CN106888968A - A kind of method of polygonatum filipes tissue culture and rapid proliferation - Google Patents
A kind of method of polygonatum filipes tissue culture and rapid proliferation Download PDFInfo
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- CN106888968A CN106888968A CN201710081087.3A CN201710081087A CN106888968A CN 106888968 A CN106888968 A CN 106888968A CN 201710081087 A CN201710081087 A CN 201710081087A CN 106888968 A CN106888968 A CN 106888968A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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Abstract
The invention belongs to technical field of plant culture, and in particular to the method for polygonatum filipes tissue culture and rapid proliferation.A kind of method of polygonatum filipes tissue culture and rapid proliferation, the method is comprised the following steps：（1）The sterilization and inoculation of polygonatum filipes bud；（2）The induction of Multiple Buds；（3）The propagation of adventitious bud；（4）The culture of rootage of regeneration plant；（5）Acclimatization and transplantses.Using the method for the present invention, compared with it has been reported that document, it is low to reduce shortening pre-treatment time, explant pollution rate, shortens adventitious bud time of origin, so as to faster obtain polygonatum filipes regeneration plant, beneficial to industrialization production.Using the inventive method, the adventitious bud induction frequency of polygonatum filipes Multiple Buds is up to 90.0%, adventitious bud number 8.4, and, up to 7.7, plantlet of transplant survival rate is up to more than 95% for adventitious bud proliferation multiple.
Description
Technical field
The invention belongs to technical field of plant culture, and in particular to the method for polygonatum filipes tissue culture and rapid proliferation.
Background technology
Polygonatum filipes (Polygonatum filipes Merr.), are Liliaceae Polygonatum herbaceos perennial, distribution
In Jiangsu, Anhui, Zhejiang, Jiangxi, Hunan, Fujian, Guangdong (the north), the dark and damp rich soil of sylvan life, shrubbery or careless slope is born in
In, because its root-like stock is largely planted in locality so that sealwort is used as medicine, while also edible.Polygonatum filipes aborning is numerous at present
Grow using seed and root-like stock, mainly based on root-like stock, have the shortcomings that breeding coefficient is low, the breeding cycle is long, therefore build
The tissue culture rapid propagation system of vertical polygonatum filipes, is the another effective way for realizing its industrialization production.
At present polygonatum filipes and sealwort, the explant of use are focused primarily upon for the tissue cultures that Liliaceae Polygonatum is carried out
Body is mainly root-like stock or the bud produced by root-like stock.Because root-like stock is grown on underground, surface carries many miscellaneous bacterias, and also deposits
In endophyte, therefore the control relative difficult of its pollution rate, need the long period just to induce adventitious bud after sterilizing in addition.Substantially
The optimal minimal medium that the report of upper all HUANGJING ZANYU CAPSULE tissue cultures is used is the MS culture mediums of various concentrations, wherein
Process before root induction mainly uses MS culture mediums, and also having carries out the report of adventitious bud inducing using 1/2MS；During root induction
It is main to use 1/2MS culture mediums, also have using MS culture mediums and 1/3MS culture mediums.The induction of Polygonatum adventitious bud is even more
Basic element of cell division 6-BA has often been used in the induction of injured tissue, and the optium concentration that many document report 6-BA are used is 4.0mg/L；
In culture of rootage be used alone auxin or its combination, or even have 2,4-D can hestening rooting report.The group of current polygonatum filipes
There is not been reported to knit culture and quick breeding.
The content of the invention
The technical problems to be solved by the invention are：In view of the shortcomings of the prior art, there is provided a kind of survival rate is high, profit
In the method for the polygonatum filipes tissue culture and rapid proliferation for realizing industrialization production.
