CN101167441B - Calamus simplicifolius clone tissue culture and fast propagation method - Google Patents

Calamus simplicifolius clone tissue culture and fast propagation method Download PDF

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CN101167441B
CN101167441B CN2007100317810A CN200710031781A CN101167441B CN 101167441 B CN101167441 B CN 101167441B CN 2007100317810 A CN2007100317810 A CN 2007100317810A CN 200710031781 A CN200710031781 A CN 200710031781A CN 101167441 B CN101167441 B CN 101167441B
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culture
bud
rootage
explant
seedling
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CN101167441A (en
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曾炳山
尹光天
李湘阳
刘�英
裘珍飞
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Research Institute of Tropical Forestry of Chinese Academy of Forestry
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Research Institute of Tropical Forestry of Chinese Academy of Forestry
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    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
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Abstract

The invention relates to a group-cultivation quick-propagation method for single-leaf calami, which is characterized in that asexual organs such as sprouting buds and the like of single-leaf calami are utilized for group cultivation and quick propagation. The specific procedure is as follows, first, sprouting buds are sawed from a stock plant with a saw in sunny days to prevent bud bodies from being damaged and the bud cores from being shattered; second, the explants of the sprouting buds are sterilized; third, anti-browning process is done to the explants of the sprouting buds to prevent browning; fourth, clumpy buds are formed by induction on a clumpy buds induction medium, after the buds are formed, the clumpy buds are transferred to a breeding culture medium for breeding and propagation; fifth, bud seedlings meeting the height requirement are cultivated on a radication culture medium for radication, bud seedlings not meeting the height requirement are first cultivated for reinforcement to grow to the required height and then are cultivated for radication; sixth, seedlings are transplanted to fields, after transplant, bactericidal agent is sprayed to the seedlings and shading nets are utilized to shade the seedlings until the seedlings are planted out of gardens. The invention solves the technical problem of group cultivation and quick propagation of the single-leaf calami clone and is applied in the breeding of the single-leaf calami clone and the quick propagation of elite seedlings, which not only promotes the development of artificial plant production of rattans and helps to achieve good economic benefit.

Description

The calamus simplicifolius clone tissue culture and fast propagation method
Technical field
The present invention relates to a kind of calamus simplicifolius clone tissue culture and fast propagation method, be applicable to that Palmae Calamus plant tissue culture is numerous soon.Belong to clonal line tissue culture technology field.
Background technology
It is that Palmae (Palmae) is economized rattan subfamily (Calamoideae) and economized the tool of rattan family (Calameae) Calamus (Calamus) liane of sting, climb, grow thickly that single leaf is economized rattan (Calamus simplicifolius Wei.), is the torrid zone, south China, the distinctive accompanying plant of the regional forest of south subtropics.Rattan has good operational characteristic, is the good raw material of making furniture and handicraft.The tender tip of rattan is rich in the multiple nutritional components of needed by human body, and it is edible to do vegetables.The seed quality is hard, is the traditional material of making " beads ".Therefore, it is the precious forest plants of multipurpose with higher economic worth and DEVELOPMENT PROSPECT that single leaf is economized rattan, also is the extensively staple rattan kind of cultivation of South China.At present, provinces and regions such as Hainan, Guangdong, Guangxi, Fujian the single leaf of commerial growing economize rattan.
In the prior art, single leaf is economized rattan and is mainly adopted seminal propagation, and the control of relevant seed storage, sowing, container nursery, nursery stock damage by disease and insect etc. all has successfully be used to the to afforest cultivation of nursery stock of detailed research report, seedling growing process.But single leaf is economized rattan seed storage tolerance not under conventional room temperature condition, even preserved six months under the optimum storaging condition of constant temperature (15 ℃), constant humidity (the media water-bearing rate remains on 50%～65%), germination rate also only can maintain about 65%.Be annual October～December the suitableeest seed collecting season that single leaf is economized rattan, owing to this in season humidity little, the easy dehydration of seed under the room temperature condition, thus lose germinating capacity rapidly, increased new difficulty to seminal propagation.
