CN109964818B - Sterile Elaeagnus mollis oil seedling GA3Rooting medium and preparation method thereof - Google Patents
Sterile Elaeagnus mollis oil seedling GA3Rooting medium and preparation method thereof Download PDFInfo
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- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
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Abstract
The invention discloses a samara oil aseptic seedling GA3A rooting culture medium and a preparation method thereof,belongs to the technical field of tissue culture and rapid propagation and detoxification of forest plants. The rooting medium per liter comprises the following components in percentage by weight: 1/2MS, indolebutyric acid 0.2-1.0mg/L, naphthylacetic acid 0.005-0.03mg/L, gibberellin 0.1-0.5mg/L, potato juice 100-200g/L, sucrose 10-30g/L, agar powder 2-10g/L, and purified water to balance. According to the technical scheme, the culture medium is more favorable for rooting induction of the samara oil sterile seedlings by optimizing various components of the culture medium, particularly adding and adjusting the concentrations of hormones of indolebutyric acid, naphthylacetic acid and gibberellin in the culture medium, and the aseptic seedlings are high in rooting rate, fast in rooting, developed in root system, good in seedling lignification and high in transplanting survival rate. The invention has simple formula, convenient use and good effect.
Description
Technical Field
The invention relates to the technical field of tissue culture and rapid propagation and detoxification of forest plants, in particular to a samara oil aseptic seedling GA3A rooting culture medium and a preparation method thereof.
Background
The plant test-tube plantlet technology is a sterile tissue culture technology, and the tissue culture medium and the cell totipotency principle can dedifferentiate plant tissues which are in a differentiation terminal or are differentiating, induce the plant tissues to form callus tissues, and form new cluster buds on the callus tissues. Namely, a technique of placing living isolated organs (such as roots, stems, leaves, stem segments, protoplasts), tissues or cells in a culture medium under artificially created aseptic conditions and placing the culture medium in an appropriate environment for continuous culture to obtain cells, tissues or individuals. The tissue culture technology is widely applied to the aspects of improving crops, propagating plants, synthesizing compounds and the like, has unique production method, and is used for large-scale artificial culture and industrial production under the conditions of artificial aseptic operation and near-temperature environment, so the tissue culture technology has special effects on quality guarantee, purity preservation and off-season production of food resources, and has important effects in industrial production of agricultural products.
The Elaeagnus mollis (Latin name) is a deciduous tree or shrub of the genus Elaeagnus of the family Elaeagnaceae. It is mainly distributed in the low mountains and hilly areas in the southwest of the mountains and the counties of the northern foot of the Qin mountain in Shaanxi, and is an ancient biological plant which remains after the action of glaciers in the fourth century. The natural forest of the global pterocarpus heterophyllus oil tree only survives in small parts of areas such as Shanxi county and is a rare plant.
After we have obtained sterile seedlings by germination of seeds, the sterile seedlings need to be transferred to a new medium to induce their rooting. The conditions under which woody and herbaceous plants root vary widely. In most cases, induction of root is easier for herbaceous plants and difficult for woody plants. The elaeagnus mollis is just woody organism, and rooting conditions have two points, namely low salt and the guarantee of auxin at a certain concentration. The rootless seedlings are soaked by indolebutyric acid (IBA) to promote rooting. Branches are picked in the field of the samara oil, the branches are difficult to root even if the branches are disinfected, aseptic seedlings are easy to root, tender aseptic seedlings are adopted, and a proper culture medium is matched to induce the aseptic seedlings to root, and the aseptic seedlings can be used for transplanting after the samara oil.
The Chinese patent with the application number of 200510114666.0 discloses a propagation method of test-tube plantlets of elaeagnus mollis. The method comprises the following steps: 1) cutting stem, root and cotyledon of the seed seedling without strain to obtain explant induced callus; 2) placing the callus on a pre-differentiation culture medium for pre-differentiation; 3) placing the callus subjected to pre-differentiation culture in the step 2) on a differentiation culture medium for adventitious bud differentiation; 4) placing the callus with the adventitious bud primordium grown in the step 3) on a bud elongation culture medium to promote the growth of buds; 5) and (3) carrying out rooting culture on the plantlets obtained in the step 4) by using an indolebutyric acid solution soaking method or a micro-cuttage method. However, the method has low seedling rooting rate, and the maximum rooting rate is 60%. Therefore, the rooting medium which is low in cost, simple, convenient and quick is provided to improve the rooting rate and the average root number of the samara oil aseptic seedlings.