To realize the purpose of the present invention, it is achieved using following technical scheme：
A kind of method of polygonatum filipes tissue culture and rapid proliferation, the method is comprised the following steps：
(1) sterilization and inoculation of polygonatum filipes bud
The bud of the polygonatum filipes not yet bloomed is taken in May then, 4 DEG C of deepfreeze 0-48h of refrigerator are put into, added after taking-up
Enter appropriate liquid detergent solution immersion 2min, after rinsing 0.5-1.0h with flowing water, stem section is transferred to 70% wine in super-clean bench
1min is soaked in smart solution, with aseptic water washing 3 times, immersion dropwise addition there are 5 drop Tween-80s, and mass fraction is 0.1% HgCl2
Soaking disinfection 9-10min in solution, is then fully embathed 3 times with sterilized water, the explant material being sterilized.
(2) induction of Multiple Buds
Explant material obtained in the previous step is put on aseptic filter paper, is cut end to end, stamen and gynoecium are removed, by petal
2cm2 sizes are cut into, are connected in the MS minimal mediums of addition TDZ 0-2.0mg/L, NAA 0-1.0mg/L, AC 0-2.0g/L,
Sucrose addition in the culture medium is 30g/L, and agar addition is 8.0g/L, and pH is 5.8-6.0, and culture medium is once in 121 DEG C
Lower sterilizing 20min, similarly hereinafter.The culture medium for connecting petal is put prior to cultivating 0-7d in dark, and cultivation temperature is 28 ± 2 DEG C；Then
Illumination cultivation is transferred to, condition of culture is：Cultivation temperature is (28 ± 2) DEG C, and light application time is 14h light/10h dark, intensity of illumination 30-
40μmol/(m2·s)；
(3) propagation of adventitious bud
The adventitious bud clump that previous step induction is obtained is cut into the 2-3 sprout tuber of adventitious bud, is inoculated into and is added with TDZ 0-
Cultivated in the MS minimal mediums of 2.0mg/L, 6-BA 0-5.0mg/L, CH 0-1.0mg/L, AC 0-2.0g/L, cultivated
Condition is：Cultivation temperature is (28 ± 2) DEG C, and light application time is 14h light/10h dark, 30-40 μm of ol/ of intensity of illumination (m2s)；
(4) culture of rootage of regeneration plant
Cut that blade is light green, grow vigorous and 3-4 pieces of blade adventitious bud, insertion is added with 6-BA0-4.0, NAA 0-
Taken root in the MS minimal mediums of the 12.5%-100% concentration of 2.0mg/L, banana puree 0-150g/L, the sugarcane added in culture medium
Sugar is 20-30g/L, and agar addition is 8.0g/L, pH 5.8-6.0, and once in the 20min that sterilized at 121 DEG C；
(5) acclimatization and transplantses
The height of seedling of the tissue-cultured seedling of under growth root is long to more than 5cm, root it is long more than 5cm when, unclamp tissue culture bottle bottle cap, toward bottle
A small amount of sterilized water is injected to prevent culture medium dry and cracked, in the later half bottle caps of moving away of 6h, and sterilized water is added, and after 18h, removes bottle
Lid, allow 30-40 μm of ol/ (m2s) light intensity light directly according to regenerated plant culture 2d, period adds sterilized water several times.Then it is small
Heart smashs culture medium to pieces, takes out regeneration plant, is carefully rinsed well the agar of residual with flowing water, plant is colonized in and was once disappeared
The perlite that poison is crossed, vegetable garden soil weight ratio is 1：In mixed-matrix 100-200), pour permeable and with transparent vinyon
Film covering is controlled between 24-28 DEG C with heat and moisture preserving, indoor temperature.Sealed polyethylene plastic corner, next day are opened after 3d
Sealed polyethylene plastic is all opened, after after young leaves expansion can balled transplanting to crop field.
Preferably：The material of inoculation is the bud not yet bloomed in the step (1), and in 4 DEG C of low temperature of refrigerator
Refrigeration 12-24h.
Preferably：MS culture mediums in the step (2) are added with TDZ 0.5mg/L, NAA 0.2mg/L, AC
0.5-1.0mg/L, illumination cultivation is transferred to petal has been inoculated with again after 3-5d is cultivated in dark.