And seminal propagation can only be bred conventional nursery stock, irreproducible clone nursery stock.Because it is cross-pollinatd plant that single leaf is economized rattan, and the domestication degree is not high, so the single leaf in natural forest and the existing forest plantation is economized the heterozygosis degree height of rattan, after the nursery stock afforestation of seminal propagation, differentiation is big at aspects such as growth, sprout tillers, wood property, both be unfavorable for the raising of average yield, be unfavorable for also that the intensive management of rattan woods and rattan gathered and sell.
Because forest genome heterozygosis degree height, the seedling forestation differentiation is big, and the road of clone breeding is normally walked in forest genetics, promptly selects select tree from breeding population, then by vegetative method reproduction improved variety.Existing can only be that the tissue culture technology of explant still can't satisfy the requirement that clone breeding and clone are promoted with the embryo.If do not solve the technical problem of calamus simplicifolius clone tissue culture and fast propagation, the pilot scale of calamus simplicifolius clone breeding, breeding and the popularization of breeding will be restricted seriously.
Last century, the nineties began, and begin one's study single leaf of Tropical Foresty Inst., Chinese Academy of Foresty Sciences is economized the group culturation rapid propagating technology of rattan.But babassu tissue culture technology research difficulty is big, through the research of two five-year plans, also can only do the success of explant ability tissue-culturing rapid propagation with embryo, and do explant with no sexual organs such as terminal bud, coppice spout, root, leaves, then can not the tissue-culturing rapid propagation success.Thereby original rattan tissue culture technology only is confined to sapling multiplication, promptly by tissue culture industrial fast breeding nursery stock, but still irreproducible clone nursery stock.Because original tissue culture technology is done explant with embryo, the hereditary basis of propagating materials can't be controlled fully, the nursery stock of breeding is compared with the nursery stock of seminal propagation does not have quality differential, but the cost of group training breeding nursery stock is than seed growing nursery stock height, be difficult on market, compete, thereby the application prospect of original tissue culture technology with kind of embryo explants is little with the nursery stock of seed growing.
Summary of the invention
Purpose of the present invention is for after the nursery stock afforestation that overcomes seminal propagation, in the big shortcoming of aspects such as growth, sprout tillers, wood property differentiation, provides a kind of calamus simplicifolius clone tissue culture and fast propagation method.
Purpose of the present invention can reach by taking following measure:
A kind of calamus simplicifolius clone tissue culture and fast propagation method is characterized in that: utilize single leaf to economize no sexual organ such as rattan coppice spout and carry out tissue-culturing rapid propagation, concrete steps are as follows:
1) gathers the coppice spout explant
Continuous sunny after 3～5 days with saw with coppice spout under maternal plant saw; In the process of saw coppice spout explant, must prevent that the sprout wound and the bud heart from shattering;
2) sterilization of explant
Adopt sterilization twice, the first step at first cleans up explant, peels off 2～3 layers of leaf sheath, with 75% alcohol disinfecting 20～25 seconds, then with 0.1% mercury chloride sterilization 10～12 minutes, with rinsed with sterile water 1 time 10 minutes; Second step, peel off 1～2 layer of leaf sheath again and be trimmed to 3～4cm length, with 0.1% mercury chloride sterilization 1～2 minute, use rinsed with sterile water again 3 times, each time was respectively 5 minutes, and 5 minutes, 10 minutes;
3) prevent brown stain
Adopt cysteine or ascorbic acid or the two to handle in conjunction with the coppice spout explant being carried out anti-brown stain, the concentration of cysteine and ascorbic acid is respectively 3mg/l and 10mg/l;
4) the clump bud is induced and enrichment culture
After explant collection and the sterilization, at first induce to form the clump bud on clump bud inducing culture, induction duration is 3～5 months, needs switching midway and changes fresh culture 2～3 times; After forming the clump bud, be transferred to and carry out enrichment culture on the proliferated culture medium, the cycle that one-step growth is cultivated is 45～50 days; Big clump bud is cut into the budlet clump that 3～4 buds are formed, is transferred to and carries out enrichment culture next time on the fresh proliferated culture medium, clone nursery stock can be expanded numerous through enrichment culture repeatedly.