Disclosure of Invention
Aiming at the defects in the prior art, the invention aims to provide a rooting medium which can effectively promote the rooting of the samara oil aseptic seedlings and has low cost, simplicity, convenience and rapidness and a preparation method thereof according to the establishment process and characteristics of the samara oil tree test-tube seedlings so as to improve the rooting rate and the average root number of the samara oil aseptic seedlings.
In order to achieve the purpose, the invention provides the following technical scheme:
sterile Elaeagnus mollis oil seedling GA3The rooting medium comprises the following components in percentage by liter: 1/2MS, indolebutyric acid 0.2-1.0mg/L, naphthylacetic acid 0.005-0.03mg/L, gibberellin 0.1-0.5mg/L, potato juice 100-200g/L, sucrose 10-30g/L, agar powder 2-10g/L, and purified water to balance.
As a still further scheme of the invention: the components of each liter of rooting culture medium and the contents of the components are as follows: 1/2MS, 0.4-0.6mg/L indolebutyric acid, 0.01-0.03mg/L naphthylacetic acid, 0.1-0.3mg/L gibberellin, 180g/L potato juice, 15-25g/L sucrose, 4-6g/L agar powder and pure water to make up the balance.
As a still further scheme of the invention: the components of each liter of rooting culture medium and the contents of the components are as follows: 1/2MS, indolebutyric acid 0.5mg/L, naphthylacetic acid 0.01mg/L, gibberellin 0.15mg/L, potato juice 150g/L, sucrose 20g/L, agar powder 5g/L, and purified water in balance.
As a still further scheme of the invention: sterile Elaeagnus mollis oil seedling GA3The preparation method of the rooting medium comprises the following steps:
weighing agar powder according to the raw material ratio, adding the agar powder into a pot filled with purified water, and boiling; dissolving 0.1 liter of 10 times of 1/2MS mother liquor with indolebutyric acid, naphthylacetic acid, potato juice and gibberellin, and adding into a pot; weighing sucrose, adding into a pot, and melting; pure water is added to the volume of 1.0 liter; adjusting the pH value; packaging, and sterilizing at high temperature and high pressure; cooling to a solid; inoculating aseptic seedling for aseptic culture.
As a still further scheme of the invention: 0.7-0.8L of purified water is filled in the pot for boiling the agar powder.
As a still further scheme of the invention: the pH was adjusted using hydrochloric acid and sodium hydroxide.
As a still further scheme of the invention: the pH value is adjusted to 5.6-5.9.
As a still further scheme of the invention: the pH was adjusted to 5.7.
As a still further scheme of the invention: the split charging is divided into 20 bottles.
As a still further scheme of the invention: the high-temperature and high-pressure sterilization conditions are high temperature of 120 ℃, high pressure of 0.10MPa and sterilization for 20 minutes.
Compared with the prior art, the invention has the following advantages:
(1) the invention cultures the samara oil aseptic seedling in the rooting culture medium containing hormone, and finally obtains the samara oil seedling by utilizing the root growth promoting and growth promoting effects of the hormone, therefore, the invention has the advantages of high safety and good sustainability.
(2) According to the invention, gibberellin is added into the culture medium when the culture medium is prepared, and experiments prove that the seedlings cultured by the culture medium added with gibberellin have good lignification and high transplanting survival rate, so that the method can greatly improve the generation of healthy samara oil seedlings, reduce loss and effectively improve the rapid propagation rate of the samara oil aseptic seedlings.
(3) The method has the advantages of simple process operation, low cost and good repeatability, can greatly improve the rooting rate and the rooting number of the samara oil aseptic seedlings, and has good economic benefit and social benefit.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments.
Example 1:
the invention relates to a process for establishing a test-tube plantlet of an elaeagnus mollis: (1) taking materials; (2) sterilizing, inoculating and culturing; (3) expanding propagation; (4) and (4) induction of roots.
According to the establishment process of the test-tube plantlet, a culture medium suitable for rooting is screened.
The three hormones in the rooting medium are selected from indolebutyric acid (IBA), naphthylacetic acid (NAA) and Gibberellin (GA)3) The rooting rate of the added hormone is obviously higher than that of the non-added hormone. The screening method of the hormone addition concentration comprises the following steps: keeping other components except hormone unchanged, keeping the concentrations of two hormones unchanged each time, and setting a concentration gradient for the residual hormone. For example, a concentration gradient is set for the naphthylacetic acid, and the concentration of other hormones is kept unchanged. The concentration gradient was set for the other two hormones in the same way. At least three bottles were made in parallel per group to reduce occasional errors in the experiment. 1/2MS + IBA 0.5mg/L + NAA 0.01mg/L + GA3The rooting rate difference of 0.15mg/L is obvious and reaches 88.1%, the average number of the roots is 3.24, the roots have the highest number, the rooting and growing effects are good, the roots are thick and strong, and the base parts have no callus.
