CN105815215A - Tissue culture propagation method for lysimachia foenum-graecum - Google Patents
Tissue culture propagation method for lysimachia foenum-graecum Download PDFInfo
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- CN105815215A CN105815215A CN201610162849.8A CN201610162849A CN105815215A CN 105815215 A CN105815215 A CN 105815215A CN 201610162849 A CN201610162849 A CN 201610162849A CN 105815215 A CN105815215 A CN 105815215A
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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 discloses a tissue culture propagation method for lysimachia foenum-graecum.The method comprises the following steps of 1, explant obtaining and disinfection, 2, obtaining of a sterile test-tube plantlet, 3, test-tube plantlet enrichment culture, 4, test-tube plantlet rooting culture and 5, test-tube plantlet acclimatization and transplanting.A stem with buds is used as an explant; by means of tissue culture propagation, the culture period can be shortened, and high-quality lysimachia foenum-graecum seedlings can be rapidly propagated.The bud propagation coefficient reaches 9.5-13.2 times after 30d of culture, the tissue culture seedling rooting rate reaches 98.7-100% through rooting culture, the survival rate is 91.3-96% after a matrix is transplanted, and the large-scale seedling culture problem is effectively solved.
Description
Technical field
The present invention relates to plant tissue culture reproduction technique field, the tissue culture propagation of a kind of Herba Lysimachiae foenum-graeci.
Background technology
Herba Lysimachiae foenum-graeci (Classification system: Lysimachiafoenum-graecumHance) has another name called Rhizoma et radix valerianae, Herba Eupatorii, Herba lysimachiae capillipedis etc., for Primulaceae Lysimachia herbaceos perennial.Grass, containing similar coumarin aromatic oil, can refine essence, as the spice such as Nicotiana tabacum L. and face cream；Can Anti-moth-eating medicated clothing in dry product cartonning；Hyoscine again, has dispersing wind and cold, wards off dirty effect, control seasonal pathogen headache due to common cold, upper qi-stagnant lumbago, abdominal distention uncomfortable in chest, seminal emission, ascarifuge etc., be famous and precious fragrant plant.
Good effect in view of Herba Lysimachiae foenum-graeci, being continuously increased of market demand, for preferably development and utilization Chinese medicine Herba Lysimachiae foenum-graeci resource, guarantee the sustainable use of Herba Lysimachiae foenum-graeci resource, accordingly, by modern biotechnology, it is carried out tissue culture rapid propagating technology to be studied, it is intended to obtain the seedling of a large amount of high-quality, produce for Herba Lysimachiae foenum-graeci and provide kind of a source, with the needs in satisfied production.Though the report in terms of existing Herba Lysimachiae foenum-graeci tissue culture, this technology breeding coefficient is higher, and production cost is lower, is more suitable for large-scale production.
Summary of the invention
It is an object of the invention to provide the tissue culture propagation of a kind of Herba Lysimachiae foenum-graeci, the problem bred for Herba Lysimachiae foenum-graeci, pass through tissue culture propagating, it is possible to shorten cultivation cycle and Fast-propagation goes out Herba Lysimachiae foenum-graeci high quality seedling, carry out large-scale production seedling, meet the market demand.
