Efficient cultivation method for honeysuckle cultivation

Abstract

The invention relates to the technical field of honeysuckle cultivation, in particular to an efficient cultivation method for honeysuckle cultivation, which comprises the following steps: step 1, explant selection: selecting a honeysuckle plant with high chlorogenic acid content; step 2, cleaning the explants; 4, differentiation culture: preparing an induction culture medium: taking an MS culture medium as a basic culture medium, and then adding cytokinin, auxin, phenylalanine and tyrosine into the basic culture medium; cutting the disinfected explants into proper lengths, inoculating the explants to a prepared induction culture medium, then putting the induction culture medium into a culture room or an illumination incubator to culture and induce dispersed cells, and differentiating to form adventitious buds or cluster buds; transferring into a rooting culture medium to grow a root system; and step 5, transplanting. By adopting a tissue culture mode, a large number of honeysuckle seedlings with high orthoacid content can be efficiently obtained.

Description

Technical Field

[0001] The present invention relates to the technical field of honeysuckle cultivation, and specifically refers to an efficient cultivation method for honeysuckle cultivation. Background Art

[0002] Honeysuckle is a commonly used Chinese medicinal material, with the effects of clearing heat and detoxifying, dispersing wind and heat, etc., and is widely used in the fields of pharmaceuticals, health products, beverages, etc.

[0003] Chlorogenic acid is the main pharmacodynamic component in honeysuckle, with a variety of biological activities. Chlorogenic acid has multiple biological activities such as antibacterial, antiviral, antioxidant, anti-inflammatory, immune enhancing, blood sugar lowering and blood lipid lowering, and is one of the main effective components for honeysuckle to exert its pharmacodynamic effects.

[0004] Traditional honeysuckle propagation methods, such as seed propagation, cutting propagation, etc., have low propagation coefficients, long seedling raising cycles, are easily infected by pests and diseases, and the content of chlorogenic acid cannot be effectively controlled.

[0005] This solution proposes an efficient cultivation method for honeysuckle cultivation, using the method of tissue culture to shorten the seedling raising cycle and increase the chlorogenic acid content. Summary of the Invention

[0006] (I) Technical Problem

[0007] The present invention aims to at least solve the problem that the chlorogenic acid content of honeysuckle propagated by traditional honeysuckle propagation methods in batches has poor consistency control.

[0008] (II) Technical Content

[0009] This solution provides an efficient cultivation method for honeysuckle cultivation, including the following steps:

[0010] Step 1, explant selection:

[0011] Select tender stems, shoot tips, buds, leaves and other parts on honeysuckle plants with high chlorogenic acid content as explants;

[0012] Step 2, explant cleaning:

[0013] Remove the surface dirt and impurities from the taken explants, and then rinse them clean with tap water;

[0014] Step 3, explant disinfection treatment:

[0015] Perform pre-disinfection on the cleaned explants, and then rinse them with sterile water; then soak them in a mercuric chloride solution;

[0016] Step 4, differentiation culture:

[0017] Induction medium preparation: Use MS medium as the basal medium, and then add cytokinin, auxin, phenylalanine, and tyrosine to the basal medium;

[0018] On the ultra-clean workbench, cut the disinfected explants into appropriate lengths and inoculate them onto the prepared induction medium; place the inoculated culture containers in the culture room or light incubator to culture and induce dispersed cells to differentiate into adventitious buds or cluster buds;

[0019] Transfer the obtained adventitious buds or cluster buds to the rooting medium and culture them under suitable temperature and light conditions to produce roots;

[0020] Step Five, transplantation:

[0021] When the test-tube seedlings grow to a certain height, take the test-tube seedlings out of the culture container and transplant them into a nutrient bowl or seedling tray filled with sterile substrate for hardening-off;

[0022] After hardening-off, transplantation can be carried out.

[0023] Preferred Technical Solution One: The induction medium uses MS medium as the basal medium, and then add cytokinin to the basal medium until the concentration reaches 1.0 - 2.0 mg / L;

[0024] Add auxin to the basal medium until the concentration reaches 0.1 - 0.5 mg / L;

[0025] Add phenylalanine to the basal medium until the concentration reaches 50 - 200 mg / L;

[0026] Add tyrosine to the basal medium until the concentration reaches 20 - 100 mg / L.