The 1/2MS formula provided by the invention is as follows: macroelements: ammonium nitrate NH4NO3825 mg/L potassium nitrate KNO3950mg/L of CaCl chloride dihydrate2·2H2O220 mg/L magnesium sulfate heptahydrate MgSO4·7H2O185 mg/L, potassium dihydrogen phosphate KH2PO4 85mg/L；
Trace elements: potassium iodide KI 0.415mg/L, boric acid H3BO33.1mg/L manganese sulfate tetrahydrate MnSO4·4H2O11.15 mg/L, zinc sulfate heptahydrate ZnSO4·7H2O4.3 mg/L, sodium molybdate dihydrate Na2MoO4·2H2O0.125 mg/L, copper sulfate pentahydrate CuSO4·5H2O0.0125 mg/L, cobalt chloride hexahydrate CoCl2·6H2O 0.0125mg/L；
Iron salt: ferrous sulfate heptahydrate FeSO4·7H2O27.8 mg/L, disodium edetate dihydrate Na2-EDTA·2H2O 37.3mg/L；
Organic matter: 100mg/L inositol, 0.5mg/L nicotinic acid VB5 or VPP, 60.5 mg/L pyridoxine hydrochloride VB, 10.1mg/L thiamine hydrochloride VB, and 2.0mg/L glycine;
30g/L of sucrose;
agar 7 g/L.
The preparation method of the potato juice comprises the following steps: taking 200g of peeled potatoes, cutting into small pieces, adding 1000 ml of water, boiling for 30 minutes, filtering out the potato pieces, and supplementing the filtrate to 1000 ml.
Sterile Elaeagnus mollis oil seedling GA3The rooting medium comprises the following components in each liter of rooting medium: 1/2MS, indolebutyric acid 0.2-1.0mg/L, naphthylacetic acid 0.005-0.03mg/L, gibberellin 0.1-0.5mg/L, potato juice 100-200g/L, sucrose 10-30g/L, agar powder 2-10g/L, and purified water to balance. The preferable components and contents of each liter of rooting medium are as follows: 1/2MS, 0.4-0.6mg/L indolebutyric acid, 0.01-0.03mg/L naphthylacetic acid, 0.1-0.3mg/L gibberellin, 180g/L potato juice, 15-25g/L sucrose, 4-6g/L agar powder and pure water to make up the balance. Further preferred components of the rooting medium per liter and the content of each component are as follows: 1/2MS, indolebutyric acid 0.5mg/L, naphthylacetic acid 0.01mg/L, gibberellin 0.15mg/L, potato juice 150g/L, sucrose20g/L, 5g/L agar powder and pure water to make up the balance.
The preparation method of the samara oil aseptic seedling rooting culture medium comprises the following steps:
weighing agar powder according to the raw material ratio, adding the agar powder into a pot filled with 0.7-0.8L of purified water, and boiling; dissolving 0.1 liter of 10 times of 1/2MS mother liquor with indolebutyric acid, naphthylacetic acid, potato juice and gibberellin, and adding into a pot; weighing sucrose, adding into a pot, and melting; pure water is added to the volume of 1.0 liter; adjusting pH to 5.6-5.9, preferably pH 5.7, with hydrochloric acid and sodium hydroxide; subpackaging into 20 bottles, sterilizing at 120 deg.C under high pressure of 0.1MPa for 20 min; cooling to a solid; inoculating aseptic seedling for aseptic culture.
Examples 2 to 15:
the rooting medium is used for culturing the samara oil aseptic seedling tissue, wherein each liter (L) of the rooting medium comprises the following components in percentage by weight, and the rooting rate and average root number are shown in the following table:
TABLE 1 design scheme of rooting medium for culturing samara oil aseptic seedling tissue, rooting rate and average root number
Agar powder is respectively weighed according to the raw material proportion in the above-mentioned embodiments 2-15 and is added into a pot filled with 0.75L of purified water to be boiled; dissolving 0.1 liter of 10 times of 1/2MS mother liquor with indolebutyric acid, naphthylacetic acid, potato juice and gibberellin, and adding into a pot; weighing sucrose, adding into a pot, and melting; pure water is added to the volume of 1.0 liter; adjusting the pH value to 5.7 by using hydrochloric acid and sodium hydroxide; subpackaging into 20 bottles, sterilizing at 120 deg.C under high pressure of 0.1MPa for 20 min; cooling to a solid; inoculating aseptic seedling for aseptic culture.
