CN102342246A - Rhododendron decorum tissue-culture quick propagation method - Google Patents
Rhododendron decorum tissue-culture quick propagation method Download PDFInfo
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- CN102342246A CN102342246A CN2010102401709A CN201010240170A CN102342246A CN 102342246 A CN102342246 A CN 102342246A CN 2010102401709 A CN2010102401709 A CN 2010102401709A CN 201010240170 A CN201010240170 A CN 201010240170A CN 102342246 A CN102342246 A CN 102342246A
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Abstract
The invention discloses a rhododendron decorum tissue-culture quick propagation method, and particularly relates to a quick propagation method which adopts a tissue culture technique to carry out the callus induction to form caespitose shoots which are differentiated to a complete plant. The rhododendron decorum tissue-culture quick propagation method comprises the following steps: a. preparation of explants; b. establishment of aseptic culture system; c. induction and propagation culture of caespitose shoots; d. elongation of the caespitose shoots and sound seedling culture; e. rooting induction culture of test-tube plantlets; and then after being cut into a single plant, each caespitose shoot with the height of 2cm to 3cm is inoculated onto rooting culture medium, the culture condition is that the culture temperature is 25 DEG C plus or minus 1, illumination time is 12h.d-1, and the illumination strength is 40mol.m-2.s-1 to 60 mol.m-2.s-1; the caespitoes shoot begins to rooting after 50 days, the rooting rate is 60 percent to 65 percent, the height of the seedling can reach 4cm to 5cm after 70 days, the quantity of the root can reach 2 to 3, and the complete plant is transplanted into substrate which is the mixture of yellow sand, perlite and humus soil to be processed in domestication culture. The rhododendron decorum tissue-culture quick propagation method has the advantages that the culture period is shortened, the method is free from being influenced by the season, the survival rate is high, the yield is high and the like.
Description
Technical field
The present invention relates to a kind of Da Bai cuckoo tissue culture quick propagation culturing method of wild alpine rose kind, belong to flowers culture technique field.
Background technology
The Da Bai cuckoo is the evergreen woody plant of Ericaceae (Ericaceae) Rhododendron (Rhododendron L.); Be China endemic species; Mainly be distributed in ground such as Sichuan, Yunnan, Guizhou, Tibet, Hubei, Hunan, be born in the sylvan life, shrubbery of height above sea level 1000～3200m.Big Rhododendron mucronatum is big, and the corolla funnel-form is bell, white or band rose look, and the green or pink spot of bennet tool in the flowers are in blossom, has a burst of delicate fragrance to blow against one's face, and gladdens the heart and refresh the mind, and ornamental value is high.The Da Bai cuckoo still is China's Chinese herbal medicine commonly used among the people, and its nature and flavor " hardship, cold, slightly poisonous " have the menses of mediation, the clear larynx of moistening lung, beneficial gas effect such as allay excitement.In addition, big Rhododendron mucronatum lobe is edible also, is rich in several amino acids and vitamin, and taste is exquisite, nutritious.At present, be main, artificial unauthorized and excessive mining mainly to the utilization of Da Bai cuckoo to gather wild resource, Da Bai cuckoo habitat is seriously damaged, resource is on the hazard.At present, seminal propagation of existing Da Bai cuckoo and cottage propagation technology are difficult to grow seedlings on a large scale because of being subjected to the restriction in resource and season, and prior art also exists problems such as cultivation cycle is long, survival rate is lower, output is lower.For better protection, utilization, exploitation Da Bai azalea resource, research Da Bai cuckoo group culturation rapid propagating technology is significant.
Summary of the invention
The objective of the invention is to, provide a kind of cultivation cycle to shorten, be not subjected to seasonal effect and the higher Da Bai cuckoo seed embryo tissue culture quick propagation culturing method of output, to overcome the deficiency of prior art.