For solving above-mentioned technical problem, the technical scheme is that the tissue culture propagation of a kind of Herba Lysimachiae foenum-graeci, comprise the following steps:
null(1) the drawing materials and sterilizing of outer implant: take Herba Lysimachiae foenum-graeci and cut into the stem with bud of 1-2cm，Carry out disinfection as outer implant，First 1-2 is dripped in the beaker that liquid detergent instills equipped with 50ml tap water，Outer implant is put into beaker and is gently mixed 5min，Outer planting surface dirt is cleaned again with Cotton Gossypii，The slight flowing water of tap water rinses 8-10min，Move to superclean bench，By 75v/v% soak with ethanol 30s，Aseptic water washing one time，It is that 0.1v/v% mercuric chloride is sterilized 6-8min by the 50ml concentration that with the addition of 1-2 and drip tween 20 the most again、Aseptic water washing 3-5 time，Finally it is placed in the rustless steel plate disinfected，Cut into the stem with bud of 0.5-1.0cm length，Obtain aseptic explant，Wherein sterilized water is the distilled water through high high-temperature high-pressure sterilizing；
(2) acquisition of in vitro cuttings: aseptic explant is placed in inducing culture carry out Induce aerosor germinate obtain in vitro cuttings, described inducing culture is with MS as minimal medium, add 5g L-1Agar, 30g L-1Sucrose, 0.5mg L-16-benzyladenine, 0.2mg L-1Naphthalene acetic acid, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 15-20d；
(3) test tube seedling proliferation is cultivated: being placed in proliferated culture medium by vitro cuttings and carry out cultivating a large amount of adventitious buds of acquisition, described proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 1.0-2.0mg L-16-benzyladenine, 0.1-0.5mg L-1Naphthalene acetic acid, 0.1-0.5mg L-1Heteroauxing, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 30d；
(4) rooting of vitro seedling cultivate: the adventitious bud list of 2-3cm is cut be placed in root media cultivation obtain complete band root, described root media is with 1/2MS as minimal medium, add 5g L-1Agar, 20g L-1Sucrose, 0.2-1.0mg L-1Naphthalene acetic acid, 0.1-0.5mg L-1Indolebutyric acid, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 25d；
(5) test tube Seedling acclimatization and transplants: screen to transplant after complete band root carries out seedling exercising and cultivate in substrate, then transplant to land for growing field crops.
Step (5) described acclimatization and transplants is carried out by following operation: obtain the whole plant of band root through step (4), treat that Seedling length is to 4-5cm, when root length is to 2-4cm, the bottle Seedling that growth selection is good, neat, healthy and strong, carry out seedling exercising in the indoor that room temperature is 23-25 DEG C, and add water in bottle and to flood culture medium, seedling exercising 5-7d, takes out test tube Seedling, cleans root culture medium, it is transplanted in substrate grow 40-50d, then transplants to land for growing field crops.
Described substrate is by weight for peat soil: the ratio of yellow sand=1:1 mixes.
The present invention has the prominent advantages that:
<1>by induction and the breeding of Herba Lysimachiae foenum-graeci adventitious bud, the character of original variety can be kept, it is possible to breeding in a short time obtains the plant that a large amount of character is consistent, meet and produce needs, and for providing the Herba Lysimachiae foenum-graeci industrial seedling rearing of merit to provide reference.
<2>used by, MS and 1/2MS culture medium comprises macro-and microelements, the chemical agents such as 6-benzyladenine, naphthalene acetic acid, heteroauxing and the indolebutyric acid added, lower cost, concentration are suitable, and concrete effect is as follows: add 6-BA, NAA and IAA in proliferated culture medium and can promote the propagation of bud；In 1/2MS root media, add NAA and IBA can promote seedling rooting, transplant to the nutrient cup filling substrate after seedling exercising.
<3>cultivating 30d bud breeding coefficient and reach 9.5～13.2 times, through root culture, tissue cultured seedling rooting rate reaches 98.7～100%, and after transplanting medium, survival rate is 91.3～96%, efficiently solves the scale breeding problem of Herba Lysimachiae foenum-graeci.
Detailed description of the invention
Below in conjunction with embodiment, technical scheme is further illustrated.