5) culture of rootage
Height is carried out culture of rootage more than or equal to the bud seedling of 3.0cm at root media, and the time of culture of rootage can be 50～60 days, and one time rooting rate can reach more than 85%; The bud seedling of not taking root after removing 1～2 layer of outer leaf sheath and base portion pruned, is carried out the culture of rootage second time, can take root; To the bud seedling of height less than 3.0cm, carry out strong seedling culture earlier, allow the bud seedling grow to 3.0cm or above height, carry out culture of rootage again;
6) transplant in the field
The height greater than 8.0cm, root length greater than 4.0cm, have 3 the most suitable transplanting of nursery stock with blade.The medium that suitable single leaf is economized the transplanting of rattan tissue cultivating seedling has fine sand, peat soil and yellow soil, also the three can be mixed in 1: 1: 2 ratio, economizes the transplanting medium of rattan tissue cultivating seedling as single leaf.Transplant medium with 1/1000 disinfecting solution of potassium permanganate, transplant and spray in turn with 1/800 tpn and carbendazim in each week of back, the bacterium that prevents to mix grows.Transplant and began fertilising in back 15 days, adopt 1/1000 spray once in per 1 week of composite fertilizer's solution.Transplanted the back 1～7 day, 95% shading screen shades, and 8～30 days, 85% shading screen shaded, and 31～60 days, 70% shading screen shaded, and adopted 50% shading screen to shade subsequently, manages outplanting afforestation always.Transplanting success is greater than 85%.
Purpose of the present invention can also reach by taking following measure:
One embodiment of the present invention are: the 1st) height of step collection coppice spout explant can be 25～45cm.
One embodiment of the present invention are: the 4th) step clump bud induce with enrichment culture after, when the growth of bud seedling is high less than the needed 3.0cm of taking root, carry out strong seedling culture at the strong seedling culture base earlier, allow the bud seedling grow into the height that is fit to culture of rootage.
One embodiment of the present invention are: carrying out the 4th) when step clump bud is induced, bud is tilted or keep flat, make it whole contact medium, to promote the growth of bud, promote the sprouting of secondary tiller bud, promote propagation.
One embodiment of the present invention are: carrying out the 5th) when going on foot culture of rootage, 2～4 bud cuttings that can bud height of seedling degree and size is approaching are transferred to and carry out culture of rootage on the root media same budlet clump, can cultivate clump bud seedling after the transplanting.
One embodiment of the present invention are: can transplant single leaf in 3～November and economize the rattan tissue cultivating seedling, wherein be the best seasonal migration that single leaf is economized the rattan tissue cultivating seedling 3～June, and temperature, humidity all be fit to the further cultivation of nursery stock, reaches afforestation height and requirement.
One embodiment of the present invention are: clump bud inducing culture is made up of macroelement, trace element, organic principle, plant hormone and sucrose, carragheen, and macroelement comprises NH 4NO 3, KNO 3, KH 2PO 4, MgSO 4, CaCl 2, trace element comprises MnSO 44H 2O, ZnSO 47H 2O, H 3BO 3, Na 2MoO 42H 2O, FeSO 47H 2O, Na 2EDTA, Kl, CuSO 45H 2O, CoCl 26H 2O, organic principle comprise amion acetic acid, thiamine hydrochloride, puridoxine hydrochloride, nicotinic acid, inositol, vitamin C and cysteine, and plant hormone comprises 6-BA, IBA, NAA, KT and 2,4-D; The content of each component is referring to table 1, and the pH value of clump bud inducing culture is 5.8.
One embodiment of the present invention are: proliferated culture medium is made up of macroelement, trace element, organic principle, plant hormone and sucrose, carragheen, and macroelement comprises NH 4NO 3, KNO 3, KH 2PO 4, MgSO 4, CaCl 2, trace element comprises MnSO 44H 2O, ZnSO 47H 2O, H 3BO 3, Na 2MoO 42H 2O, FeSO 47H 2O, Na 2EDTA, Kl, CuSO 45H 2O, CoCl 26H 2O, organic principle comprise amion acetic acid, thiamine hydrochloride, puridoxine hydrochloride, nicotinic acid, inositol, vitamin C and cysteine, and plant hormone comprises 6-BA, IBA, NAA, KT and 2,4-D; The content of each component is referring to table 1, and the pH value of proliferated culture medium is 5.8.
