Method for accelerating reproduction of chrysanthemum by using liquid culture medium

Abstract

The invention relates to the technical field of chrysanthemum breeding methods, in particular to a method for accelerating chrysanthemum breeding by using a liquid culture medium, which comprises the following steps: cutting chrysanthemum stalks into square blocks of 1cm * 1cm in a culture box, flatly laying the stalks upwards on an induction culture solution soaking base, cutting stem tissues with axillary buds into small sections of 2-3cm, and obliquely inserting the small sections into the induction culture solution for culturing, mS + 6-BA 2.0 mg / L +, NAA 0.1 mg / L, alpha-naphthylacetic acid 2-4 mg / L, 6-benzylaminopurine and cane sugar are selected as culture solutions, in the propagation culture of the chrysanthemum, stem seedling sections grow fast, each section generates a plurality of clustered seedlings, the average multiplication coefficient is 4-7 times, the height of the stem seedlings is different, the clustered seedlings are 5-7 cm high when the seedlings are cultured in the nutrient solution for 2 months, and the adventitious bud multiplication rate of the chrysanthemum seedlings is increased. The method has the effect of accelerating chrysanthemum reproduction.

Description

Technical Field

[0001] The present application relates to the technical field of chrysanthemum propagation methods, and specifically to a method for accelerating chrysanthemum propagation by utilizing a liquid culture medium. Background Art

[0002] Chrysanthemum is a perennial herbaceous plant of the genus Chrysanthemum in the family Asteraceae. It is native to China and is regarded as the national flower of Japan. It evolved from some wild chrysanthemums through interspecific hybridization; the stems are branched and woody at the base; the leaves are single and alternate, mostly oval, with coarse serrations or deep fissures on the edges; the inflorescence is capitulate, with ligulate flowers on the periphery, which vary greatly in size and shape, including flat petals, spoon-shaped petals, etc.; the center is a tubular flower, which is often rare or absent; different petal shapes form different flower shapes, colors, and varieties. Below the inflorescence is the involucre, the ligulate flowers are mostly male flowers, the tubular flowers are bisexual flowers, and the pistil stigma is bifurcated.

[0003] At present, a method for cultivating strong gerbera seedlings is disclosed, which is also a commonly used technology, and includes the following steps: Ⅰ. Inoculate gerbera tissue culture seedlings into a rooting medium for cultivation. Ⅱ. Select the gerbera tissue culture seedlings cultured in step Ⅰ and transplant them into a cultivation medium. Ⅲ. Regularly water the gerbera tissue culture seedlings transplanted into the cultivation medium with a bacterial suspension. The preparation process of the bacterial suspension is to take the Indian pear-shaped spore solution cultured in PDB liquid culture medium, wash it with ddH2O after centrifugation, add ddH2O and crush it, and adjust the absorbance value. Ⅳ. Perform a seedling acclimatization treatment on the gerbera seedlings. Ⅴ. After the gerbera seedlings after the seedling acclimatization treatment are fully watered, natural drought treatment is adopted, and the drought treatment is carried out until the gerbera seedlings fully unfold the third leaf.

[0004] Another disclosed method of inducing chrysanthemum seedlings in one step with high efficiency using low-concentration chlorine dioxide is also an increasingly mature technology, comprising the following steps: adding low-concentration chlorine dioxide to the chrysanthemum culture medium to induce the simultaneous growth of chrysanthemum stems, leaves and roots, and completing chrysanthemum tissue culture without staged culture; the chlorine dioxide concentration is 1-20 μg / L, the culture medium contains or does not contain exogenous hormones, and chrysanthemum plants with strong growth and well-developed root systems can be obtained after 20 days of culture, and the transplant survival rate reaches 100%. By promoting plant growth and rooting with low-concentration chlorine dioxide, while efficiently inducing efficient regeneration of chrysanthemums, it gets rid of the current dependence on exogenous hormones in tissue culture.