Technical scheme of the present invention; A kind of Da Bai cuckoo tissue culture quick propagation culturing method of the present invention is to adopt tissue culture technique that the cotyledon of seed embryo and plumular axis are carried out callus induction to form a kind of tissue culture quick propagation culturing method that the bud of growing thickly then is divided into whole plant then, and its process comprises the following steps:
The preparation of a, explant
Get the still uncracked capsule of Da Bai cuckoo; Under flowing water, scrub the capsule surface with brush; On superclean bench with 75% ethanol disinfection 30S; Inoculation is with 0.1% mercuric chloride solution sterilization 10min; Aseptic water washing 1 time soaks 3～5min, aseptic water washing 5～6 times with 10% hydrogenperoxide steam generator again; Blot surface moisture with sterilized filter paper at last, subsequent use;
The foundation of b, aseptic culture system
Capsule having been gone out behind the bacterium, with scissors and tweezers strip off capsule, seed has been trembled on the sterile seed germination medium that is sprinkling upon the bacterium of having gone out, is that 25 (± 1) ℃, light application time are 12hd in cultivation temperature then
-1With intensity of illumination be 40～60 mo1m
-2S
-1Condition of culture under cultivate;
C, inducing clumping bud and enrichment culture
All sprout when seed and to finish, when cotyledon all launches, aseptic seedling is divided into cotyledon and plumular axis two parts are inoculated into respectively on the inducing clumping bud medium, its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination be 40～60 mo1m
-2S
-1
Behind the one and a half months, elongation growth appears in cotyledon, becomes 2 true leaves, and a small amount of yellow callus appears in the cotyledon base portion; The plumular axis edge begins protuberance and expands, and yellow callus of small quantities of particles shape and green callus occur; After 2 months; Callus obviously increases, and the callus that cotyledon forms differentiates the bud of growing thickly, at this moment; The callus that plumular axis forms a large amount of green bud points that may be seen indistinctly, the callus that will grow thickly bud and plumular axis form continue to be transferred to and carry out the shoot proliferation cultivation on the shoot proliferation medium; Its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1Be 1～February the blanking time that shoot proliferation is cultivated, and its reproduction coefficient can reach 6.5;
The elongation of d, the bud of growing thickly and strong seedling culture
The bud of will growing thickly cuts down from callus, and the bud of growing thickly is divided into fritter and is transferred on grow thickly bud elongation and the strong seedling culture base, and its condition of culture is that cultivation temperature is 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1
On grow thickly bud elongation and strong seedling culture base, bud seedling robust growth after 30 days, the individual plant bud of growing thickly is extending to about 2 centimetres, and the budlet seedling on the bud clump can rise to 1.0 centimetres;
E, rooting of vitro seedling inducing culture
After the bud seedling of growing thickly of high 2～3 cm is cut into individual plant, be inoculated on the root media, its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1
Begin behind the 50d to take root, rooting rate is that height of seedling can reach 4～5 cm behind 60%～65%, 70 d, and 2～3 of radicals are transplanted to whole plant and are refined seedling in the matrix that yellow sand, perlite, humus soil mix and cultivate.
Above-mentioned sterile seed germination medium is 1/4MS medium, sucrose 30g/L, zeatin 0.5mg/L, methyl 0.01mg/L.
Above-mentioned inducing clumping bud medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2～2.0mg/L, methyl 1.0mg/L.
Above-mentioned shoot proliferation medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2 ~ 2.0mg/L, methyl 1.0mg/L.
The above-mentioned bud of growing thickly extends and the strong seedling culture base is 1/4MS medium, sucrose 30g/L, benzthiazuron 0.5mg/L, gibberellin 1.0～2.0mg/L.
Above-mentioned root media is 1/4MS medium, sucrose 25g/L, indolebutyric acid 1.0mg/L, methyl 3.0mg/L, active carbon 0.8 mg/L.
In above-mentioned sterile seed germination medium, inducing clumping bud medium, shoot proliferation medium, the bud of growing thickly elongation and strong seedling culture base and root media, all contain agar 10g/L and pH5.2～5.4 respectively.
Owing to adopted technique scheme, the present invention to have the advantages that cultivation cycle is short, the branch branch growth is good, be not subjected to seasonal effect.Usually Da Bai cuckoo reproduction coefficient is low, and reproduction coefficient of the present invention can reach 6.5, has improved Da Bai cuckoo coefficient greatly, can obtain a large amount of Da Bai cuckoo seedlings in a short time.
