Tissue culture method for bulbils of brachygyne tenuifolia
Through the tissue culture method of the fern bead buds of light leaf fern, mature bead buds are used as explants to disinfect and prepare sterile culture medium and light incubator culture, the problem of long cultivation cycle and seasonal limitations of bead bud soil is solved, and a rapid expansion and simplified culture process is achieved.
Patent Information
- Application Number
- CN202510507233.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-04-22
- Publication Date
- 2025-06-06
AI Technical Summary
The bead bud soil cultivation cycle of the light-leaf fern has a long period of time and is slow to grow, and is restricted by seasons, making it difficult to achieve rapid expansion and proliferation.
The mature bead buds of the light leaf fern are used as explants, and the tissue culture of the bead buds is achieved by disinfection, preparation of sterile solid culture medium and light incubator culture.
The breeding cycle of the bead buds of the light leaf fern has been shortened, and the rapid expansion and reproduction has been achieved without seasonal restrictions. The disinfection time is short, the cultivation process is simplified, and the cost is reduced.
Smart Images
Figure CN120092710A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the technical field of fern propagation and cultivation, in particular to a tissue culture method for bulbils of Pteris glabra. Background Art
[0002] As a national first-class protected wild plant, since its discovery, researchers have carried out a series of basic research and conservation and rescue plans, such as habitat protection, growth monitoring and plant breeding measures, but no substantial progress has been made, especially in plant breeding.
[0003] At present, the only method for propagating bulbils of Pteris glabra is soil culture, but this method has a long cycle, slow growth, and is restricted by seasons, which is not conducive to propagation. Summary of the invention
[0004] The invention aims to provide a tissue culture method for the bulbils of Pteris glabra, so as to solve the shortcomings of the bulbils of Pteris glabra, such as long soil culture period, slow growth and seasonal restrictions.
[0005] To achieve the above object, the present invention provides the following technical solutions:
[0006] A tissue culture method for Pteris glabra bulbils, the tissue culture method for Pteris glabra bulbils specifically comprising: S1: selecting an explant
[0007] Selecting mature bulbils of Pteris glabra as explants, collecting the bulbils of Pteris glabra and placing them in a container filled with water;
[0008] S2: Explant sterilization
[0009] The light-leaved fern bulbils are rinsed under running water; the rinsed light-leaved fern bulbils are placed in a tea bag sterilized at high temperature, disinfected with a mixed solution, and then rinsed with sterile water; the sterilized light-leaved fern bulbils are placed in a high temperature sterilized plate for later use;
[0010] S3: Preparation of sterile solid culture medium
[0011] Adding culture medium and agar to the culture bottle to obtain a culture medium solution, adjusting the pH value of the culture medium solution; then sterilizing at high temperature, cooling to room temperature, and obtaining the sterile solid culture medium;
[0012] S4: Inoculation and cultivation
[0013] Inoculating the bulbils of Pteris glabra into the sterile solid culture medium, and culturing in a light incubator to obtain Pteris glabra seedlings;
[0014] S5: Transfer seedlings
[0015] When the seedlings are observed to have fully expanded their leaves, they can be transferred.
[0016] Preferably, in said S1, said bulbils of Pteris glabra are bulbils of Pteris glabra that mature in July every year.
[0017] Preferably, in S2, the bulbils of Pteris glabra are rinsed under tap water for 3 hours.
[0018] Preferably, in S2, the mixed solution is a mixture of 100 mL of 0.1% HgCl2 and Tween 80, the disinfection time is 30 seconds, and the sterile water is rinsed 3 to 5 times.
[0019] Preferably, in S3, the culture medium is 1 / 2MS culture medium, does not contain agar and sucrose, and has a concentration of 2.47 g / L.
[0020] Preferably, in S3, the concentration of the agar in the sterile solid culture medium is 4.8 g / L.
[0021] Preferably, in S3, the pH value of the culture medium solution is adjusted to 5.8-5.9.
[0022] Preferably, in S3, the high temperature sterilization is to sterilize the prepared culture medium solution at 121° C. for 15 to 20 minutes.
[0023] Preferably, in S4, the culture conditions of the bulbils of the Pteris glabra are a light cycle of 12h light / 12h dark, a light intensity of 40%, and a temperature of 18±1°C.
[0024] Preferably, in S4, during the culture process, the culture medium needs to be replaced every 45 to 60 days to allow the plants to continuously absorb nutrients.
[0025] Compared with the prior art, the present invention has the following beneficial effects:
[0026] 1. The present invention is the first to use bulbils as explants to carry out tissue culture of Pteris glabra.
[0027] 2. Not restricted by seasons: Since the light-leaved fern is a winter-dead plant, traditional seeding and seedling cultivation and bulbil soil culture will be affected by various factors such as season and climate, resulting in a long breeding cycle. However, bulbil tissue culture will not be affected, shortening the culture cycle and achieving rapid expansion.
[0028] 3. Short disinfection time: Compared with the existing tissue culture technology, the present invention has an obvious time advantage in the explant disinfection link, and the mixed solution only needs 30 seconds for disinfection.