Preferably：MS culture mediums are added with TDZ 0.05mg/L, 6-BA 2.0mg/L, CH in the step (3)
0.3-0.4g/L, AC 0.5-1.0mg/L.
Preferably：In the step (4) prescription of rooting medium be 50%MS, 6-BA0.2, NAA 0.5mg/L,
Banana puree 80-100g/L.
Compared with prior art, the beneficial effects of the invention are as follows：Using the method for the present invention, compared with it has been reported that text
Offer, it is low to reduce shortening pre-treatment time, explant pollution rate, shortens adventitious bud time of origin, so as to faster obtain stalk long
Sealwort regeneration plant, beneficial to industrialization production.Using the inventive method, the adventitious bud induction frequency of polygonatum filipes Multiple Buds is up to
90.0%, adventitious bud number 8.4, up to 7.7, plantlet of transplant survival rate is up to more than 95% for adventitious bud proliferation multiple.
Initialism in the present invention：
AC activated carbons
6-BA 6- benzyl aminoadenines
CH caseinhydrolysates
NAA methyl α-naphthyl acetates
TDZ Thidiazurons (Thidiazuron)
Specific embodiment
Embodiment 1
A kind of method of polygonatum filipes tissue culture and rapid proliferation, comprises the following steps successively：
(1) sterilization of polygonatum filipes bud
The bud of the polygonatum filipes not yet bloomed is taken in May then, 4 DEG C of deepfreeze 12-24h of refrigerator is put into, after taking-up
Appropriate liquid detergent solution immersion 2min is added, after rinsing 0.5-1.0h with flowing water, stem section 70% is transferred in super-clean bench
1min is soaked in alcoholic solution, with aseptic water washing 3 times, immersion dropwise addition there are 5 drop Tween-80s, and mass fraction is 0.1%
HgCl2Soaking disinfection 9-10min in solution, is then fully embathed 3 times with sterilized water, and the explant material being sterilized disappears
Malicious success rate is up to more than 90%.
(2) induction of Multiple Buds
Explant material obtained in the previous step is put on aseptic filter paper, is cut end to end, stamen and gynoecium are removed, by petal
It is cut into 2cm2Size, is connected in the MS minimal mediums of addition TDZ 0.5mg/L, NAA 0.2mg/L, AC 0.5-1.0g/L, should
Sucrose addition in culture medium is 30g/L, and agar addition is 8.0g/L, and pH is 5.8-6.0, and culture medium is once at 121 DEG C
Sterilizing 20min, similarly hereinafter.The culture medium for connecting petal is put prior to cultivating 3-5d in dark, and cultivation temperature is 28 ± 2 DEG C；Then turn
Enter illumination cultivation, condition of culture is：Cultivation temperature is (28 ± 2) DEG C, and light application time is 14h light/10h dark, intensity of illumination 30-40
μmol/(m2·s)；Starting to occur petal section during culture 15d has protuberance, and adventitious bud clump can have been found during 20d, indefinite during 30d
Bud can grow to 3cm or so, and up to 90.0%, the adventitious bud number of the adventitious bud clump of induction has 8.4 to the inductivity of adventitious bud.
(3) propagation of adventitious bud
The adventitious bud clump that previous step induction is obtained is cut into the 2-3 sprout tuber of adventitious bud, is inoculated into and is added with TDZ
Cultivated in the MS minimal mediums of 0.05mg/L, 6-BA2.0mg/L, CH 0.3-0.4mg/L, AC 0.5-1.0g/L, trained
Foster condition is：Cultivation temperature is (28 ± 2) DEG C, and light application time is 14h light/10h dark, 30-40 μm of ol/ (m of intensity of illumination2·s)；
Counted after culture 30d, proliferation times 7.7, adventitious bud grows fine.
(4) culture of rootage of regeneration plant
Cut that blade is light green, grow vigorous and 3-4 pieces of blade adventitious bud, insertion is added with 6-BA 0.2, NAA
Taken root in the MS minimal mediums of 50% concentration of 0.5mg/L, banana puree 80-100g/L, the sucrose added in culture medium is
20-30g/L, agar addition is 8.0g/L, pH 5.8-6.0, and once in the 20min that sterilized at 121 DEG C；Culture 30d has found to take root
Rate forms spherosome up to more than 95%, and root is generated thereon, and root is about 6cm, sturdy, and length has many root hairs.