One embodiment of the present invention are: the strong seedling culture base is made up of macroelement, trace element, organic principle and sucrose, carragheen, and macroelement comprises NH 4NO 3, KNO 3, KH 2PO 4, MgSO 4, CaCl 2, trace element comprises MnSO 44H 2O, ZnSO 47H 2O, H 3BO 3, Na 2MoO 42H 2O, FeSO 47H 2O, Na 2EDTA, Kl, CuSO 45H 2O, CoCl 26H 2O, organic principle comprise amion acetic acid, thiamine hydrochloride, puridoxine hydrochloride, nicotinic acid, inositol, vitamin C and cysteine; The content of each component is referring to table 1, and the pH value of strong seedling culture base is 5.8.
One embodiment of the present invention are: root media is made up of macroelement, trace element, organic principle, plant hormone and sucrose, carragheen, and macroelement comprises NH 4NO 3, KNO 3, KH 2PO 4, MgSO 4, CaCl 2, trace element comprises MnSO 44H 2O, ZnSO 47H 2O, H 3BO 3, Na 2MoO 42H 2O, FeSO 47H 2O, Na 2EDTA, Kl, CuSO 45H 2O, CoCl 26H 2O, organic principle comprise amion acetic acid, thiamine hydrochloride, puridoxine hydrochloride, nicotinic acid and inositol, and plant hormone comprises IBA and NAA; The content of each component is referring to table 1, and the pH value of root media is 5.8.
The present invention has following beneficial effect:
1, the present invention utilizes no sexual organs such as coppice spout to carry out tissue-culturing rapid propagation, solve the technical barrier of calamus simplicifolius clone tissue culture and fast propagation, enlarge single leaf and economize the range of application of rattan tissue culture technology, make it can be used for single leaf and economize the numerous soon of the clone breeding of rattan and breeding nursery stock, economize the numerous soon of rattan choiceness nursery stock by single leaf, both can promote the development of palm rattan artificial planting industry, can obtain good economic benefit again.
2, compare with original seed seedling-raising, the present invention has the advantage that sapling multiplication speed is fast, seedling production is not vulnerable to seasonal effect.With what delivered is that the tissue culture technology of explant is compared with the embryo, and the present invention has the advantage that can breed choiceness.This method is done explant with coppice spout, can breed the choiceness through seed selection and mensuration, production be the nursery stock of breeding, have the powerful market competitiveness, thereby present technique has broad application prospects, can produce huge economic benefit.
Embodiment
Describe the present invention below in conjunction with specific embodiment.
The calamus simplicifolius clone tissue culture and fast propagation that the present invention relates to, concrete grammar can carry out as follows:
1) coppice spout explant collection
The key of gathering the coppice spout explant is to prevent shattering of the sprout wound and the bud heart.If sprout has wound, earth enters sprout inside easily, the success of can't sterilizing.After the bud heart shattered, the wound was rapid brown stain, easily causes the death of the bud heart.Thereby, can not coppice spout be dug from maternal plant with hoe, can not cut with cutter, must coppice spout be sawed from maternal plant with saw.Coppice spout is among the soil, must avoid gathering in 1～3 day behind rainy day and rain as far as possible.Otherwise pollution rate obviously rises.
The size of suitable coppice spout explant is 25～45cm height.Coppice spout is too small, and explant sterilization back is easily dead, and does not form enough secondary coppice spout original hases, occurs the high growth of simple bud that continues after the cultured in vitro easily but does not sprout the phenomenon of secondary coppice spout.Coppice spout is excessive, and behind the especially super 60cm, coppice spout has begun to put forth, and stem apex is not at base portion.Keep stem apex, then explant is excessive, and the Nutrient Absorption supply is not gone up, and stem apex can't be grown, and the part brown stain of exposing outside medium is serious, finally causes the death of whole bud.Do not keep stem apex, explant is stem apex for want of, and its secondary tiller bud is difficult to sprout.