Da Bai cuckoo seed embryo tissue culture quick propagation culturing method of the present invention is that the back of a series of test below inventor's process is resulting:
Sterile seed germination research
Different hormone kinds of table 1 and consumption are to the influence of sterile seed germination
Top table 1 data show adds plant hormone and does not add plant hormone and all can induce sterile seed germination in the 1/4MS medium.But induce difference on effect bigger.ZT0.5 mgL
-1+ NAA0.01 mgL
-1Combined effect is better, and the blank medium effect of 1/4MS is relatively poor.At ZT0.5 mgL
-1+ NAA0.01 mgL
-1In the medium, begin about aseptic seed 17d to sprout, cultivate all to sprout after 50 days and finish, the cotyledon full expand, seed can be grown on medium, and seed germination rate is low under this moment other prescription, seed only can lie low on medium.Therefore, filter out 1/4MS+ZT0.5 mgL
-1+ NAA0.01 mgL
-1Culture medium prescription is fit to Da Bai cuckoo seed germination.
Inducing clumping bud and proliferation research
All sprout when seed and to finish, when cotyledon all launches, aseptic seedling is divided into cotyledon and plumular axis two parts is inoculated into respectively on the callus inducing medium and (sees table 2).Behind 1 first quarter moon, elongation growth appears in cotyledon, becomes 2 true leaves, and a small amount of yellow callus appears in the cotyledon base portion; The plumular axis marginal swell is expanded, and yellow callus of small quantities of particles shape and green callus occur.
Following table 2 data show ZT0.5 mgL
-1To ZT2.0 mgL
-1Be equipped with the NAA (0.1 ~ 1.0mgL of variable concentrations
-1) all can produce by evoked callus, but induce difference on effect bigger.Low concentration ZT (0.5 mgL
-1) inductivity lower, raise with ZT concentration, inductivity becomes to increase progressively trend, when ZT concentration reaches 2.0 mgL
-1The time, inductivity can reach more than 85%.Find ZT2.0 mgL through screening test
-1+ NAA1.0 mgL
-1Combination is better to the callus induction effect, and callus induction rate can reach 100% in the time of 60 days.Therefore, optional 1/4MS+ZT2.0mgL
-1+ NAA1.0 mgL
-1Culture medium prescription carries out callus induction and cultivates.
Different hormone concentrations of table 2 and proportioning are to the influence of callus induction
The callus of Da Bai cuckoo seed embryogenesis need not to induce, and on former medium, can differentiate seedling voluntarily.The callus that first generation is induced continues to cultivate on former medium, and after 2 months, the callus that cotyledon forms differentiates the bud of growing thickly, the callus that plumular axis forms a large amount of green bud points that may be seen indistinctly.For obtaining a large amount of seedlings, need that also grow thickly bud and callus are carried out shoot proliferation and cultivate.On initial culture base basis, optimize and revise, the bud of will growing thickly cuts into single seedling, and callus cuts into fritter, is inoculated into shoot proliferation and cultivates upward (seeing table 3).Following table 3 result shows that callus initial culture base is 1/4MS+ZT2.0 mgL
-1+ NAA1.0 mgL
-1Culture medium prescription is fit to that also the bud of growing thickly is carried out shoot proliferation to be cultivated, and on this medium, after 1 month, the seedling and bud proliferation rate can reach 100%, and reproduction coefficient can reach 6.5; The callus that cotyledon forms increases obviously, and top dissolves the more bud seedling of growing thickly; Plumular axis forms callus and all breaks up the clump that sprouts, but bud is extremely short and small, thin and weak, the elongation and the strong seedling culture of the bud of need growing thickly.
Different hormone concentrations of table 3 and proportioning are to the bud subculture influence of growing thickly
The bud of growing thickly elongation and strong seedling culture
The bud of will growing thickly cuts down from callus, and the bud clump that grows thickly is divided into fritter and is transferred to shown in the table 4 on the medium, observes bud seedling upgrowth situation after 1 month.TDZ0.5 mgL
-1Be equipped with GA1.0 ~ 2.0 mgL
-1Suit the bud of growing thickly is extended strong seedling culture, on this medium, bud seedling robust growth behind the 30d.The individual plant bud of growing thickly is extending to about 2.5 centimetres, and the budlet seedling on the bud clump that grows thickly can rise to 1.0 centimetres.