Comparative example:
rooting culture medium: 1/2MS, benzylaminopurine 5.0mg/L and indolebutyric acid 3.0mg/L are adopted in the prior art.
The culture medium prepared by the above examples 2-15 and comparative examples is used for rooting culture of the samara oil aseptic seedling, and the results in the following table show that the technical indexes of the examples are far superior to those of the comparative examples.
Table 2 comparison of the effects of the design of the examples and the comparative examples
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and the inventive concepts thereof according to the present invention should be equivalent or changed within the scope of the present invention.
Claims (10)
1. A rooting medium for sterile seedlings of samara oil GA3 is characterized in that each liter of rooting medium comprises the following components in percentage by weight: 1/2MS, indolebutyric acid 0.2-1.0mg/L, naphthylacetic acid 0.005-0.03mg/L, gibberellin 0.1-0.5mg/L, potato juice 100-200g, sucrose 10-30g/L, agar powder 2-10g/L, and purified water to make up the rest;
wherein, 1/2MS formula is: 1/2MS culture medium is MS culture medium with half of macroelements and microelements;
the preparation method of the potato juice comprises the following steps: taking 200g of peeled potatoes, cutting into small pieces, adding 1000 ml of water, boiling for 30 minutes, filtering out the potato pieces, and supplementing the filtrate to 1000 ml.
2. The rooting medium for the samara oil sterile seedling GA3 of claim 1, wherein the components and the content of each component in each liter of rooting medium are as follows: 1/2MS, indolebutyric acid 0.4-0.6mg/L, naphthylacetic acid 0.01-0.03mg/L, gibberellin 0.1-0.3mg/L, potato juice 120-180g, sucrose 15-25g/L, agar powder 4-6g/L, and purified water to balance.
3. The rooting medium for the samara oil sterile seedling GA3 of claim 2, wherein the components and the content of each component in each liter of rooting medium are as follows: 1/2MS, indolebutyric acid 0.5mg/L, naphthylacetic acid 0.01mg/L, gibberellin 0.15mg/L, potato juice 150g, sucrose 20g/L, agar powder 5g/L, and purified water in balance.
4. The preparation method of the rooting culture medium of the sterile seedling GA3 of samara oil based on any one of claims 1-3 is characterized by comprising the following steps:
weighing agar powder according to the raw material ratio, adding the agar powder into a pot filled with purified water, and boiling; dissolving 0.1 liter of 10 times of 1/2MS mother liquor with indolebutyric acid, naphthylacetic acid, potato juice and gibberellin, and adding into a pot; weighing sucrose, adding into a pot, and melting; pure water is added to the volume of 1.0 liter; adjusting the pH value; after subpackaging, sterilizing at high temperature and high pressure; cooling to a solid; inoculating aseptic seedling for aseptic culture.
5. The preparation method of the samara oil aseptic seedling GA3 rooting medium according to claim 4, wherein 0.7-0.8L of purified water is contained in a pot for boiling agar powder.
6. The method for preparing the rooting medium of the samara oil aseptic seedling GA3 of claim 4, wherein the pH is adjusted by using hydrochloric acid and sodium hydroxide.
7. The method for preparing the rooting medium of the samara oil aseptic seedling GA3 of claim 4, wherein the pH value is adjusted to 5.6-5.9.
8. The method for preparing the rooting medium of the samara oil sterile seedling GA3 as claimed in claim 4, wherein the pH value is adjusted to 5.7.
9. The preparation method of the samara oil sterile seedling GA3 rooting medium according to claim 4, wherein the split charging is performed into 20 bottles.
10. The preparation method of the samara oil sterile seedling GA3 rooting medium according to claim 4, wherein the high temperature and high pressure sterilization conditions are high temperature 120 ℃, high pressure 0.10MPa, and sterilization is performed for 20 minutes.
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| CN100346689C (en) * | 2005-10-25 | 2007-11-07 | 山西师范大学 | Breeding method for test tube seedling of elaeagnus molllis |
| CN106305428B (en) * | 2016-08-30 | 2018-11-02 | 南京林业大学 | A kind of elaeagnus mollis bud multiplication and plant regeneration method |
| CN106613974A (en) * | 2016-12-14 | 2017-05-10 | 东方上彩现代农业有限公司 | Scindapsus aureus rooting medium and scindapsus aureus rooting culture method |
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