CN109329062A - A kind of tissue culture technique of cottonrose hibiscus leaf - Google Patents
A kind of tissue culture technique of cottonrose hibiscus leaf Download PDFInfo
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- CN109329062A CN109329062A CN201811428637.5A CN201811428637A CN109329062A CN 109329062 A CN109329062 A CN 109329062A CN 201811428637 A CN201811428637 A CN 201811428637A CN 109329062 A CN109329062 A CN 109329062A
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- culture
- cottonrose hibiscus
- hibiscus leaf
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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
-
- 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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- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a kind of tissue culture technique of cottonrose hibiscus leaf, cottonrose hibiscus leaf is malvaceae plant, the dried leaf of cotton rose.Main product is in wash rice river, Jiangsu.The harvesting of two season of summer and autumn, removes impurity, dries or dry.Gas is micro-, lightly seasoned.With removing toxicity for detumescence, cool blood, analgesic.For pain subcutaneous ulcer boil, venomous snake bite, cream pain, traumatic injury, burn and scald.Cottonrose hibiscus leaf obtains the method for culturing seedlings of high quality seedling by rapid propagation in vitro technology.The present invention is using cottonrose hibiscus leaf stem with bud as explant, by processes such as explant disinfection, inducing clumping bud, culture of rootage, acclimatization and transplants to establish cottonrose hibiscus leaf tissue-culturing quick-propagation system, the popularization of cottonrose hibiscus leaf breeding and resources development and utilization is accelerated to have important practical significance.
Description
Technical field
The present invention relates to the methods of Plant Tissue Breeding in Plant Biotechnology, specifically, being related to a kind of cottonrose hibiscus leaf
Tissue culture technique.
Background technique
Cottonrose hibiscus leaf is malvaceae plant, the dried leaf of cotton rose.Main product is in wash rice river, Jiangsu.The harvesting of two season of summer and autumn, removes impurity elimination
Matter is dried or is dried.Gas is micro-, lightly seasoned.With removing toxicity for detumescence, cool blood, analgesic.For pain subcutaneous ulcer boil, venomous snake bite, cream pain is fallen
Beat damage, burn and scald.Cottonrose hibiscus leaf mainly carries out sapling multiplication by seed, but since seminal propagation characters of progenies separates,
It is unfavorable for high-quality plasm resource protection, constrains the popularizing planting of high-quality cottonrose hibiscus leaf.Therefore, it is necessary to establish the tissue of cottonrose hibiscus leaf
Culture technique quickly breeds for its breeding and provides technical support and guarantee, also to meet the needs of China is to cottonrose hibiscus leaf seedling, adds
Fast its resource expansion has important practical significance.
Summary of the invention
The purpose of the present invention is to provide a kind of tissue culture techniques of cottonrose hibiscus leaf out, and the present invention is with cottonrose hibiscus leaf stem with bud
For explant, cottonrose hibiscus leaf group is established by processes such as explant disinfection, inducing clumping bud, culture of rootage, acclimatization and transplants
Culture rapid propagation system is knitted, to realize the purpose of the present invention.
A kind of tissue culture technique of cottonrose hibiscus leaf of the invention, includes the following steps:
Step 1, inducing clumping bud: choosing robust growth without the treelet current year of disease pest raw stem with bud as explant, with washing
Clean essence aqueous solution soaking 12min, then rinses 32min with tap water, dry it is spare after surface moisture, in superclean bench with
70% ethyl alcohol impregnates 12s, and 0.1% mercuric chloride solution sterilizes 22min, then with sterile water wash 8 times, and it is 2.2cm that length is cut into after drying
Stem section be inoculated into induced medium, first it is dark culture 25 days full under the conditions of 25 ~ 28 DEG C, it is small to be subsequently placed in daily illumination 12
When, intensity of illumination 1800lx, cultivation temperature is cultivated under the conditions of being 23 DEG C, until induced synthesis Multiple Buds；Induced medium are as follows:
MS+8mg/L6-BA+3mg/L NAA+0.7mmol/L La(NO3)2+ 0.2% active carbon of+0.6% agar of+4.0% sucrose, pH value are
5.6;
Step 2, Multiplying culture: the resulting Multiple Buds of step 1 are transferred on proliferated culture medium and carry out Multiplying culture, are first existed after inoculation
Full dark culture germinates a lateral bud at axil under the conditions of 23 DEG C, is subsequently placed in daily illumination 18 hours, intensity of illumination is
2000lx, culture under conditions of cultivation temperature is 23 DEG C is until each Multiple Buds are elongated to 5cm；Proliferated culture medium are as follows: MS+3mg/
+ 0.3% active carbon of+0.8% agar of L6-BA+0.6mg/L NAA+5.2% sucrose, pH value 5.6;
Step 3, culture of rootage: the Multiple Buds bud that step 1 or step 2 height obtained are about 4cm is cut and is inoculated into life
Culture of rootage is carried out on root culture medium, it is first dark culture 12 days full under the conditions of 23 DEG C after inoculation, it is small to be subsequently placed in daily illumination 14
When, intensity of illumination 3200lx, cultivation temperature is cultivated under conditions of being 23 DEG C to taking root；Root media are as follows: 1/2MS+6mg/L
+ 0.2% active carbon of+0.8% agar of NAA+4.0% sucrose, pH value 5.6;
Step 4, acclimatization and transplants: after taking root in cottonrose hibiscus leaf bottle 30 days, after being placed in natural lighting lower refining seedling 10 days, root culture is cleaned
Base is transplanted by yellow soil: in the matrix that river sand=3:1 is mixed into.