Explant collection season, growth and the inducing of bud of clump to sterilization success rate, bud all had remarkable influence.The explant that each month from February to September gathers shows: the explant of gathering with September at the beginning of 2～3 months, because of antecedent soil moisture, to sterilize successfully easily, and 4～August, rainwater was more, and temperature is higher, and the explant of gathering behind the rain is difficult to sterilize successfully.Do not grow for a long time on medium but 2～March and September gather explant, be difficult to the induced bundle bud.Although what gathered 4～August is difficult to sterilize successfully, explant is sprouted easily, and the secondary tiller bud of its base portion is also easily sprouted, thereby breeds easily.Therefore, be suitable for gathering the coppice spout explant 4～June.
2) sterilization of explant
By the characteristics that the multilayer leaf sheath wraps up, adopt sterilization at the bud heart of coppice spout explant twice.At first, with washing powder explant is cleaned up, peel off 2～3 layers of leaf sheath, 75% alcohol disinfecting 20～25 seconds, 0.1% mercury chloride sterilization 10～12 minutes, rinsed with sterile water 1 time 10 minutes is peelled off 1～2 layer of leaf sheath again and is trimmed to the 3～4cm length that is suitable for cultivating, 0.1% mercury chloride sterilization 1～2 minute, rinsed with sterile water 3 times, be respectively 5 minutes, 5 minutes, 10 minutes.The sterilization success rate is greater than 35%.
3) prevent brown stain
Palm rattan coppice spout explant sterilization back brown stain is very serious, and it is dead rapidly not add the processing meeting.Find through series of studies, adopt methods such as dark place reason, additional activity carbon to be difficult to produce a desired effect, use polyvinylpyrrolidone then can cause explant to produce serious white secretion, adopt cysteine and ascorbic acid can suppress the brown stain of bud explant effectively, the two is in conjunction with better effects if.Be suitable for preventing that the cysteine of the anti-brown stain of coppice spout explant and the concentration of ascorbic acid are respectively 3mg/l and 10mg/l.
4) the clump bud is induced and enrichment culture
After explant collection and the sterilization, at first induce to form the clump bud on clump bud inducing culture, induction duration is 3～5 months, needs switching midway and changes fresh culture 2～3 times, the clump bud induction rate about 15% of explant.After forming the clump bud, be transferred to and carry out enrichment culture on the proliferated culture medium, the cycle of enrichment culture is 45～50 days, and the propagation multiple is 3～4 times.Big clump bud is cut into the budlet clump that 3～4 buds are formed, is transferred to and carries out enrichment culture next time on the fresh proliferated culture medium, clone nursery stock can be expanded numerous through enrichment culture repeatedly.The prescription of clump bud inducing culture and the prescription of proliferated culture medium see Table 1.
When the clump bud was induced, inoculation is induced the clump bud of coppice spout explant with cultural method also certain influence.After the explant sterilization, base portion 0.2～0.5cm is dead because of killing and wounding with brown stain, and dead part has a strong impact on the absorption of nutrition, bud must be tilted or keep flat, make it whole contact medium, can significantly propose the growth that promotes bud, promote the sprouting of secondary tiller bud, thereby promote propagation.
5) strong seedling culture
On proliferated culture medium, the growth of most bud seedling is less than the needed 3.0cm height of taking root, and therefore need carry out strong seedling culture for height less than the bud seedling of 3.0cm, allows the bud seedling grow into the height that is fit to culture of rootage.The prescription of strong seedling culture base sees Table 1.Time of strong seedling culture needs 45～50 days, can reach the needed height of culture of rootage through the bud seedling more than 75% after 1 strong seedling culture.The bud seedling that does not reach the culture of rootage height need carry out the strong seedling culture second time.
6) culture of rootage
Height is fit to culture of rootage the most greater than the bud seedling of 3.0cm, and the prescription of root media sees Table 1.The time of culture of rootage needs 50～60 days, and rooting rate can reach more than 85%.The bud seedling of not taking root carries out the culture of rootage second time after removing 1～2 layer of outer leaf sheath and base portion pruned, can take root.