(5) acclimatization and transplantses
The height of seedling of the tissue-cultured seedling of under growth root is long to more than 5cm, root it is long more than 5cm when, unclamp tissue culture bottle bottle cap, toward bottle
A small amount of sterilized water is injected to prevent culture medium dry and cracked, in the later half bottle caps of moving away of 6h, and sterilized water is added, and after 18h, removes bottle
Lid, allows 30-40 μm of ol/ (m2S) directly according to regenerated plant culture 2d, period adds sterilized water several times to the light of light intensity.Then it is small
Heart smashs culture medium to pieces, takes out regeneration plant, is carefully rinsed well the agar of residual with flowing water, plant is colonized in and was once disappeared
The perlite that poison is crossed, vegetable garden soil weight ratio is 1：In the mixed-matrix of 100-200, pour permeable and thin with transparent vinyon
Film covering is controlled between 24-28 DEG C with heat and moisture preserving, indoor temperature.Sealed polyethylene plastic corner is opened after 3d, next day will
Sealed polyethylene plastic is all opened, after after young leaves expansion can balled transplanting to crop field.Practice shoot survival percent and transplanting survival rate
Up to more than 98%.
Embodiment 2
The present embodiment is step (1) with the difference of embodiment 1：The sterilization of polygonatum filipes bud：Taken not yet in May then
The bud of the polygonatum filipes bloomed, is put into 4 DEG C of deepfreeze 0-48h of refrigerator, adds appropriate liquid detergent solution to soak after taking-up
2min, after rinsing 0.5-1.0h with flowing water, 1min is soaked during stem section to be transferred to 70% alcoholic solution in super-clean bench, with aseptic
Water is rinsed 3 times, and immersion dropwise addition has 5 drop Tween-80s, and mass fraction is 0.1% HgCl2Soaking disinfection 9-10min in solution, so
Fully embathed with sterilized water 3 times afterwards, the explant material being sterilized.Find out in inoculation 30d statisticses, afterwards using bud
The pollution rate for obtaining is carried out disinfection as explant will be well below root-like stock be used as explant, while bud is used as outer
It is that root is also significantly larger than on explant, and adventitious bud number that the time of implant evoking adventive bud also will significantly faster than use root-like stock
Shape stem is explant, and in the comparing of Cold pretreatment, is all petal as explant, and pollution rate is basically unchanged, but enters
The inductivity of Multiple Buds has significantly raised after row Cold pretreatment, and adventitious bud number has a small amount of increase.From table 1, low temperature is pre-
Treatment 24-48h has optimal inducing clumping bud rate and adventitious bud number.