[0005] With respect to the above-mentioned related technologies, the inventors believe that the following defects exist: the breeding and cultivation method of gerbera is relatively complicated, which affects the cultivation of chrysanthemums during the breeding process; the cultivation of this kind of spore liquid may cause root rot of chrysanthemums, directly affecting the growth of chrysanthemums. Summary of the invention

[0006] In order to improve the technical problem that the cultivation of spore liquid may cause root rot of chrysanthemum and directly affect the growth of chrysanthemum, the present application provides a method for accelerating the reproduction of chrysanthemum using liquid culture medium.

[0007] The present application provides a method for accelerating chrysanthemum reproduction using a liquid culture medium, which adopts the following technical solution:

[0008] A method for accelerating chrysanthemum reproduction using a liquid culture medium comprises the following steps:

[0009] Step 1: Cut the chrysanthemum stems into 1 cm × 1 cm squares in a culture box, lay the stems upward on the base soaked in induction culture solution, cut the stem tissue with axillary buds into small sections of 2 to 3 cm, and obliquely insert them into the induction culture solution for culture. The culture solution is selected from MS + 6-BA 2.0 mg / L +, NAA 0.1 mg / L, α-naphthyl acetic acid, 2-4 mg / L, 6-benzylaminopurine, sucrose;

[0010] Step 2: After the chrysanthemum stem tubers and stem tissues with axillary buds are induced to sprout, the tender shoots are cut out for further cultivation, and new chrysanthemum seedlings are continued to be propagated. MS is selected as the culture medium, and the hormones MS+6-BA2.0mg / L+, NAA0.1mg / L and 6-benzylaminopurine, potassium dihydrogen phosphate are added. The root growth culture solution is selected from MS, and NAA0.3mg / L base solution is added. Agar 4.5g / L and sucrose 20g / L are added to the culture box;

[0011] Step 3: The temperature for chrysanthemum reproduction and cultivation is 10-15°C, the light intensity is 1000-1500lx, and the lighting time is 6h / d.

[0012] Step 4. Transport the chrysanthemum seedlings with rhizomes of 1 to 2 cm to a room with a room temperature of 10° to 24°. Control the temperature at about 23°C, expose them to sunlight for 5 days, receive sufficient photosynthesis, spray nutrient solution manually, cover with plastic transparent film to keep warm and prevent water loss. When cultivating and growing, wash the chrysanthemum seedlings with warm water, wash and remove the culture solution on the rhizomes, transplant the chrysanthemum seedlings into the seedling tray sterilized with potassium permanganate, spray water manually, build an arch shed with plastic film on the seedling tray to keep warm and moisturize, gradually increase the photosynthetic exposure after 3 days of cultivation, sterilize and disinfect and fertilize once every 3 days.

[0013] By adopting the above technical scheme, the liquid culture medium can be used efficiently and quickly to accelerate the reproduction process of chrysanthemum, significantly improve the reproduction efficiency, and ensure the healthy development of chrysanthemum seedlings at different growth stages. First, by cutting the chrysanthemum stems and inducing culture in the liquid culture medium, the stem tissue with axillary buds is promoted to differentiate and sprout, and the chrysanthemum seedlings are quickly propagated; secondly, in the process of induction culture and propagation culture, the plant hormones and nutrients in the culture solution are precisely regulated, such as MS culture medium, 6-BA, NAA, sucrose, etc., to optimize the growth environment of chrysanthemum and promote the healthy development of stem tissue and seedlings; thirdly, by strictly controlling the culture environment Temperature, light intensity and light time provide ideal growth conditions for the rapid reproduction of chrysanthemums, ensuring the smooth growth of seedlings during the cultivation process; fourth, seedlings that have passed the breeding period are transplanted to a greenhouse environment, covered with transparent plastic film for insulation, and regularly sprayed with nutrient solution and sterilized to ensure the good development and disease resistance of the seedlings; finally, by gradually increasing light and fertilization, the root development and growth of the seedlings are effectively promoted, ultimately achieving a high survival rate of chrysanthemum reproduction and rapid cultivation, which not only increases the speed of chrysanthemum reproduction, but also ensures environmental control and resource optimization during the reproduction process, which is suitable for large-scale production of chrysanthemum planting.