If want to cultivate clump bud seedling, 2～4 bud cuttings that can bud height of seedling degree and size is approaching are transferred to and carry out culture of rootage on the root media same budlet clump, can cultivate clump bud seedling after the transplanting.
7) transplant in the field
The height greater than 8.0cm, root length greater than 4.0cm, have 3 the most suitable transplanting of nursery stock with blade.The medium that suitable single leaf is economized the transplanting of rattan tissue cultivating seedling has fine sand, peat soil and yellow soil, also the three can be mixed in 1: 1: 2 ratio, economizes the transplanting medium of rattan tissue cultivating seedling as single leaf.Transplant medium with 1/1000 disinfecting solution of potassium permanganate, transplant and spray in turn with 1/800 tpn and carbendazim in each week of back, the bacterium that prevents to mix grows.Transplant and began fertilising in back 15 days, adopt 1/1000 spray once in per 1 week of composite fertilizer's solution.Transplanted the back 1～7 day, 95% shading screen shades, and 8～30 days, 85% shading screen shaded, and 31～60 days, 70% shading screen shaded, and adopted 50% shading screen to shade subsequently, manages outplanting afforestation always.Transplanting success is greater than 85%.
When transplanting in the field, economize the rattan tissue cultivating seedling at 3～11 monthly portable list leaves, wherein be the best seasonal migration that single leaf is economized the rattan tissue cultivating seedling 3～June, and temperature, humidity all be fit to the further cultivation of nursery stock, reaches afforestation height and requirement.In 7～September, temperature is too high, and transplanting success is lower slightly, and by the intensity fertilising, nursery stock can reach the afforestation height.10～November can transplanting survival, but seedling growth season mistake, spring next year, nursery stock still can't reach the afforestation height.
The single leaf of table 1 is economized the culture medium prescription of rattan coppice spout tissue-culturing rapid propagation
The claimed quick breeding of the present invention also can comprise other examples of implementation.

Claims (6)

1. calamus simplicifolius clone tissue culture and fast propagation method is characterized in that: utilizing single leaf to economize the rattan coppice spout does not have sexual organ and carries out tissue-culturing rapid propagation, and concrete steps are as follows:
1) gathers the coppice spout explant, when fine day, use and saw coppice spout under the maternal plant saw; In the process of saw coppice spout explant, must prevent that the sprout wound and the bud heart from shattering;
2) sterilization is adopted in the sterilization of explant twice, and the first step at first cleans up explant, peels off outer leaf sheath, uses 75% alcohol disinfecting, with the sterilization of 0.1% mercury chloride, uses rinsed with sterile water more then; Second step, peel off the middle level leaf sheath again and be trimmed to 3～4cm length, with the sterilization of 0.1% mercury chloride, use the rinsed with sterile water several times again;
3) prevent brown stain, adopt cysteine or ascorbic acid or the two to handle in conjunction with the coppice spout explant being carried out anti-brown stain, the concentration of cysteine and ascorbic acid is respectively 3mg/l and 10mg/l;
