Culture medium for tissue culture of ampelopsis japonica

Abstract

The invention belongs to the technical field of plant biology, and discloses a culture medium for tissue culture of ampelopsis japonica, the culture medium is a one-step seedling culture medium, and the culture medium is composed of inorganic salt, an organic supplement, a growth regulator and a plant substrate; the method is specially used for efficient tissue culture of the ampelopsis japonica, the tissue culture efficiency of the ampelopsis japonica is improved, meanwhile, the operation process of plant culture is simplified, the growth time in the seedling stage is remarkably shortened, the ampelopsis japonica tissue is effectively prevented from being infected by microorganisms, and necrosis of the ampelopsis japonica tissue is effectively prevented.

Description

Technical Field

[0001] The invention relates to the field of plant biotechnology, in particular to a culture medium for cultivating Bletilla striata tissue. Background Art

[0002] Bletilla striata is a traditional Chinese medicinal material widely used in the field of traditional Chinese medicine. It has a wide range of medicinal value and nutrients and is often used to treat a variety of diseases. With the development of modern science and technology, research on Bletilla striata and its active ingredients has gradually increased, but due to its slow natural growth rate and difficulty in collection, its market supply is insufficient. Therefore, it is particularly important to develop efficient breeding methods.

[0003] Tissue culture is a technology that uses plant cells or tissues to proliferate and differentiate in a sterile environment. This technology can not only achieve efficient reproduction, but also maintain the genetic characteristics and active ingredients of the plant. In plant tissue culture, the composition and formulation of the culture medium have an important influence on the growth, differentiation and synthesis of active ingredients of the tissue. The appropriate culture medium can provide the nutrients and growth regulators required for plant growth, thereby significantly improving the success rate of tissue culture.

[0004] At present, the tissue culture of Bletilla striata is still in the research stage, and there is a lack of special culture medium formulas tailored to its characteristics. The research on special culture medium for Bletilla striata is still insufficient, and currently it usually relies on the nutrient formula of other plants, failing to fully consider the special needs of Bletilla striata in cell growth and differentiation.

[0005] Therefore, there is an urgent need to develop an efficient tissue culture medium formula specifically for Bletilla striata, aiming to improve the tissue culture efficiency of Bletilla striata. Summary of the invention

[0006] The invention aims to provide a culture medium for tissue culture of Bletilla striata, so as to carry out efficient tissue culture specifically for Bletilla striata and improve the tissue culture efficiency of Bletilla striata.

[0007] In order to achieve the above object, the present invention provides the following technical solutions:

[0008] A culture medium for tissue culture of Ampelopsis radiata, the culture medium being a one-step seedling culture medium, and the culture medium consisting of the following components:

[0009] Including inorganic salts: potassium nitrate 850mg / L, ammonium nitrate 160mg / L, magnesium nitrate 180mg / L, potassium dihydrogen phosphate 280mg / L, calcium nitrate 150mg / L, calcium chloride 190mg / L, manganese sulfate dihydrate 10mg / L, zinc sulfate 1mg / L, boric acid 6mg / L, potassium iodide 1mg / L, calcium chloride hexahydrate 0.1mg / L, copper sulfate pentahydrate 0.3mg / L, sodium sulfate dihydrate 0.1mg / L;

[0010] Organic supplements: Na2-EDFA 16mg / L, ferrous sulfate heptahydrate 20mg / L, inositol 100mg / L, B16 mg / L, B64 mg / L, hydrochloric acid 5mg / L, amino acids 10mg / L, ascorbic acid 5mg / L, glutamic acid 10mg / L, ribosyl acid 3mg / L, folic acid 5mg / L;

[0011] Growth regulators: 6-BA 1.6mg / L, 2,4-D 0.8mg / L, IBA 0.3mg / L, rooting 2.5mg / L;

[0012] Plant matrix: potato extract 100g / L, activated carbon powder 1g / L, carrageenan 8g / L, white sugar 20g / L.

[0013] In the embodiment of the present invention, within the scope known to those skilled in the art, Na2-EDFA is sodium ethylenediamine di-o-phenyl acetate; B1 is vitamin B1, i.e. thiamine hydrochloride; B6 is vitamin B6, i.e. pyridoxine; 6-BA is 6-benzylaminopurine; 2,4-D is 2,4-dichlorophenoxyacetic acid; and IBA is indolebutyric acid.

[0014] The beneficial effects of the basic technical solution are:

[0015] 1. The present invention is a one-step seedling culture medium, which provides all the elements required in a single step, from sterile culture to the formation of seedlings, eliminating the cumbersome multiple steps in the traditional culture system, simplifying the operation process, significantly shortening the seedling growth time, improving the seedling rate, and reducing resource consumption.

[0016] 2. Compared with the segmented operation of traditional multi-step culture medium, the one-step seedling culture medium avoids microbial infection during the segmented operation and effectively prevents plant tissue necrosis. DETAILED DESCRIPTION

[0017] The present invention is further described in detail below in conjunction with the embodiments:

[0018] Example 1 Preparation of culture medium

[0019] Prepare the culture medium using the following recipe:

[0020] Inorganic salts: potassium nitrate 850mg / L, ammonium nitrate 160mg / L, magnesium nitrate 180mg / L, potassium dihydrogen phosphate 280mg / L, calcium nitrate 150mg / L, calcium chloride 190mg / L, manganese sulfate dihydrate 10mg / L, zinc sulfate 1mg / L, boric acid 6mg / L, potassium iodide 1mg / L, calcium chloride hexahydrate 0.1mg / L, copper sulfate pentahydrate 0.3mg / L, sodium sulfate dihydrate 0.1mg / L;

[0021] Organic supplements: Na2-EDFA 16mg / L, ferrous sulfate heptahydrate 20mg / L, inositol 100mg / L, B16mg / L, B64mg / L, hydrochloric acid 5mg / L, amino acids 10mg / L, ascorbic acid 5mg / L, glutamic acid 10mg / L, ribosyl acid 3mg / L, folic acid 5mg / L;

[0022] Growth regulators: 6-BA 1.6 mg / L, 2,4-D 0.8 mg / L, IBA 0.3 mg / L, rooting 2.5 mg / L;

[0023] Plant matrix: potato extract 100g / L, activated carbon powder 1g / L, carrageenan 8g / L, white sugar 20g / L.