Different hormone combinations of table 4 and concentration are to the bud elongation and the influence in strong sprout of growing thickly
Culture of rootage
After the bud seedling of growing thickly about high 2～3 cm is cut into individual plant, be inoculated on the root media.
Different hormone combinations of table 5 and concentration are to the tissue cultivating seedling influence of taking root
Top table 5 result shows that NAA content is taken root to Da Bai cuckoo tissue cultivating seedling has very big influence, and low concentration NAA content can not induce tissue cultivating seedling to take root.IBA is taken root to tissue cultivating seedling has certain promotion.Cooperate IBA1.0 mgL at NAA3.0
-1Being fit to tissue cultivating seedling takes root.In medium, add 0.08% active carbon and help to improve the quality of taking root.On this medium, begin to take root behind the tissue cultivating seedling 50d, rooting rate is that height of seedling can reach 4～5 cm behind 60.5%, 70 d, 2～3 of radicals.Whole plant can be transplanted to and refine seedling in the matrix that yellow sand, perlite, humus soil mix and cultivate.
Through above test, obtained technical scheme of the present invention.Through evidence, the present invention compared with prior art, the present invention has cultivation cycle and shortens, is not subjected to seasonal effect, survival rate height and output than advantages such as height.
Embodiment
The present invention is further illustrated below in conjunction with embodiment.
Embodiments of the invention: during a kind of Da Bai cuckoo tissue culture quick propagation culturing method of embodiment of the present invention, comprise adopt the seed embryo cotyledon with plumular axis the grow thickly method for quickly breeding of bud of formation, its process is made up of the following step:
The preparation of a, explant
Get the still uncracked capsule of Da Bai cuckoo; Under flowing water, scrub the capsule surface with brush; On superclean bench with 75% ethanol disinfection 30S; Inoculation is with 0.1% mercuric chloride solution sterilization 10min; Aseptic water washing 1 time soaks 3～5min, aseptic water washing 5～6 times with 10% hydrogenperoxide steam generator again; Blot surface moisture with sterilized filter paper at last, subsequent use;
The foundation of b, aseptic culture system
Capsule has been gone out behind the bacterium, with scissors and tweezers strip off capsule, seed has been trembled on the sterile seed germination medium that is sprinkling upon the bacterium of having gone out, this sterile seed germination medium is 1/4MS medium, sucrose 30g/L, zeatin 0.5mg/L, methyl 0.01mg/L; In the sterile seed germination medium, contain agar 10g/L, pH5.2～5.4; Be that 25 (± 1) ℃, light application time are 12hd in cultivation temperature then
-1With intensity of illumination be 40～60 mo1m
-2S
-1Condition of culture under cultivate;
C, inducing clumping bud and enrichment culture
When the whole sproutings of seed finish; When cotyledon all launches; Aseptic seedling is divided into cotyledon and plumular axis two parts are inoculated into respectively on the inducing clumping bud medium, this inducing clumping bud medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2～2.0mg/L, methyl 1.0mg/L; In the inducing clumping bud medium, contain agar 10g/L, pH5.2～5.4; Its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination be 40～60 mo1m
-2S
-1
Behind the one and a half months, elongation growth appears in cotyledon, becomes 2 true leaves, and a small amount of yellow callus appears in the cotyledon base portion; The plumular axis edge begins protuberance and expands, and yellow callus of small quantities of particles shape and green callus occur; After 2 months; Callus obviously increases; The callus that cotyledon forms differentiates the bud of growing thickly; At this moment; The callus that plumular axis forms a large amount of green bud points that may be seen indistinctly; The callus that bud and plumular axis form of will growing thickly continues to be transferred to and carries out shoot proliferation on the shoot proliferation medium and cultivate, and its shoot proliferation medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2～2.0mg/L, methyl 1.0mg/L; In the shoot proliferation medium, contain agar 10g/L, pH5.2～5.4; Its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1Be 1～February the blanking time that shoot proliferation is cultivated, and its reproduction coefficient can reach 6.5;
The elongation of d, the bud of growing thickly and strong seedling culture
