林建设有限公司 | Leaf culturing and plant regeneration method for dendrobenthamia hongkongensis |
| CN106879466A (en) * | 2017-03-13 | 2017-06-23 | 玉林师范学院 | A kind of rapid propagation method of the Miaos medicinal materials green grass or young crops pod leaf |
| CN107494266A (en) * | 2017-09-19 | 2017-12-22 | 南京林业大学 | A kind of anti-browning of Tokyo Dendronenthamia japonica var.chinensis and tissue culture enrichment procedure |
| CN109287484A (en) * | 2018-11-02 | 2019-02-01 | 杭州市园林绿化股份有限公司 | Dwarf form bigleaf hydrangea method for tissue culture |
| CN110679483A (en) * | 2019-09-19 | 2020-01-14 | 江苏农林职业技术学院 | Method for inducing Chinese flowering dogwood callus formation |
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| 唐佳佳等: "狭叶四照花茎段的离体培养与植株再生", 《中国农学通报》 * |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104982334A (en) * | 2015-06-26 | 2015-10-21 | 江苏绿苑园林建设有限公司 | Leaf culturing and plant regeneration method for dendrobenthamia hongkongensis |
| CN106879466A (en) * | 2017-03-13 | 2017-06-23 | 玉林师范学院 | A kind of rapid propagation method of the Miaos medicinal materials green grass or young crops pod leaf |
| CN107494266A (en) * | 2017-09-19 | 2017-12-22 | 南京林业大学 | A kind of anti-browning of Tokyo Dendronenthamia japonica var.chinensis and tissue culture enrichment procedure |
| CN107494266B (en) * | 2017-09-19 | 2018-11-02 | 南京林业大学 | A kind of anti-browning of Tokyo Dendronenthamia japonica var.chinensis and tissue culture enrichment procedure |
| CN109287484A (en) * | 2018-11-02 | 2019-02-01 | 杭州市园林绿化股份有限公司 | Dwarf form bigleaf hydrangea method for tissue culture |
| CN110679483A (en) * | 2019-09-19 | 2020-01-14 | 江苏农林职业技术学院 | Method for inducing Chinese flowering dogwood callus formation |
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