[0024] Example 1: Experiment on clone propagation of Ampelopsis pilosula tissue

[0025] S1. Prepare the Bletilla striata tissue: select a well-growing and virus-free Bletilla striata mother plant, perform disinfection and tissue cutting treatments in sequence, and finally obtain the Bletilla striata tissue;

[0026] S2. Inoculating the Bletilla striata tissue: placing the Bletilla striata tissue into the culture medium in a culture bottle under a sterile environment;

[0027] S3. Cultivating Bletilla striata tissue: placing the inoculated culture medium into the culture rack of the culture room, and adjusting the environment of the culture room to: temperature 25°C, humidity below 45, 16 hours of light, and 8 hours of darkness.

[0028] S4. Record the cultivation results: After 3 months of cultivation, count the number of inocula that can be inoculated and calculate the reproduction coefficient.

[0029] Comparative Example 1: Effects of different concentrations of 6-BA on the meristem reproduction of Bletilla striata

[0030] The specific implementation is the same as that of Example 1, except that when configuring the culture medium,

[0031] In Example 1, the concentration of 6-BA was 1.6 mg / L;

[0032] In Comparative Example 1, the concentration of 6-BA was 1.3 mg / L and 2.0 mg / L;

[0033] The data of meristem propagation of Bletilla striata tissues are shown in Table 1 ;

[0034] Table 1 Effects of different concentrations of 6-BA on tissue meristem reproduction of Bletilla striata

[0035]

[0036] As shown in Table 1, the tissues of Bletilla striata can grow in the 6-BA concentration of 1.3-2.0 mg / L. In the culture medium with 6-BA concentration of 1.6 mg / L, the meristem reproduction effect of the tissues of Bletilla striata is the best.

[0037] Example 2 Preparation of culture medium

[0038] The specific implementation is the same as that of Example 1;

[0039] Example 2 Rooting experiment of Ampelopsis pilosula tissue

[0040] The specific implementation is the same as that of Example 1, except that in step S4;

[0041] Step S4 of Example 1, after 3 months of cultivation, counting the inocula and calculating the reproduction coefficient;

[0042] Step S4 of Example 2, after 3 months of cultivation, counting the number of roots;

[0043] Comparative Example 2: Effects of different concentrations of IBA on the number of rooting tissues of Ampelopsis pilosula

[0044] The specific implementation is the same as that of Example 2, except that when configuring the culture medium,

[0045] In Example 2, the concentration of IBA was 0.3 mg / L;

[0046] In Comparative Example 1, the concentrations of IBA were 0.25 mg / L, 0.3 mg / L, and 0.35 mg / L;

[0047] The data on the number of rooting tissues of Bletilla striata are shown in Table 2;

[0048] Table 2 Effects of different concentrations of IBA on the number of roots in Bletilla striata tissues

[0049]

[0050] As shown in Table 2, the Bletilla striata tissue can take root in the medium with IBA concentration of 0.25-0.35 mg / L, and the rooting effect of the Bletilla striata tissue is the best in the medium with IBA concentration of 0.30 mg / L.

[0051] The above is only an embodiment of the present invention, and the common knowledge such as the known specific technical solutions or characteristics in the solution is not described in detail here. It should be pointed out that for those skilled in the art, several modifications and improvements can be made without departing from the technical solution of the present invention, which should also be regarded as the protection scope of the present invention, and these will not affect the effect of the implementation of the present invention and the practicality of the patent. The scope of protection required by this application shall be based on the content of its claims, and the specific implementation methods and other records in the specification can be used to interpret the content of the claims.

Claims

1. A culture medium for tissue culture of Ampelopsis pilosula, characterized in that: The culture medium is a one-step seedling culture medium, and the culture medium is composed of the following components: Including inorganic salts: potassium nitrate 850mg / L, ammonium nitrate 160mg / L, magnesium nitrate 180mg / L, potassium dihydrogen phosphate 280mg / L, calcium nitrate 150mg / L, calcium chloride 190mg / L, manganese sulfate dihydrate 10mg / L, zinc sulfate 1mg / L, boric acid 6mg / L, potassium iodide 1mg / L, calcium chloride hexahydrate 0.1mg / L, copper sulfate pentahydrate 0.3mg / L, sodium sulfate dihydrate 0.1mg / L; Organic supplements: Na2-EDFA 16mg / L, ferrous sulfate heptahydrate 20mg / L, inositol 100mg / L, B16 mg / L, B64 mg / L, hydrochloric acid 5mg / L, amino acids 10mg / L, ascorbic acid 5mg / L, glutamic acid 10mg / L, ribosyl acid 3mg / L, folic acid 5mg / L; Growth regulators: 6-BA 1.6mg / L, 2,4-D 0.8mg / L, IBA 0.3mg / L, rooting 2.5mg / L; Plant matrix: potato extract 100g / L, activated carbon powder 1g / L, carrageenan 8g / L, white sugar 20g / L.

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