The bud of will growing thickly cuts down from callus; The bud of growing thickly is divided into fritter and is transferred on grow thickly bud elongation and the strong seedling culture base, and this bud of growing thickly extends and the strong seedling culture base is 1/4MS medium, sucrose 30g/L, benzthiazuron 0.5mg/L, gibberellin 1.0～2.0mg/L; In grow thickly bud elongation and strong seedling culture base, contain agar 10g/L, pH5.2～5.4; Its condition of culture is that cultivation temperature is 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1On grow thickly bud elongation and strong seedling culture base, bud seedling robust growth after 30 days, the individual plant bud of growing thickly is extending to about 2 centimetres, and the budlet seedling on the bud clump can rise to 1.0 centimetres;
E, rooting of vitro seedling inducing culture
After the bud seedling of growing thickly of high 2～3 cm is cut into individual plant, be inoculated on the root media, this root media is 1/4MS medium, sucrose 25g/L, indolebutyric acid 1.0mg/L, methyl 3.0mg/L, active carbon 0.8 mg/L; In root media, contain agar 10g/L, pH5.2～5.4; Its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1
Begin behind the 50d to take root, rooting rate is that height of seedling can reach 4～5 cm behind 60%～65%, 70 d, and 2～3 of radicals are transplanted to whole plant and are refined seedling in the matrix that yellow sand, perlite, humus soil mix and cultivate.
Used in the present embodiment MS medium, sucrose, zeatin, methyl, benzthiazuron, gibberellin, indolebutyric acid, active carbon all adopt finished product of the prior art.
Claims (7)
1. Da Bai cuckoo tissue culture quick propagation culturing method; It is characterized in that: adopt tissue culture technique that the cotyledon of seed embryo and plumular axis are carried out callus induction and form a kind of tissue culture quick propagation culturing method that the bud of growing thickly then is divided into whole plant then, its process comprises the following steps:
The preparation of a, explant
Get the still uncracked capsule of Da Bai cuckoo; Under flowing water, scrub the capsule surface with brush; On superclean bench with 75% ethanol disinfection 30S; Inoculation is with 0.1% mercuric chloride solution sterilization 10min; Aseptic water washing 1 time soaks 3～5min, aseptic water washing 5～6 times with 10% hydrogenperoxide steam generator again; Blot surface moisture with sterilized filter paper at last, subsequent use;
The foundation of b, aseptic culture system
Capsule having been gone out behind the bacterium, with scissors and tweezers strip off capsule, seed has been trembled on the sterile seed germination medium that is sprinkling upon the bacterium of having gone out, is that 25 (± 1) ℃, light application time are 12hd in cultivation temperature then
-1With intensity of illumination be 40～60 mo1m
-2S
-1Condition of culture under cultivate;
C, inducing clumping bud and enrichment culture
All sprout when seed and to finish, when cotyledon all launches, aseptic seedling is divided into cotyledon and plumular axis two parts are inoculated into respectively on the inducing clumping bud medium, its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination be 40～60 mo1m
-2S
-1
Behind the one and a half months, elongation growth appears in cotyledon, becomes 2 true leaves, and a small amount of yellow callus appears in the cotyledon base portion; The plumular axis edge begins protuberance and expands, and yellow callus of small quantities of particles shape and green callus occur; After 2 months; Callus obviously increases, and the callus that cotyledon forms differentiates the bud of growing thickly, at this moment; The callus that plumular axis forms a large amount of green bud points that may be seen indistinctly, the callus that will grow thickly bud and plumular axis form continue to be transferred to and carry out the shoot proliferation cultivation on the shoot proliferation medium; Its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1Be 1～February the blanking time that shoot proliferation is cultivated, and its reproduction coefficient can reach 6.5;
The elongation of d, the bud of growing thickly and strong seedling culture
The bud of will growing thickly cuts down from callus, and the bud of growing thickly is divided into fritter and is transferred on grow thickly bud elongation and the strong seedling culture base, and its condition of culture is that cultivation temperature is 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1
On grow thickly bud elongation and strong seedling culture base, bud seedling robust growth after 30 days, the individual plant bud of growing thickly is extending to about 2 centimetres, and the budlet seedling on the bud clump can rise to 1.0 centimetres;
E, rooting of vitro seedling inducing culture
After the bud seedling of growing thickly of high 2～3 cm is cut into individual plant, be inoculated on the root media, its condition of culture is cultivation temperature 25 (± 1) ℃, light application time 12hd
-1With intensity of illumination 40～60 mo1m
-2S
-1
Begin behind the 50d to take root, rooting rate is that height of seedling can reach 4～5 cm behind 60%～65%, 70 d, and 2～3 of radicals are transplanted to whole plant and are refined seedling in the matrix that yellow sand, perlite, humus soil mix and cultivate.
2. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: the sterile seed germination medium is 1/4MS medium, sucrose 30g/L, zeatin 0.5mg/L, methyl 0.01mg/L.
3. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: the inducing clumping bud medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2～2.0mg/L, methyl 1.0mg/L.
4. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: the shoot proliferation medium is 1/4MS medium, sucrose 30g/L, zeatin 1.2 ~ 2.0mg/L, methyl 1.0mg/L.
5. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: the bud of growing thickly elongation and strong seedling culture base are 1/4MS medium, sucrose 30g/L, benzthiazuron 0.5mg/L, gibberellin 1.0～2.0mg/L.
6. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: root media is 1/4MS medium, sucrose 25g/L, indolebutyric acid 1.0mg/L, methyl 3.0mg/L, active carbon 0.8 mg/L.
7. Da Bai cuckoo tissue culture quick propagation culturing method according to claim 1 is characterized in that: in sterile seed germination medium, inducing clumping bud medium, shoot proliferation medium, the bud of growing thickly elongation and strong seedling culture base and root media, all contain agar 10g/L and pH5.2～5.4 respectively.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102792893A (en) * | 2012-08-27 | 2012-11-28 | 云南农业大学 | Tissue culture propagating method of Rhododendron agastum |
| CN103814736A (en) * | 2014-03-07 | 2014-05-28 | 安徽理工大学 | Cuckoo greenhouse seedling thinning transplantation method |
| CN104054429A (en) * | 2014-06-26 | 2014-09-24 | 浙江大学 | Seedling culturing method of rhododendra |
| CN107079815A (en) * | 2017-05-25 | 2017-08-22 | 贵州师范大学 | A kind of method for inducing charming cuckoo and Rhododendron delavayi cenospecies callus |
| CN111134123A (en) * | 2019-12-19 | 2020-05-12 | 东莞市东阳光冬虫夏草中药有限公司 | Seed germination treatment agent and application |
| CN116649215A (en) * | 2023-06-21 | 2023-08-29 | 滨州学院 | Tissue culture and rapid propagation method of phyllanthus niruri |
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| CN102792893A (en) * | 2012-08-27 | 2012-11-28 | 云南农业大学 | Tissue culture propagating method of Rhododendron agastum |
| CN102792893B (en) * | 2012-08-27 | 2013-10-02 | 云南农业大学 | Tissue culture propagating method of Rhododendron agastum |
| CN103814736A (en) * | 2014-03-07 | 2014-05-28 | 安徽理工大学 | Cuckoo greenhouse seedling thinning transplantation method |
| CN104054429A (en) * | 2014-06-26 | 2014-09-24 | 浙江大学 | Seedling culturing method of rhododendra |
| CN104054429B (en) * | 2014-06-26 | 2015-12-09 | 浙江大学 | A kind of spring cuckoo seedling-cultivating method |
| CN107079815A (en) * | 2017-05-25 | 2017-08-22 | 贵州师范大学 | A kind of method for inducing charming cuckoo and Rhododendron delavayi cenospecies callus |
| CN107079815B (en) * | 2017-05-25 | 2019-06-11 | 贵州师范大学 | A method of inducing charming cuckoo and Rhododendron delavayi cenospecies callus |
| CN111134123A (en) * | 2019-12-19 | 2020-05-12 | 东莞市东阳光冬虫夏草中药有限公司 | Seed germination treatment agent and application |
| CN116649215A (en) * | 2023-06-21 | 2023-08-29 | 滨州学院 | Tissue culture and rapid propagation method of phyllanthus niruri |
| CN116649215B (en) * | 2023-06-21 | 2024-04-19 | 滨州学院 | Tissue culture and rapid propagation method of phyllanthus niruri |
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