Compared with prior art the invention has the advantages that the present invention obtains the nursery of high quality seedling by rapid propagation in vitro technology
Method.Using cottonrose hibiscus leaf stem with bud as explant, waited by explant disinfection, inducing clumping bud, culture of rootage, acclimatization and transplants
Journey accelerates the popularization of cottonrose hibiscus leaf breeding and resources development and utilization to have to establish cottonrose hibiscus leaf tissue-culturing quick-propagation system
Important realistic meaning.
Specific embodiment
It the following examples are further illustrations of the invention, is not limitation of the present invention.
Embodiment 1:
(1) inducing clumping bud: the stem with bud of treelet current year life of the robust growth without disease pest is chosen as explant, uses dish washing liquid
Then aqueous solution soaking 10min rinses 12min with tap water, dry spare after surface moisture.In superclean bench with 75%
Ethyl alcohol impregnates 2s, and 0.1% mercuric chloride solution sterilizes 15min, then with sterile water wash 6 times, and the stem that length is 2.5cm is cut into after drying
Section is inoculated into induced medium, first dark culture 16 days full under the conditions of 26 DEG C, is subsequently placed in daily illumination 12 hours, illumination is strong
Degree be 1500x, cultivation temperature be 26 DEG C under the conditions of cultivate 41 days can induced synthesis Multiple Buds, inductivity 70%.The induction
Culture medium is MS+5mg/L6-BA+1.0mg/L NAA+0.5mmol/L La (NO3)2+ 0.55% agar+0.15% of+3.5% sucrose is living
Property charcoal, pH value 5.8.
(2) Multiplying culture: step (1) resulting Multiple Buds are transferred on proliferated culture medium and carry out Multiplying culture, after inoculation
First dark culture full under the conditions of 26 DEG C germinates a lateral bud at axil, is subsequently placed in daily illumination 15 hours, intensity of illumination
For 1500lx, cultivation temperature, which cultivates 32 days each Multiple Buds under conditions of being 26 DEG C, can be elongated to 6cm, proliferation rate 6.0.It is described
Proliferated culture medium is+0.2% active carbon of+0.55% agar of MS+3mg/L6-BA+0.6mg/L NAA+3.0% sucrose, pH value 5.8.
(3) culture of rootage: the Multiple Buds bud that step (1) or step (2) height obtained are about 6cm is cut and is inoculated with
Culture of rootage is carried out on to root media, it is first dark culture 8 days full under the conditions of 26 DEG C after inoculation, it is subsequently placed in daily illumination 16
Hour, intensity of illumination 2000lx, cultivation temperature is cultivated 41 days and can be taken root under conditions of being 26 DEG C, rooting rate 72%.
(4) acclimatization and transplants: after taking root in cottonrose hibiscus leaf bottle 25 days, after being placed in natural lighting lower refining seedling 6 days, root culture is cleaned
Base is transplanted by yellow soil: in the matrix that river sand=2:1 is mixed into.Transplanting survival rate is 88%.
Embodiment 2:
(1) inducing clumping bud: the stem with bud of treelet current year life of the robust growth without disease pest is chosen as explant, uses dish washing liquid
Then aqueous solution soaking 15min rinses 18min with tap water, dry spare after surface moisture.In superclean bench with 75%
Ethyl alcohol impregnates 16s, and 0.1% mercuric chloride solution sterilizes 16min, then with sterile water wash 8 times, and the stem that length is 2.6cm is cut into after drying
Section is inoculated into induced medium, first dark culture 20 days full under the conditions of 27 DEG C, is subsequently placed in daily illumination 16 hours, illumination is strong
Degree be 1600x, cultivation temperature be 27 DEG C under the conditions of cultivate 40 days can induced synthesis Multiple Buds, inductivity 76%.The induction
Culture medium is MS+6mg/L6-BA+1.1mg/L NAA+0.5mmol/L La (NO3)2+ 0.2% activity of+3.6%+0.5% agar of sucrose
Charcoal, pH value 5.9.
(2) Multiplying culture: step (1) resulting Multiple Buds are transferred on proliferated culture medium and carry out Multiplying culture, after inoculation
First dark culture full under the conditions of 28 DEG C germinates a lateral bud at axil, is subsequently placed in daily illumination 15 hours, intensity of illumination
For 1600lx, cultivation temperature, which cultivates 28 days each Multiple Buds under conditions of being 27 DEG C, can be elongated to 5cm, proliferation rate 5.5.It is described
Proliferated culture medium is+0.2% active carbon of+0.1% agar of MS+2mg/L6-BA+0.8mg/L NAA+2.5% sucrose, pH value 5.9.