[0029] 4. One-step seedling formation from tissue culture seedlings: The present invention does not have the time-consuming, costly and cumbersome multi-stage transfer process in existing plant tissue culture. Specifically for the method of one-step seedling formation from tissue culture seedlings of Pteris glabra, a simplified culture process is adopted, which effectively shortens the culture cycle, reduces costs, and enhances the consistency and health of plant growth. BRIEF DESCRIPTION OF THE DRAWINGS
[0030] Figure 1 The mature bulbils of Pteris glabra of the present invention;
[0031] Figure 2 for germinating the bulbils of the present invention;
[0032] Figure 3 The bulbils of the present invention are sprouted;
[0033] Figure 4 The bulbils of the present invention begin to unfold leaves;
[0034] Figure 5 The light-leaved fern seedlings grown from the bulbils of the present invention;
[0035] Figure 6 The light-leaved fern seedling of the present invention;
[0036] Figure 7 A large number of Pteris glabra seedlings cultured in the laboratory of the present invention. DETAILED DESCRIPTION
[0037] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. The experimental methods described in the following embodiments are conventional methods unless otherwise specified; the reagents and materials, unless otherwise specified, can be obtained from commercial channels.
[0038] Example
[0039] 1. Selection of explants: Select the bulbils of Pteris glabra that mature in July every year (the top forms a protrusion and the part in contact with the leaves turns brown) as explants. After collection, they should be placed in a bottle filled with water or a plastic bag and brought back.
[0040] 2. Treatment of explants: Place the brought back pearl buds into a glass bottle 8-12 cm high, tie a filter to the mouth of the bottle (the filter mouth should be smaller than the size of the pearl buds to prevent the pearl buds from being washed away), rinse under tap water for 3 hours, and then place in a high-temperature sterilized tea bag.
[0041] 3. Explant disinfection: In a clean bench, use 100mL 0.1% HgCl2 Sterilize the bulbils with a mixed solution of 1% tween 80 and Tween 80 for 30 seconds, then rinse with sterile water for 3 to 5 times. Place the washed explants in a high-temperature sterilized dish for later use.
[0042] 4. Prepare sterile solid culture medium: add 1 / 2MS culture medium (without agar and sucrose) to the culture bottle, add agar to prepare the culture medium solution. The concentration of 1 / 2MS is 2.47g / L, the concentration of agar is 4.8g / L, and the pH value of the culture medium solution is adjusted to 5.8-5.9; then sterilize at 121℃ for 15-20min, take out and cool to room temperature to obtain sterile solid culture medium for use.
[0043] 5. Inoculation and culture: Inoculate the bulbils into sterile solid culture medium, with one bulbil inoculated per bottle. After inoculation, place the bulbils in a BSG-800 light incubator for culture, set a certain temperature, humidity and light, the light cycle is 12h light / 12h dark, the light intensity is 40%, and the temperature is controlled at 18±1°C to obtain the light leaf fern seedlings.
[0044] 6 Transfer seedlings: When the seedlings are observed to have fully expanded their leaves, they can be transferred to accelerate their growth and rooting.
[0045] Generally speaking, germination begins 7 days after inoculation, sprouting begins 15 days after inoculation, and leafing begins 20 days after inoculation. Note that during the culture process, the culture medium needs to be replaced every 45 to 60 days to ensure continuous nutrient absorption by the plant.
[0046] Finally, it should be noted that the above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art can still modify the technical solutions described in the aforementioned embodiments or replace some of the technical features therein by equivalents. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.
Claims
1. A method for tissue culture of Pteris glabra bulbils, characterized in that: The tissue culture method of the bulbils of Pteris glabra is specifically as follows: S1: Selection of explants Selecting mature bulbils of Pteris glabra as explants, collecting the bulbils of Pteris glabra and placing them in a container filled with water; S2: Explant sterilization The light-leaved fern bulbils are rinsed under running water; the rinsed light-leaved fern bulbils are placed in a tea bag sterilized at high temperature, disinfected with a mixed solution, and then rinsed with sterile water; the sterilized light-leaved fern bulbils are placed in a high temperature sterilized plate for later use; S3: Preparation of sterile solid culture medium Adding culture medium and agar to the culture bottle to obtain a culture medium solution, adjusting the pH value of the culture medium solution; then sterilizing at high temperature, cooling to room temperature, and obtaining the sterile solid culture medium; S4: Inoculation and cultivation Inoculating the bulbils of Pteris glabra into the sterile solid culture medium, placing the culture medium in a light incubator for cultivation, and obtaining Pteris glabra seedlings; S5: Transfer seedlings When the seedlings are observed to have fully expanded their leaves, they can be transferred.
2. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In the S1, the bulbils of Pteris glabra are bulbils of Pteris glabra that mature in July every year.
3. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In the S2, the bulbils of the Pteris glabra are rinsed under tap water for 3 hours.
4. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In S2, the mixed solution is a mixture of 100 mL of 0.1% HgCl2 and Tween 80, the disinfection time is 30 seconds, and the sterile water is rinsed 3 to 5 times.
5. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In the S3, the culture medium is 1 / 2MS culture medium without agar and sucrose, and the concentration is 2.47 g / L.
6. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In S3, the concentration of the agar in the sterile solid culture medium is 4.8 g / L.
7. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In S3, the pH value of the culture medium solution is adjusted to 5.8-5.
9.
8. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In S3, the high temperature sterilization is to sterilize the prepared culture medium solution at 121° C. for 15 to 20 minutes.
9. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In the S4, the culture conditions of the bulbils of the Pteris glabra are a light cycle of 12h light / 12h dark, a light intensity of 40%, and a temperature of 18±1°C.
10. The tissue culture method of Pteris glabra bulbils according to claim 1, characterized in that: In S4, during the culture process, the culture medium needs to be replaced every 45 to 60 days to allow the plants to continuously absorb nutrients.