[0014] Optionally, for some chrysanthemum rhizomes differentiation, adding 0.03-0.6 mg / L can promote the induced differentiation of cluster seedlings. The test on the induced growth of axillary buds of new chrysanthemum rhizomes, the induction rate of adding N treatment for 10 days reached 60%, while the induction rate of not adding N treatment was only 20%.

[0015] By adopting the above technical scheme, the amount of nitrogen source (N) added in the culture medium can be precisely controlled to significantly increase the induction rate of axillary buds of chrysanthemum rhizomes, thereby promoting the rapid differentiation and reproduction of chrysanthemum seedlings, and ensuring that chrysanthemums can grow efficiently during the rhizome differentiation process. In the experiment, adding 0.03-0.6 mg / L nitrogen source can significantly improve the induction growth effect of axillary buds of chrysanthemum rhizomes, reaching an induction rate of 60%, while the induction rate is only 20% when no nitrogen source is added. By optimizing the use of nitrogen source, the chrysanthemum reproduction efficiency is effectively improved, and a higher seedling survival rate can be obtained in a shorter time, which provides important technical support for the large-scale reproduction and production of chrysanthemums.

[0016] Optionally, under the premise of appropriate N and KT concentrations and ratios, different chrysanthemum reproduction and growth rates can be obtained by adjusting the concentration of the culture solution. The culture solution is mainly composed of MS+6-BA2.0mg / L+ and NAA0.1mg / L, and the concentration should be between 0.03-4mg / L to maintain reproduction coordination.

[0017] By adopting the above technical scheme, the growth rate of chrysanthemum reproduction can be precisely controlled by adjusting the concentration of the culture medium under the optimized nitrogen source (N) and 6-benzylaminopurine (KT) concentrations and their ratios, thereby maintaining the coordination during the reproduction process. The culture medium is based on MS culture medium, and appropriate amounts of 6-BA and NAA are added. The concentration is controlled between 0.03-4 mg / L, which can adjust the growth rate and development state of chrysanthemum and ensure healthy growth and stable development during the reproduction process.

[0018] Optionally, the method for disinfecting and sterilizing the explants comprises the following steps: washing the explants with running water, washing powder solution, ethanol solution and sodium hypochlorite solution respectively and then washing with sterile water.

[0019] By adopting the above technical scheme, the explants can be effectively disinfected and sterilized, and contamination by microorganisms such as bacteria and fungi can be prevented, the cleanliness and sterile environment of the explants during chrysanthemum reproduction can be improved, and the healthy growth of the explants in subsequent cultivation can be ensured. By using running water, laundry detergent solution, ethanol solution and sodium hypochlorite solution for cleaning in sequence, and then rinsing thoroughly with sterile water, impurities and microorganisms on the surface of the explants can be effectively removed, thereby ensuring the establishment of sterile culture conditions.

[0020] Optionally, seedlings are produced by inducing buds from chrysanthemum stems, wherein leaves begin to swell 5 days after inoculation, and axillary buds begin to germinate 3 to 5 days after inoculation of stem segments with axillary buds. As the number of days increases, adventitious buds are induced from the base of the chrysanthemum stems. Compared with the method of inducing adventitious buds from chrysanthemum stems, the adventitious buds induced from stem segments with chrysanthemum stems produce fewer buds, but the bud formation time is shorter.

[0021] By adopting the above technical scheme, adventitious buds can be effectively induced on chrysanthemum stems and stem segments with axillary buds, thereby promoting the rapid growth of chrysanthemum seedlings. The leaves begin to swell 5 days after inoculation, the axillary buds germinate 3 to 5 days after inoculation of the stem segments with axillary buds, and adventitious buds can be induced at the base of the chrysanthemum stems. In comparison, although the adventitious buds induced by the stem segments with chrysanthemum stems produce fewer buds, the bud formation time is shorter, thereby accelerating the reproduction process of chrysanthemums and improving the reproduction efficiency.