(3) culture of rootage: the Multiple Buds bud that step (1) or step (2) height obtained are about 4cm is cut and is inoculated with
Culture of rootage is carried out on to root media, it is first dark culture 8 days full under the conditions of 27 DEG C after inoculation, it is subsequently placed in daily illumination 10
Hour, intensity of illumination 1900lx, cultivation temperature is cultivated 36 days and can be taken root under conditions of being 27 DEG C, rooting rate 82%.
(4) acclimatization and transplants: after taking root in cottonrose hibiscus leaf bottle 25 days, after being placed in natural lighting lower refining seedling 9 days, root culture is cleaned
Base is transplanted by yellow soil: in the matrix that river sand=3:2 is mixed into.Transplanting survival rate is 92%.
Claims (1)
1. a kind of tissue culture technique of cottonrose hibiscus leaf, it is characterised in that the following steps are included:
Step 1, inducing clumping bud: choosing robust growth without the treelet current year of disease pest raw stem with bud as explant, with washing
Clean essence aqueous solution soaking 12min, then rinses 32min with tap water, dry it is spare after surface moisture, in superclean bench with
70% ethyl alcohol impregnates 12s, and 0.1% mercuric chloride solution sterilizes 22min, then with sterile water wash 8 times, and it is 2.2cm that length is cut into after drying
Stem section be inoculated into induced medium, first it is dark culture 25 days full under the conditions of 25 ~ 28 DEG C, it is small to be subsequently placed in daily illumination 12
When, intensity of illumination 1800lx, cultivation temperature is cultivated under the conditions of being 23 DEG C, until induced synthesis Multiple Buds；Induced medium are as follows:
MS+8mg/L6-BA+3mg/L NAA+0.7mmol/L La(NO3)2+ 0.2% active carbon of+0.6% agar of+4.0% sucrose, pH value are
5.6;
Step 2, Multiplying culture: the resulting Multiple Buds of step 1 are transferred on proliferated culture medium and carry out Multiplying culture, are first existed after inoculation
Full dark culture germinates a lateral bud at axil under the conditions of 23 DEG C, is subsequently placed in daily illumination 18 hours, intensity of illumination is
2000lx, culture under conditions of cultivation temperature is 23 DEG C is until each Multiple Buds are elongated to 5cm；Proliferated culture medium are as follows: MS+3mg/
+ 0.3% active carbon of+0.8% agar of L6-BA+0.6mg/L NAA+5.2% sucrose, pH value 5.6;
Step 3, culture of rootage: the Multiple Buds bud that step 1 or step 2 height obtained are about 4cm is cut and is inoculated into life
Culture of rootage is carried out on root culture medium, it is first dark culture 12 days full under the conditions of 23 DEG C after inoculation, it is small to be subsequently placed in daily illumination 14
When, intensity of illumination 3200lx, cultivation temperature is cultivated under conditions of being 23 DEG C to taking root；Root media are as follows: 1/2MS+6mg/L
+ 0.2% active carbon of+0.8% agar of NAA+4.0% sucrose, pH value 5.6;
Step 4, acclimatization and transplants: after taking root in cottonrose hibiscus leaf bottle 30 days, after being placed in natural lighting lower refining seedling 10 days, root culture is cleaned
Base is transplanted by yellow soil: in the matrix that river sand=3:1 is mixed into.
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| CN201811428637.5A CN109329062A (en) | 2018-11-27 | 2018-11-27 | A kind of tissue culture technique of cottonrose hibiscus leaf |
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| CN201811428637.5A CN109329062A (en) | 2018-11-27 | 2018-11-27 | A kind of tissue culture technique of cottonrose hibiscus leaf |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686341A (en) * | 2015-02-22 | 2015-06-10 | 梁仕华 | Tissue culture technique of aquilaria sinensis |
| CN107864862A (en) * | 2017-12-18 | 2018-04-03 | 曲靖师范学院 | A kind of method for rapidly and efficiently obtaining kapok tissue-cultured seedling |
| CN108077067A (en) * | 2016-11-21 | 2018-05-29 | 四川七彩林业开发有限公司 | A kind of tissue culture and rapid propagation method of cotton rose |
| CN108541592A (en) * | 2018-05-18 | 2018-09-18 | 象州县科学技术情报研究所 | The tissue culture mating system of kapok |
-
2018
- 2018-11-27 CN CN201811428637.5A patent/CN109329062A/en not_active Withdrawn
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686341A (en) * | 2015-02-22 | 2015-06-10 | 梁仕华 | Tissue culture technique of aquilaria sinensis |
| CN108077067A (en) * | 2016-11-21 | 2018-05-29 | 四川七彩林业开发有限公司 | A kind of tissue culture and rapid propagation method of cotton rose |
| CN107864862A (en) * | 2017-12-18 | 2018-04-03 | 曲靖师范学院 | A kind of method for rapidly and efficiently obtaining kapok tissue-cultured seedling |
| CN108541592A (en) * | 2018-05-18 | 2018-09-18 | 象州县科学技术情报研究所 | The tissue culture mating system of kapok |
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Application publication date: 20190215 |