Embodiment 1
One example of the tissue culture propagation of Herba Lysimachiae foenum-graeci of the present invention, comprises the steps:
(1) the drawing materials and sterilizing of outer implant: take Herba Lysimachiae foenum-graeci and cut into the stem with bud of 1-2cm, carry out disinfection as outer implant, first 1-2 is dripped in the beaker that liquid detergent instills equipped with 50ml tap water, outer implant is put into beaker and is gently mixed 5min, cleaning outer planting surface dirt with Cotton Gossypii again, the slight flowing water of tap water rinses 8-10min；Dislocation superclean bench, by 75v/v% soak with ethanol 30s, aseptic water washing one time, it is 0.1v/v% mercuric chloride sterilization 6-8min, aseptic water washing 3-5 time by the 50ml concentration that with the addition of 1-2 and drip tween 20 the most again, finally it is placed on the rustless steel plate disinfected, cutting into the stem with bud of 0.5-1.0cm length, it is thus achieved that aseptic explant, wherein sterilized water is the distilled water through high high-temperature high-pressure sterilizing；
(2) acquisition of in vitro cuttings: the outer implant disinfected is placed in inducing culture and carries out Induce aerosor germination, it is 23-25 DEG C in temperature, intensity of illumination 1500-2000lux, light application time is cultivation 15-20d under conditions of 10-12h/d, obtains in vitro cuttings；
(3) test tube seedling proliferation is cultivated: is placed in proliferated culture medium by vitro cuttings and cultivates, it is 23-25 DEG C in temperature, intensity of illumination 1500-2000lux, light application time is cultivation 30d under conditions of 10-12h/d, obtaining a large amount of adventitious bud, adventitious bud proliferation coefficient is 9.5；
(4) rooting of vitro seedling is cultivated: is cut by the adventitious bud list of 2-3cm and is placed in 1/2MS root media cultivation 25d, it is 23-25 DEG C in temperature, intensity of illumination 1500-2000lux, light application time is cultivation 25d under conditions of 10-12h/d, obtaining complete band root, rooting rate is 98.7%；
(5) test tube Seedling acclimatization and transplants: the complete band root obtained through step (4), treat Seedling length to 4-5cm, during root length to 3-5cm, the bottle Seedling that growth selection is good, neat, healthy and strong, seedling exercising is carried out in the indoor that room temperature is 23-25 DEG C, and in bottle, add a small amount of tap water flood culture medium, seedling exercising 5-7d, takes out test tube Seedling with tweezers, clean root culture medium, being transplanted in substrate grow 40-50d, then transplant to land for growing field crops, survival rate is 91.3%.Described substrate is in peat soil: the mixing of the ratio of yellow sand=1:1, and substrate loose ventilative, draining is good.
Wherein, the culture medium prescription that each stage uses is as follows:
Described inducing culture is with MS as minimal medium, adds 5g L-1Agar, 30g L-1Sucrose, 1.0mg L-16-benzyladenine, Medium's PH Value is 5.8；
Described proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 1.0mg L-16-benzyladenine, 0.1mg L-1Naphthalene acetic acid, 0.1mg L-1Heteroauxing, Medium's PH Value is 5.8；
Described root media is with 1/2MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 0.2mg L-1Naphthalene acetic acid, 0.1mg L-1Indolebutyric acid, Medium's PH Value is 5.8.
Embodiment 2
Another example of the tissue culture propagation of Herba Lysimachiae foenum-graeci of the present invention, comprises the steps:
Other conditions are with embodiment 1, and only herein below is different:
In step (3), proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 1.5mg L-16-benzyladenine, 0.3mg L-1Naphthalene acetic acid, 0.3mg L-1Heteroauxing, Medium's PH Value is 5.8；Adventitious bud proliferation coefficient is 13.2；
In step (4), root media is with 1/2MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 0.3g L-1Activated carbon, 0.5mg L-1Naphthalene acetic acid, 0.3mg L-1Indolebutyric acid, Medium's PH Value is 5.8, and rooting rate is 99.6%.
In step (5), survival rate is 93%.
Embodiment 3
Another example of the tissue culture propagation of Herba Lysimachiae foenum-graeci of the present invention, comprises the steps:
Other conditions are with embodiment 1, and only herein below is different:
In step (3), proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 1.5mg L-16-benzyladenine, 0.3mg L-1Naphthalene acetic acid, 0.5mg L-1Heteroauxing, Medium's PH Value is 5.8；Adventitious bud proliferation coefficient is 12.5；
In step (4), root media is with 1/2MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 0.3g L-1Activated carbon, Medium's PH Value is 5.8, and adds 1.0mg L-1Naphthalene acetic acid, 0.3mg L-1Indolebutyric acid, rooting rate is 100%.
In step (5), survival rate is 96%.
Embodiment 4
Another example of the tissue culture propagation of Herba Lysimachiae foenum-graeci of the present invention, comprises the steps:
Other conditions are with embodiment 1, and only herein below is different:
In step (3), proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 2.0mg L-16-benzyladenine, 0.5mg L-1Naphthalene acetic acid, 0.5mg L-1Heteroauxing, Medium's PH Value is 5.8, and adventitious bud proliferation coefficient is 6.2；
In step (4), root media is with 1/2MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 0.5g L-1Activated carbon, 1.0mg L-1Naphthalene acetic acid, 0.5mg L-1Indolebutyric acid, Medium's PH Value is 5.8, and rooting rate is 100%.
In step (5), survival rate is 95.7%.