[0027] Preferred Technical Solution Two: In Step One, before selecting the explants, first perform mutation induction on the explant plants. The process is as follows:

[0028] Use chemical mutagens to treat Lonicera japonica Thunb. to induce mutants, and then screen out mutant strains with significantly increased chlorogenic acid content from them.

[0029] Preferred Technical Solution Three: The sterile substrate is one or a mixture of several of perlite, vermiculite, peat soil, etc.

[0030] Preferred Technical Solution Four: The chemical mutagen is one or a mixture of several of ethyl methanesulfonate, sodium azide, colchicine, etc.

[0031] Preferred Technical Solution Five: Sucrose is also added to the basal medium, and the sucrose concentration is added to 20 - 30 g / L.

[0032] Preferred Technical Solution Six: Agar is also added to the basal medium, and the agar concentration is adjusted to 7-9 g / L.

[0033] (III) Technical Effects

[0034] Adopting the above structure enables this solution to have the following beneficial effects:

[0035] 1. By adopting the tissue culture method, a large number of honeysuckle seedlings with high content of original acid can be efficiently obtained;

[0036] 2. Shorten the seedling raising cycle and reduce the production cost;

[0037] 3. The content of original acid can be effectively controlled. Specific Embodiments

[0038] Next, the technical solutions in the embodiments of the present invention will be described clearly and completely. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments; based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0039] Embodiment

[0040] An efficient cultivation method for honeysuckle includes the following steps:

[0041] Step 1, explant selection:

[0042] Before the explant is selected, the explant plant is first subjected to mutation induction, and the process is as follows:

[0043] The honeysuckle is treated with a chemical mutagen to induce mutants, and then mutants with significantly increased chlorogenic acid content are screened out;

[0044] Among them, the chemical mutagen is one or a mixture of ethyl methanesulfonate, sodium azide, and colchicine;

[0045] Select healthy, pest-free, and high-chlorogenic acid-content honeysuckle plants from the above mutants;

[0046] Select parts such as tender stems, shoot tips, buds, and leaves on the plants as explants. For example, select shoot tips or stem segments with axillary buds on the semi-lignified branches of the current year;

[0047] Step 2, explant cleaning:

[0048] Put the taken explants into a dishwashing liquid solution and soak for a few minutes to remove surface dirt and impurities, and then rinse them thoroughly with tap water;

[0049] Step 3, explant disinfection treatment:

[0050] Put the cleaned explants into 75% alcohol and soak for 5 - 30 seconds for pre - disinfection, then rinse with sterile water 2 - 3 times;

[0051] Then soak in 0.1% - 0.2% mercuric chloride solution for 2 - 10 minutes. The specific time depends on the size and texture of the explants. Gently shake during soaking to make the disinfection more thorough, and finally rinse with sterile water multiple times to remove the residual disinfectant;

[0052] Step four, differentiation culture:

[0053] Configuration of induction medium: Based on MS medium as the basal medium, then add cytokinin, auxin, phenylalanine, and tyrosine to the basal medium. Among them:

[0054] Add cytokinin to the basal medium to a concentration of 1.0 - 2.0 mg / L;

[0055] Add auxin to the basal medium to a concentration of 0.1 - 0.5 mg / L;

[0056] Add phenylalanine to the basal medium to a concentration of 50 - 200 mg / L;

[0057] Add tyrosine to the basal medium to a concentration of 20 - 100 mg / L;

[0058] Add sucrose to the basal medium to a concentration of 20 - 30 g / L;

[0059] Add agar to the basal medium to a concentration of 7 - 9 g / L;

[0060] On the ultra - clean workbench, cut the disinfected explants into appropriate lengths and inoculate them onto the prepared induction medium;

[0061] Put the inoculated culture containers into the culture room or light incubator to culture and induce dispersed cells to differentiate into adventitious buds or cluster buds. The temperature in the incubator is generally controlled at about 25°C, provide 14 - 16 hours of light per day, and the light intensity is 2000 - 3000 lx;

[0062] Transfer the obtained adventitious buds or cluster buds to the rooting medium and culture them under suitable temperature (about 23 - 27°C) and light conditions to produce roots;

[0063] Step five, transplantation:

[0064] When the test-tube plantlets grow to a certain height and have well-developed roots, take the test-tube plantlets out of the culture container, wash off the culture medium attached to the roots, and then transplant them into a nutrient bowl or seedling tray filled with sterile substrate. Water thoroughly, cover with a plastic film to keep moisture, and carry out acclimatization in a greenhouse or under natural light. Shade appropriately for the first few days, and then gradually increase the light intensity and duration to allow the test-tube plantlets to gradually adapt to the external environment.