[0022] Optionally, use chrysanthemum stems and stem segments with axillary buds for induction experiments, determine that stem segments with chrysanthemum stems have the best effect in inducing adventitious buds, subculture and root culture the induced chrysanthemum seedlings, determine that the suitable subculture culture medium is MS+6-BA2.0mg / L+NAA0.1mg / L, and the suitable rooting culture medium is 1 / 2MS+NAA0.5mg / L, and after hardening the seedlings, transplant them into the culture medium, control the light intensity, temperature and humidity, and spraying of nutrient solution.

[0023] By adopting the above technical scheme, the chrysanthemum breeding process can be optimized, adventitious buds can be induced by using stem segments with chrysanthemum stems, the best chrysanthemum seedling breeding effect can be obtained, and the healthy development of seedlings can be ensured through subculture and rooting culture. By selecting suitable subculture culture medium (MS+6-BA2.0mg / L+NAA0.1mg / L) and rooting culture medium (1 / 2MS+NAA0.5mg / L), as well as accurately controlling light intensity, temperature and humidity, and nutrient solution spraying, the survival rate and growth rate of chrysanthemum seedlings can be effectively improved.

[0024] Optionally, the induced chrysanthemum adventitious buds are cut off individually, cut into stem segments of about 2.0 cm, and then inoculated into subculture proliferation medium for cultivation. The stem seedlings are cut into segments for transfer reproduction each time thereafter, and each segment has 3 buds. In the reproduction and cultivation of chrysanthemum, the stem seedling segments grow fast, and each segment produces multiple clustered seedlings, with an average proliferation coefficient of 4 to 7 times. The height of the stem seedlings is also uneven. After culturing in nutrient solution for 2 months, the height of the clustered seedlings is mostly 5 to 7 cm, and the proliferation rate of the adventitious buds of chrysanthemum seedlings is improved.

[0025] By adopting the above technical scheme, the reproduction efficiency of chrysanthemum can be significantly improved. The adventitious buds are cut and transferred to the subculture proliferation medium. Each cut is cultured with 3 buds, thereby promoting the rapid growth of stem seedlings and the efficient proliferation of clustered seedlings. The rapid proliferation of chrysanthemum stem seedlings is achieved, and the proliferation coefficient reaches 47cm, which significantly improves the proliferation rate of adventitious buds of chrysanthemum seedlings.

[0026] Optionally, MS+6-BA2.0mg / L+NAA0.1mg / L is a suitable subculture culture medium. Chrysanthemum seedlings over 3.0cm in height are cut and inoculated in the culture medium. Roots will appear after 5 days, and 60% of them will appear after 8 days. After 15 days, the roots will grow to 1-2.5cm, and the seedlings will grow to 3-5.0cm. The rooting situation will be observed after 20 days. The rooting rate of the culture medium can reach 100%.

[0027] By adopting the above technical scheme, the rooting and growth of chrysanthemum seedlings can be effectively promoted. Subculture is carried out in MS+6-BA2.0mg / L+NAA0.1mg / L culture medium, and efficient rooting of chrysanthemum seedlings is successfully achieved. Rooting begins within 5 days, 60% of the seedlings take root after 8 days, the roots are 1 to 2.5cm long after 15 days, and the rooting rate reaches 100% within 20 days, which greatly improves the survival rate and growth rate of chrysanthemum seedlings, and provides an efficient technical method for large-scale reproduction and planting of chrysanthemums.

[0028] In summary, the present application includes at least one of the following beneficial technical effects:

[0029] 1. Seedlings were produced by inducing buds from chrysanthemum stems. The leaves began to swell 5 days after inoculation, and the axillary buds of the chrysanthemum stems began to germinate 3 to 5 days after inoculation of the stem segments with axillary buds. As the number of days increased, adventitious buds were induced from the base of the chrysanthemum stems. Compared with the method of inducing adventitious buds from chrysanthemum stems, the adventitious buds induced from the stem segments with chrysanthemum stems produced fewer buds, but the bud formation time was shorter.