The influence of table 1 different explants and Cold pretreatment to pollution rate and inducing clumping bud
Embodiment 3
The present embodiment is step (2) with the difference of embodiment 1：The induction of Multiple Buds：By explant obtained in the previous step
Material is put on aseptic filter paper, is cut end to end, removes stamen and gynoecium, and petal is cut into 2cm2Size, is connected to addition TDZ 0-
In the MS minimal mediums of 2.0mg/L, NAA 0-1.0mg/L, AC 0-2.0g/L, the sucrose addition in the culture medium is
30g/L, agar addition is 8.0g/L, and pH is 5.8-6.0, culture medium once in the 20min that sterilized at 121 DEG C, similarly hereinafter.Connect petal
Culture medium put prior to cultivating 0-7d in dark, cultivation temperature is 28 ± 2 DEG C；Illumination cultivation is then continued at, condition of culture is：Training
It is (28 ± 2) DEG C to support temperature, and light application time is 14h light/10h dark, 30-40 μm of ol/ (m of intensity of illumination2·s)；Hair is counted after 30d
It is existing,；The different disposal of TDZ concentration shows, the TDZ (0.1mg/L) of low concentration has and lured than the TDZ 0.5mg/L low adventitious buds for the treatment of
Conductance and adventitious bud number, the growth of adventitious bud are also weaker, and the TDZ (2.0mg/L) of high concentration treatment is compared with TDZ 0.5mg/L treatment
Slightly higher adventitious bud inducing and slightly lower adventitious bud number, the growth of adventitious bud show vitrification phenomenon；AC can play absorption to be had
The effect of evil material, then AC additions are excessive but absorbable nutrition, and the AC of debita spissitudo has high indefinite as can be seen from Table 2
Bud induction rate and adventitious bud number, will also result in the decline of adventitious bud induction frequency and adventitious bud number due to browning during without AC, and
Adventitious bud induction frequency and adventitious bud number can be also excessively reduced, slowing down for the adventitious bud speed of growth is also brought along；With MS culture medium phases
Than the adventitious bud induction frequency and adventitious bud number for being incubated at the SH culture mediums of identical hormone and concentration of activated carbon combination are weaker, no
The growth of normal bud is similar.Therefore the MS culture mediums of addition TDZ 0.5mg/L, NAA 0.2mg/L, AC 0.5g/L have highest not
Normal bud inductivity and adventitious bud number, and the growth of adventitious bud is best, show as that leaf color is dark green, and stem is high and sturdy.
Influence of the different disposal of table 2 to inducing clumping bud
Embodiment 4
The present embodiment is step (3) with the difference of embodiment 1：The propagation of adventitious bud：Step (2) induction is obtained not
Normal bud clump is cut into the 2-3 sprout tuber of adventitious bud, is inoculated into and is added with TDZ 0-2.0mg/L, 6-BA 0-5.0mg/L, CH 0-
Cultivated in the MS minimal mediums of 1.0mg/L, AC 0-2.0g/L, condition of culture is：Cultivation temperature is (28 ± 2) DEG C, light
It is 14h light/10h dark, 30-40 μm of ol/ (m of intensity of illumination according to the time2·s)；Counted after 30d and found, adventitious bud in each culture medium
Proliferation times between 4.5-8.6, but proliferation times difference substantially, with the rising of TDZ concentration, although improve propagation
Multiple, but serious vitrifying is brought, the leaf of growth deformity, stem also water stainization, the adventitious bud of acquisition is in culture of rootage
Can not take root completely；But compared with (proliferation times 4.5) without TDZ, the presence of TDZ promotes propagation (propagation really
Multiple 7.7)；Study and also found, the propagation being added with beneficial to adventitious bud of CH, but finite concentration above effect is not further added by.
The concentration of 6-BA also has an impact for the propagation of adventitious bud, and the effect of low concentration is also weaker than debita spissitudo, high concentration but
Bring the vitrifying of adventitious bud.Table 3 in general, with the addition of TDZ 0.05mg/L, 6-BA 2.0mg/L, CH 0.4g/L, AC
The MS minimal mediums of 0.5g/L are best suitable for the propagation of adventitious bud.