4) the clump bud is induced and enrichment culture, after explant collection and the sterilization, at first induces to form the clump bud on clump bud inducing culture; Secondly after forming the clump bud, be transferred to and carry out enrichment culture on the proliferated culture medium, after one-step growth is cultivated, big clump bud is cut into the budlet clump that 3～4 buds are formed, be transferred to and carry out enrichment culture next time on the fresh proliferated culture medium, numerous through behind the enrichment culture repeatedly clone nursery stock being expanded;
5) culture of rootage is carried out the culture of rootage first time to the bud seedling that highly reaches 3.0cm at root media, to the bud seedling through also not taking root after the culture of rootage for the first time, after removing 1～2 layer of outer leaf sheath and base portion being pruned, carries out the culture of rootage second time; To highly not reaching the bud seedling of 3.0cm, carry out strong seedling culture at the strong seedling culture base earlier, allow the bud seedling grow to 3.0cm after, carry out culture of rootage again;
6) transplant in the field, height after the culture of rootage is carried out the field more than or equal to 8.0cm, root length more than or equal to 4.0cm, nursery stock with at least 3 blades transplant; The medium that carries out transplanting in the field is fine sand, peat soil and yellow soil;
7) in the clump bud inducing culture, the content of each component is respectively: NH 4NO 3825mg/L, KNO 33800mg/L, KH 2PO 4340mg/L, MgSO 4740mg/L, CaCl 2664mg/L, MnSO 44H 2O 22.3mg/L, ZnSO 47H 2O 8.6mg/L, H 3BO 36.2mg/L, Na 2MoO 42H 2O 0.25mg/L, FeSO 47H 2O 27.8mg/L, Na 2EDTA 37.3mg/L, KI 0.83mg/L, CuSO 45H 2O 0.025mg/L, CoCl 26H 2O 0.025mg/L, amion acetic acid 2mg/L, thiamine hydrochloride 0.2mg/L, puridoxine hydrochloride 0.75mg/L, nicotinic acid 0.5mg/L, inositol 100mg/L, vitamin C 10mg/L and cysteine 2mg/L, 6-BA 8mg/L, IBA 1mg/L, NAA 0.25mg/L, KT 0.5mg/L and 2,4-D 0.4mg/L; Sucrose 40g/L, carragheen 8.5～9.2g/L; The pH value of clump bud inducing culture is 5.8;
In the proliferated culture medium, the content of each component is respectively: NH 4NO 3825mg/L, KNO 33800mg/L, KH 2PO 4340mg/L, MgSO 4740mg/L and CaCl 2664mg/L, MnSO 44H 2O 22.3mg/L, ZnSO 47H 2O 8.6mg/L, H 3BO 36.2mg/L, Na 2MoO 42H 2O 0.25mg/L, FeSO 47H 2O 27.8mg/L, Na 2EDTA 37.3mg/L, KI 0.83mg/L, CuSO 45H 2O 0.025mg/L, CoCl 26H 2O 0.025mg/L, amion acetic acid 2mg/L, thiamine hydrochloride 0.2mg/L, puridoxine hydrochloride 0.75mg/L, nicotinic acid 0.5mg/L, inositol 100mg/L, vitamin C 10mg/L and cysteine 2mg/L, 6-BA 4mg/L, I BA 0.5mg/L, NAA 0.125mg/L, KT 0.25mg/L and 2,4-D 0.2mg/L; Sucrose 40g/L, carragheen 8.5～9.2g/L; The pH value of proliferated culture medium is 5.8;
In the strong seedling culture base, the content of each component is respectively: NH 4NO 3825mg/L, KNO 33800mg/L, KH 2PO 4340mg/L, MgSO 4740mg/L and CaCl 2664mg/L, MnSO 44H 2O 22.3mg/L, ZnSO 47H 2O 8.6mg/L, H 3BO 36.2mg/L, Na 2MoO 42H 2O 0.25mg/L, FeSO 47H 2O 27.8mg/L, Na 2EDTA 37.3mg/L, KI 0.83mg/L, CuSO 45H 2O 0.025mg/L, CoCl 26H 2O 0.025mg/L, amion acetic acid 2mg/L, thiamine hydrochloride 0.2mg/L, puridoxine hydrochloride 0.75mg/L, nicotinic acid 0.5mg/L, inositol 100mg/L, vitamin C 10mg/L and cysteine 2mg/L; Sucrose 40g/L, carragheen 8.5～9.2g/L; The pH value of strong seedling culture base is 5.8;
In the root media, the content of each component is respectively: NH 4NO 3825mg/L, KNO 3950mg/L, KH 2PO 485mg/L, MgSO 4185mg/L and CaCl 2166mg/L, MnSO 44H 2O 22.3mg/L, ZnSO 47H 2O 8.6mg/L, H 3BO 36.2mg/L, Na 2MoO 42H 2O 0.25mg/L, FeSO 47H 2O 27.8mg/L, Na 2EDTA 37.3mg/L, KI 0.83mg/L, CuSO 45H 2O 0.025mg/L, CoCl 26H 2O 0.025mg/L, amion acetic acid 2mg/L, thiamine hydrochloride 0.1mg/L, puridoxine hydrochloride 0.5mg/L, nicotinic acid 0.5mg/L and inositol 100mg/L, IBA 1～2mg/L, NAA 0.5～1.0mg/L; Sucrose 40g/L, carragheen 8.5～9.2g/L; The pH value of root media is 5.8.