[0065] Among them, the sterile substrate is one or a mixture of several of perlite, vermiculite, peat soil, etc.

[0066] After a period of acclimatization, when the test-tube plantlets grow strong and have well-developed roots, they can be transplanted into the field or flowerpots. When transplanting, pay attention to selecting suitable soil and planting time to ensure the normal growth of the transplanted honeysuckle.

[0067] It should be noted that in this text, the term "including", "comprising" or any other variant thereof is intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements not only includes those elements, but also includes other elements not expressly listed, or further includes elements inherent to such process, method, article or device.

[0068] Although the embodiments of the present invention have been shown and described, for those of ordinary skill in the art, it can be understood that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principle and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. An efficient cultivation method for cultivating honeysuckle, characterized in that: The steps include: Step 1: Explant selection: Choose honeysuckle plants that are healthy, pest-free, and high in chlorogenic acid; Select tender stems, stem tips, buds, leaves and other parts of the plant as explants; Step 2: Explant cleaning: Remove dirt and impurities from the surface of the explant; Step 3: Disinfection of explants: The cleaned explants were pre-sterilized and then rinsed with sterile water to remove residual disinfectant; Step 4: Differentiation culture: Induction medium configuration: MS medium is used as the basic medium, and then cytokinin, auxin, phenylalanine and tyrosine are added to the basic medium; On a clean bench, cut the sterilized explants into appropriate lengths and inoculate them onto the prepared induction medium; The inoculated culture container is placed in a culture room or a light incubator to culture and induce dispersed cells to differentiate into adventitious buds or clustered buds; The obtained adventitious buds or clustered buds are transferred to a rooting medium to cultivate and grow roots; Step 5: Transplanting: When the test-tube seedlings grow to a certain height, they are taken out of the culture container and transplanted into a nutrient pot or seedling tray filled with a sterile substrate for seedling hardening. After hardening the seedlings, they can be transplanted.

2. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: In step 4, cytokinin is added to the basal medium to a concentration of 1.0-2.0 mg / L; Adding auxin to the basal medium to a concentration of 0.1-0.5 mg / L; Adding phenylalanine to the basal medium to a concentration of 50-200 mg / L; Tyrosine is added to the basal medium to a concentration of 20-100 mg / L.

3. The efficient cultivation method for honeysuckle cultivation according to claim 2, characterized in that: Sucrose was also added to the basal medium, and the concentration of sucrose was added to 20 to 30 g / L.

4. The efficient cultivation method for honeysuckle cultivation according to claim 3, characterized in that: Agar is also added to the basal medium, and the agar concentration is added to 7 to 9 g / L.

5. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: In step 1, before the explants are selected, mutation induction is first performed on the explants, and the process is as follows: The honeysuckle was treated with chemical mutagens to induce mutants, and then the mutants with significantly increased chlorogenic acid content were screened out.

6. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: In step 3, the cleaned explants are pre-sterilized by soaking them in 75% alcohol for 5-30 seconds, and then rinsed with sterile water 2-3 times; Then soak in 0.1%-0.2% mercuric chloride solution for 2-10 minutes.

7. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: In step 4, the temperature in the incubator is generally controlled at about 25°C, and 14-16 hours of light are provided every day, with a light intensity of 2000-3000lx.

8. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: The chemical mutagen is one of ethyl methanesulfonate, sodium azide and colchicine or a mixture of several of them.

9. The efficient cultivation method for honeysuckle cultivation according to claim 5, characterized in that: In step 5, the sterile matrix is ​​one or a mixture of perlite, vermiculite, and peat soil.

10. The efficient cultivation method for honeysuckle cultivation according to claim 1, characterized in that: In step 4, the culture temperature of the rooting medium is 23-27°C.