[0030] 2. In the reproduction and cultivation of chrysanthemum, the stem seedling segments grow fast, and each segment produces multiple clustered seedlings, with an average proliferation coefficient of 4 to 7 times. The height of the stem seedlings also varies. After culturing in nutrient solution for 2 months, the height of the clustered seedlings is mostly 5 to 7 cm, and the proliferation rate of adventitious buds of chrysanthemum seedlings is improved. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 It is a schematic diagram of the process of the present invention;

[0032] Figure 2 It is the chrysanthemum growth chart of the present invention. DETAILED DESCRIPTION

[0033] The following is combined with Figure 1-2 Further details of this application

[0034] The present application discloses a method for accelerating the reproduction of chrysanthemums using a liquid culture medium. Figure 1-Figure 2 As shown, a method for accelerating chrysanthemum propagation using liquid culture medium comprises the following steps:

[0035] Step 1: Cut the chrysanthemum stems into 1 cm × 1 cm squares in a culture box, lay the stems upward on the base soaked in induction culture solution, cut the stem tissue with axillary buds into small sections of 2 to 3 cm, and obliquely insert them into the induction culture solution for culture. The culture solution is selected from MS + 6-BA 2.0 mg / L +, NAA 0.1 mg / L, α-naphthyl acetic acid, 2-4 mg / L, 6-benzylaminopurine, sucrose;

[0036] Step 2: After the chrysanthemum stem tubers and stem tissues with axillary buds are induced to sprout, the tender shoots are cut out for further cultivation, and new chrysanthemum seedlings are continued to be propagated. MS is selected as the culture medium, and the hormones MS+6-BA2.0mg / L+, NAA0.1mg / L and 6-benzylaminopurine, potassium dihydrogen phosphate are added. The root growth culture solution is selected from MS, and NAA0.3mg / L base solution is added. Agar 4.5g / L and sucrose 20g / L are added to the culture box;

[0037] Step 3: The temperature for chrysanthemum reproduction and cultivation is 10-15°C, the light intensity is 1000-1500lx, and the lighting time is 6h / d.

[0038] Step 4. Transport the chrysanthemum seedlings with rhizomes of 1 to 2 cm to a room with a room temperature of 10° to 24°. Control the temperature at about 23°C, expose them to sunlight for 5 days, receive sufficient photosynthesis, spray nutrient solution manually, cover with plastic transparent film to keep warm and prevent water loss. When cultivating and growing, wash the chrysanthemum seedlings with warm water, wash and remove the culture solution on the rhizomes, transplant the chrysanthemum seedlings into the seedling tray sterilized with potassium permanganate, spray water manually, build an arch shed with plastic film on the seedling tray to keep warm and moisturize, gradually increase the photosynthetic exposure after 3 days of cultivation, sterilize and disinfect and fertilize once every 3 days.

[0039] Some chrysanthemum rhizomes differentiate, adding 0.03-0.6 mg / L can promote the induced differentiation of cluster seedlings. The test on the induced growth of axillary buds of new chrysanthemum rhizomes, the induction rate of adding N treatment for 10 days reached 60%, and the induction rate of not adding N treatment was only 20%.

[0040] Under the premise of appropriate N and KT concentrations and ratios, different chrysanthemum reproduction and growth rates can be obtained by adjusting the concentration of the culture medium. The culture medium is mainly composed of MS+6-BA2.0mg / L+ and NAA0.1mg / L, and the concentration should be between 0.03-4mg / L to maintain reproduction coordination.

[0041] The method for disinfecting and sterilizing explants comprises the following steps: washing the explants with running water, washing powder solution, ethanol solution and sodium hypochlorite solution respectively and then washing with sterile water.

[0042] Seedlings were produced by inducing buds from chrysanthemum stems. The leaves began to swell 5 days after inoculation, and the axillary buds of the chrysanthemum stems began to germinate 3 to 5 days after inoculation of the stem segments with axillary buds. As the days went by, adventitious buds were induced from the base of the chrysanthemum stems. Compared with the method of inducing adventitious buds from chrysanthemum stems, the adventitious buds induced from the stem segments with chrysanthemum stems produced fewer buds, but the bud formation time was shorter.