2. calamus simplicifolius clone tissue culture and fast propagation method as claimed in claim 1 is characterized in that: the 1st) in the step, gathering highly is the coppice spout explant of 25～45cm; The 2nd) in the step when carrying out the first step of secondary sterilization, peel off outer 2～3 layers of leaf sheath,, sterilized 10～12 minutes with 0.1% mercury chloride then, used rinsed with sterile water again 1 time 10 minutes with 75% alcohol disinfecting 20～25 seconds; When the 2nd) carrying out second step of secondary sterilization in the step, peel off 1～2 layer of leaf sheath and be trimmed to 3～4cm length, with 0.1% mercury chloride sterilization 1～2 minute, use rinsed with sterile water again 3 times, each time was respectively 5 minutes, and 5 minutes, 10 minutes.
3. calamus simplicifolius clone tissue culture and fast propagation method as claimed in claim 1 is characterized in that: the 4th) when the step, the clump bud was induced, induction duration was 3～5 months, need switching midway and change fresh culture 2～3 times; The cycle that one-step growth is cultivated is 45～50 days; The 5th) when going on foot culture of rootage, the time of culture of rootage is 50～60 days for the first time; The 6th) when the step, transplanted in the field, transplant medium with 1/1000 disinfecting solution of potassium permanganate, transplant the back and spray in each week with bactericidal agent, prevent that assorted bacterium from growing; Transplant the per 1 week fertilising of beginning in back 15 days once, promptly need after the transplanting to shade, manage outplanting afforestation always.
4. calamus simplicifolius clone tissue culture and fast propagation method as claimed in claim 1 is characterized in that: carrying out the 4th) when step clump bud is induced, bud is tilted or keep flat, make it whole contact medium, to promote the growth of bud, promote the sprouting of secondary tiller bud, promote propagation.
5. calamus simplicifolius clone tissue culture and fast propagation method as claimed in claim 1, it is characterized in that: carrying out the 5th) when going on foot culture of rootage, 2～4 bud cuttings that bud height of seedling degree and size is approaching are same budlet clump, be transferred to and carry out culture of rootage on the root media, can cultivate clump bud seedling after the transplanting.
6. calamus simplicifolius clone tissue culture and fast propagation method as claimed in claim 1, it is characterized in that: the 6th) step field transplanting, transplant and begin fertilising after back 15 days, adopt 1/1000 spray once in per 1 week of composite fertilizer's solution, transplanted the back 1～7 day, 95% shading screen shades, 8～30 days, 85% shading screen shaded, 31～60 days, 70% shading screen shades, and adopts 50% shading screen to shade subsequently.
CN2007100317810A 2007-11-28 2007-11-28 Calamus simplicifolius clone tissue culture and fast propagation method Expired - Fee Related CN101167441B (en)

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CN103380682A (en) * 2012-05-02 2013-11-06 海南宏祺藤业科技股份有限公司 Cultivating method of strong seedlings of Hainan herba fibraureae recisae and calamus simplicifolius
CN103380681A (en) * 2012-05-02 2013-11-06 海南宏祺藤业科技股份有限公司 Dregea volubilis introduction and cultivation technology
CN102696482B (en) * 2012-06-18 2013-06-12 湖南省林业科学院 Method for detoxifying clonal tissue culture explant of Camellia oleifera Abel
CN102986533A (en) * 2012-12-14 2013-03-27 苏州和美生物科技有限公司 Culture medium special for preventing brown stain of butterfly orchids
CN102972277B (en) * 2012-12-27 2013-12-04 中国林业科学研究院热带林业研究所 Method for quickly raising clonal seedlings of teakwood by light matrix tray
CN106135006A (en) * 2016-08-28 2016-11-23 李志勇 A kind of foundation of palm tissue culture quick breeding technical system
CN112106656A (en) * 2020-09-24 2020-12-22 安徽农业大学 Direct organ generation type regeneration method of rhizoma acori graminei

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