Influence of the different disposal of table 3 to adventitious bud proliferation
| Culture medium | Proliferation times | Adventitious bud growing state |
| MS, TDZ 0.05,6-BA 2.0, CH 0.4, AC 0.5 | 7.7 | Leaf color is dark green, and stem is thick, and growth is quick fast |
| MS, TDZ 0.5,6-BA 2.0, CH 0.4, AC 0.5 | 8.6 | Vitrifying |
| MS, TDZ 0.05,6-BA 2.0, AC 0.5 | 7.1 | Leaf color is dark green, and stem is thick, fast growth |
| MS, 6-BA 2.0, CH 0.4, AC 0.5 | 4.5 | Leaf color is dark green, and stem is thick, fast growth |
| MS, TDZ 0.05,6-BA 1.0, CH 0.4, AC 0.5 | 6.4 | Leaf color is dark green, and stem is thicker, and the speed of growth is very fast |
Embodiment 5
The present embodiment is step (4) with the difference of embodiment 1：The culture of rootage of regeneration plant：Cut blade light green, raw
Vigorous and 3-4 pieces of blade the adventitious bud of length, insertion is added with 6-BA 0-4.0, NAA 0-2.0mg/L, banana puree 0-150g/L's
Taken root in the MS minimal mediums of 12.5%-100% concentration, the sucrose added in culture medium is 20-30g/L, agar addition
It is 8.0g/L, pH 5.8-6.0, and once in the 20min that sterilized at 121 DEG C.During 30d count find, rooting rate in 22.5-95.0%,
Earliest rootage duration 12-22d.MS minimal medium rooting rates without the low concentration (12.5%) of any hormone are extremely low, and
The growth of tissue-cultured seedling is not good, and 100%MS is also unfavorable for taking root, and 50%MS is suitable；The addition of 6-BA, although for rooting rate
It is several without influence, but it is favourable for the growth of tissue-cultured seedling overground part, promotes the growth of overground part；NAA is served for taking root
Crucial effect, the NAA of low concentration can cause thin and delicate root system, and with increasing for NAA concentration, root is thicker to shorten, therefore also uncomfortable
Work as high concentration；Addition AC and banana puree considerably increase rooting rate, AC root growth can be provided needed for dark surrounds, and banana
Mud may also function as promoting the effect of tissue-cultured seedling growth, it may be possible to because it is internal with growth promoting substance.Consolidated statement 4, it is optimal
50%MS, 6-BA 0.2mg/L, NAA 0.5mg/L are combined as, banana puree 80g/L, sucrose concentration is 20-30g/L, its rooting rate
Reachable 95%, radical 5.6, root is slightly moderate, is suitable to acclimatization and transplantses.
Influence of the different disposal of table 4 to inducing clumping bud
Claims (5)
1. a kind of method of polygonatum filipes tissue culture and rapid proliferation, it is characterised in that the method is comprised the following steps：
1）The sterilization and inoculation of polygonatum filipes bud
The bud of the polygonatum filipes not yet bloomed is taken in May then, 4 DEG C of deepfreeze 0-48 h of refrigerator are put into, added after taking-up
Appropriate liquid detergent solution soaks 2 min, after rinsing 0.5-1.0 h with flowing water, stem section is transferred to 70% alcohol in super-clean bench
1 min is soaked in solution, with aseptic water washing 3 times, immersion dropwise addition there are 5 drop Tween-80s, and mass fraction is 0.1% HgCl2It is molten
Soaking disinfection 9-10 min in liquid, are then fully embathed 3 times with sterilized water, the explant material being sterilized；
2）The induction of Multiple Buds
Explant material obtained in the previous step is put on aseptic filter paper, is cut end to end, remove stamen and gynoecium, petal is cut into
2 cm2Size, is connected in the MS minimal mediums of addition TDZ 0-2.0 mg/L, NAA 0-1.0 mg/L, AC 0-2.0 g/L,
Sucrose addition in the culture medium is 30 g/L, and agar addition is 8.0 g/L, and pH is 5.8-6.0, and culture medium is once in 121
Sterilize 20 min at DEG C；The culture medium for connecting petal is put prior to cultivating 0-7 d in dark, and cultivation temperature is 28 ± 2 DEG C；Then turn
Enter illumination cultivation, condition of culture is：Cultivation temperature is 28 ± 2 DEG C, and light application time is 14 h light/10 h dark, intensity of illumination 30-40
μmol/(m2·s)；
3）The propagation of adventitious bud