[0043] Chrysanthemum stems and stem segments with axillary buds were used for induction experiments, and it was determined that the stem segments with chrysanthemum stems had the best effect in inducing adventitious buds. The induced chrysanthemum seedlings were subcultured and rooted, and it was determined that the suitable subculture medium was MS+6-BA2.0mg / L+NAA0.1mg / L, and the suitable rooting medium was 1 / 2MS+NAA0.5mg / L. After hardening the seedlings, they were transplanted into the culture medium, and the light intensity, temperature and humidity, and the spraying of nutrient solution were controlled.

[0044] The present application discloses a method for accelerating the reproduction of chrysanthemums using liquid culture medium, which significantly improves the reproduction efficiency and growth quality of chrysanthemums. Through systematic culture steps, optimized hormone ratios and precise environmental control, the technical effect of rapid reproduction and growth of chrysanthemums is successfully achieved. First, during the culture process, the chrysanthemum stems are cut into small segments and placed in an induction culture solution. The formula of the culture solution (such as MS+6-BA2.0mg / L+, NAA0.1mg / L) is optimized to effectively stimulate the differentiation of chrysanthemum axillary buds and the rapid germination of new buds. This step promotes the rapid germination of stem segments under the action of hormones at appropriate concentrations, and the number and quality of new buds are relatively high, thereby accelerating the reproduction rate of chrysanthemums. By transplanting chrysanthemum seedlings into subculture culture medium and rooting culture medium, the chrysanthemum is successfully promoted. The growth and root development of chrysanthemum seedlings. In the subculture stage, the appropriate culture medium MS+6-BA2.0mg / L+NAA0.1mg / L and the rooting medium 1 / 2MS+NAA0.5mg / L effectively promoted the reproduction and rooting of chrysanthemum seedlings. After cultivation and hardening, the survival rate and rooting rate of the seedlings were significantly improved. Secondly, the liquid culture medium method not only accelerated the growth of chrysanthemum, but also improved its reproduction coefficient. Under the condition of an average proliferation coefficient of 41500lx, the rapid growth of chrysanthemum seedlings was successfully promoted. During the rooting process, the hormone concentration in the culture solution had a significant effect on the root development. The root system of the seedlings was 52.5cm, reaching a rooting rate of 100%, providing high-quality root development support for later cultivation.

[0045] In addition, it also includes effective disinfection and sterilization steps for explants, using running water, laundry detergent liquid, ethanol solution and sodium hypochlorite solution for thorough cleaning, effectively removing pathogens and ensuring a sterile environment, thereby improving the survival rate and health of chrysanthemum reproduction, reducing the breeding of pathogens, and avoiding cultivation failures due to contamination; it optimizes the technical conditions of each link in the chrysanthemum reproduction process, and through reasonable hormone ratios and environmental control, greatly improves the chrysanthemum reproduction efficiency and seedling growth quality, and realizes rapid and large-scale reproduction of chrysanthemums.

[0046] Example 1

[0047] Some chrysanthemum rhizomes differentiate, adding 0.05-0.8 mg / L can promote the induced differentiation of cluster seedlings. The test on the induction growth of axillary buds of new chrysanthemum rhizomes, the induction rate of adding N for 12 days reached 80%, and the induction rate of not adding N was only 40%.

[0048] Under the premise of appropriate N and KT concentrations and ratios, different chrysanthemum reproduction and growth rates can be obtained by adjusting the concentration of the culture medium. The culture medium is mainly composed of MS+6-BA2.0mg / L+ and NAA0.1mg / L, and the concentration should be between 0.05-6mg / L to maintain reproduction coordination.

[0049] Example 2

[0050] Some chrysanthemum rhizomes differentiate, adding 0.07-1mg / L can promote the induced differentiation of cluster seedlings. The test on the induction growth of axillary buds of new chrysanthemum rhizomes, the induction rate of adding N treatment for 14 days reached 100%, and the induction rate of not adding N treatment was only 60%.