Claims (3)
1. the tissue culture propagation of a Herba Lysimachiae foenum-graeci, it is characterised in that comprise the following steps:
null(1) the drawing materials and sterilizing of outer implant: take Herba Lysimachiae foenum-graeci and cut into the stem with bud of 1-2cm，Carry out disinfection as outer implant，First 1-2 is dripped in the beaker that liquid detergent instills equipped with 50ml tap water，Outer implant is put into beaker and is gently mixed 5min，Outer planting surface dirt is cleaned again with Cotton Gossypii，The slight flowing water of tap water rinses 8-10min，Move to superclean bench，By 75v/v% soak with ethanol 30s，Aseptic water washing one time，It is that 0.1v/v% mercuric chloride is sterilized 6-8min by the 50ml concentration that with the addition of 1-2 and drip tween 20 the most again、Aseptic water washing 3-5 time，Finally it is placed in the rustless steel plate disinfected，Cut into the stem with bud of 0.5-1.0cm length，Obtain aseptic explant，Wherein sterilized water is the distilled water through high high-temperature high-pressure sterilizing；
(2) acquisition of in vitro cuttings: aseptic explant is placed in inducing culture carry out Induce aerosor germinate obtain in vitro cuttings, described inducing culture is with MS as minimal medium, add 5g L-1Agar, 30g L-1Sucrose, 0.5mg L-16-benzyladenine, 0.2mg L-1Naphthalene acetic acid, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 15-20d；
(3) test tube seedling proliferation is cultivated: being placed in proliferated culture medium by vitro cuttings and carry out cultivating a large amount of adventitious buds of acquisition, described proliferated culture medium is with MS as minimal medium, adds 5g L-1Agar, 20g L-1Sucrose, 1.0-2.0mg L-16-benzyladenine, 0.1-0.5mg L-1Naphthalene acetic acid, 0.1-0.5mg L-1Heteroauxing, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 30d；
(4) rooting of vitro seedling cultivate: the adventitious bud list of 2-3cm is cut be placed in root media cultivation obtain complete band root, described root media is with 1/2MS as minimal medium, add 5g L-1Agar, 20g L-1Sucrose, 0.2-1.0mg L-1Naphthalene acetic acid, 0.1-0.5mg L-1Indolebutyric acid, Medium's PH Value is 5.8, and condition of culture is: temperature is 22-24 DEG C, intensity of illumination 1500-2000lux, and light application time is 10-12h/d, and incubation time is 25d；
(5) test tube Seedling acclimatization and transplants: screen to transplant after complete band root carries out seedling exercising and cultivate in substrate, then transplant to land for growing field crops.
The tissue culture propagation of Herba Lysimachiae foenum-graeci the most according to claim 1, it is characterized in that, step (5) described acclimatization and transplants is carried out by following operation: obtain the whole plant of band root through step (4), treat that Seedling length is to 4-5cm, when root length is to 2-4cm, the bottle Seedling that growth selection is good, neat, healthy and strong, seedling exercising is carried out in the indoor that room temperature is 23-25 DEG C, and add water in bottle and to flood culture medium, seedling exercising 5-7d, takes out test tube Seedling, cleans root culture medium, it is transplanted in substrate grow 40-50d, then transplants to land for growing field crops.
The tissue culture propagation of Herba Lysimachiae foenum-graeci the most according to claim 2, it is characterised in that: described substrate is by weight for peat soil: the ratio of yellow sand=1:1 mixes.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110800561A (en) * | 2019-11-14 | 2020-02-18 | 韦东 | Improvement method of arrowhead variety |
| CN116806702A (en) * | 2023-07-26 | 2023-09-29 | 新疆中亚果树产业研究院有限公司 | Method for rapid propagation of vanilla tissue |
-
2016
- 2016-03-21 CN CN201610162849.8A patent/CN105815215A/en active Pending
Non-Patent Citations (2)
| Title |
|---|
| 张宏志等: ""过路黄的组织培养"", 《湖南农业科学》 * |
| 黎海利等: ""聚花过路黄的组织培养和快速繁殖"", 《江苏农业科学》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110800561A (en) * | 2019-11-14 | 2020-02-18 | 韦东 | Improvement method of arrowhead variety |
| CN116806702A (en) * | 2023-07-26 | 2023-09-29 | 新疆中亚果树产业研究院有限公司 | Method for rapid propagation of vanilla tissue |
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Application publication date: 20160803 |