The adventitious bud clump that previous step induction is obtained is cut into the 2-3 sprout tuber of adventitious bud, is inoculated into and is added with TDZ 0-2.0 mg/
Cultivated in the MS minimal mediums of L, 6-BA 0-5.0 mg/L, CH 0-1.0 mg/L, AC 0-2.0 g/L, condition of culture
For：Cultivation temperature is 28 ± 2 DEG C, and light application time is 14 h light/10 h dark, 30-40 μm of ol/ (m of intensity of illumination2·s)；
4）The culture of rootage of regeneration plant
Cut that blade is light green, grow vigorous and 3-4 pieces of blade adventitious bud, insertion is added with 6-BA 0-4.0, NAA 0-2.0
Taken root in the MS minimal mediums of the 12.5%-100% concentration of mg/L, banana puree 0-150 g/L, the sucrose added in culture medium is
20-30 g/L, agar addition is 8.0 g/L, pH 5.8-6.0, and once in 20 min that sterilized at 121 DEG C；
5）Acclimatization and transplantses
The height of seedling of the tissue-cultured seedling of under growth root is long to 5 more than cm, root it is long more than 5 cm when, unclamp tissue culture bottle bottle cap, toward bottle note
Enter a small amount of sterilized water to prevent culture medium dry and cracked, in the later half bottle caps of moving away of 6 h, and add sterilized water, after 18 h, remove bottle
Lid, allows 30-40 μm of ol/ (m2S) d of light direct irradiation regenerated plant culture 2 of light intensity, period adds sterilized water to prevent several times
Only culture medium season cracking；Then carefully culture medium is smashed to pieces, takes out regeneration plant, rushed the agar for remaining with flowing water is careful
Wash clean, plant is colonized in the perlite once sterilized, and vegetable garden soil weight ratio is 1：In the mixed-matrix of 100-200, irrigate
Water is simultaneously covered with heat and moisture preserving with transparent sealed polyethylene plastic, and indoor temperature is controlled between 24-28 DEG C；Opened after 3 d
Sealed polyethylene plastic corner, next day all opens sealed polyethylene plastic, after after young leaves expansion can balled transplanting to big
Field.
2. the method for a kind of polygonatum filipes tissue culture and rapid proliferation according to claim 1, it is characterised in that described
Step 1）The material of middle inoculation is the bud not yet bloomed, and in 4 DEG C of deepfreeze 12-24 h of refrigerator.
3. the method for a kind of polygonatum filipes tissue culture and rapid proliferation according to claim 1, it is characterised in that described
Step 2）In MS culture mediums be added with TDZ 0.5 mg/L, NAA 0.2 mg/L, AC 0.5-1.0 mg/L, be inoculated with flower
Valve is transferred to illumination cultivation again after 3-5 d are cultivated in dark.
4. the method for a kind of polygonatum filipes tissue culture and rapid proliferation according to claim 1, it is characterised in that described
Step 3）Middle MS culture mediums are added with TDZ 0.05 mg/L, 6-BA 2.0 mg/L, CH 0.3-0.4 g/L, AC 0.5-1.0
mg/L。
5. the method for a kind of polygonatum filipes tissue culture and rapid proliferation according to claim 1, it is characterised in that described
Step 4）Middle prescription of rooting medium is 50%MS, 6-BA 0.2, NAA 0.5 mg/L, banana puree 80-100 g/L.
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| CN102550413A (en) * | 2011-12-26 | 2012-07-11 | 福建省三明市农业科学研究所 | Rapid propagation method for tube seedlings of high-grade polygonatum cyrtonema hua |
| CN105532471A (en) * | 2016-01-13 | 2016-05-04 | 福建柘参种业有限公司 | Novel high-yield, high-quality and large-scale polygonatum cyrtonema cultivation method |
| CN105594426A (en) * | 2015-12-23 | 2016-05-25 | 安徽森沣农业综合开发有限公司 | Method for planting polygonatum cyrtonema |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102550413A (en) * | 2011-12-26 | 2012-07-11 | 福建省三明市农业科学研究所 | Rapid propagation method for tube seedlings of high-grade polygonatum cyrtonema hua |
| CN105594426A (en) * | 2015-12-23 | 2016-05-25 | 安徽森沣农业综合开发有限公司 | Method for planting polygonatum cyrtonema |
| CN105532471A (en) * | 2016-01-13 | 2016-05-04 | 福建柘参种业有限公司 | Novel high-yield, high-quality and large-scale polygonatum cyrtonema cultivation method |
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