[0051] Under the premise of appropriate N and KT concentrations and ratios, different chrysanthemum reproduction and growth rates can be obtained by adjusting the concentration of the culture medium. The culture medium is mainly composed of MS+6-BA2.0mg / L+ and NAA0.1mg / L, and the concentration should be between 0.07-8mg / L to maintain reproduction coordination.

[0052] Comparative Example 1

[0053] The induced chrysanthemum adventitious buds are cut off individually, cut into stem segments of about 2.0 cm, and then inoculated into subculture proliferation medium for cultivation. Each time thereafter, the stem seedlings are cut into segments for transfer reproduction, and each segment has 3 buds. In the reproduction and cultivation of chrysanthemum, the stem seedling segments grow fast, and each segment produces multiple clustered seedlings, with an average proliferation coefficient of 4 to 7 times. The height of the stem seedlings is also uneven. After culturing in nutrient solution for 2 months, the height of the clustered seedlings is mostly 5 to 7 cm, and the proliferation rate of the adventitious buds of chrysanthemum seedlings is improved.

[0054] MS+6-BA2.0mg / L+NAA0.1mg / L is a suitable subculture culture medium. Chrysanthemum seedlings over 3.0cm in height were cut and inoculated in the culture medium. They took root after 5 days, and 60% took root after 8 days. After 15 days, the roots grew to 1-2.5cm, and the seedlings grew to 3-5.0cm. The rooting was observed after 20 days, and the rooting rate of the culture medium reached 100%.

[0055] The above are all preferred embodiments of the present application, and the protection scope of the present application is not limited thereto. Therefore, any equivalent changes made according to the structure, shape, and principle of the present application should be included in the protection scope of the present application.

Claims

1. A method for accelerating chrysanthemum reproduction using a liquid culture medium, characterized in that: The following steps are involved: Step 1: Cut the chrysanthemum stems into 1 cm × 1 cm squares in a culture box, lay the stems upward on the base soaked in induction culture solution, cut the stem tissue with axillary buds into small sections of 2 to 3 cm, and obliquely insert them into the induction culture solution for culture. The culture solution is selected from MS + 6-BA 2.0 mg / L +, NAA 0.1 mg / L, α-naphthyl acetic acid, 2-4 mg / L, 6-benzylaminopurine, sucrose; Step 2: After the chrysanthemum stem tubers and stem tissues with axillary buds are induced to sprout, the tender shoots are cut out for further cultivation, and new chrysanthemum seedlings are continued to be propagated. MS is selected as the culture medium, and the hormones MS+6-BA2.0mg / L+, NAA0.1mg / L and 6-benzylaminopurine, potassium dihydrogen phosphate are added. The root growth culture solution is selected from MS, and NAA0.3mg / L base solution is added. Agar 4.5g / L and sucrose 20g / L are added to the culture box; Step 3: The temperature for chrysanthemum propagation and cultivation is 10-15°C, the light intensity is 1000-1500lx, and the light time is 6h / d; Step 4. Transport the chrysanthemum seedlings with rhizomes of 1 to 2 cm to a room with a room temperature of 10° to 24°. Control the temperature at about 23°C, expose them to sunlight for 5 days, receive sufficient photosynthesis, spray nutrient solution manually, cover with plastic transparent film to keep warm and prevent water loss. When cultivating and growing, wash the chrysanthemum seedlings with warm water, wash and remove the culture solution on the rhizomes, transplant the chrysanthemum seedlings into the culture seedling tray sterilized with potassium permanganate, spray water manually, build an arch shed with plastic film on the seedling tray to keep warm and moisturize. After 3 days of cultivation, gradually increase the photosynthetic exposure, and sterilize and fertilize once every 3 days.

2. The method for accelerating chrysanthemum reproduction using liquid culture medium according to claim 1, characterized in that: Some chrysanthemum rhizomes differentiate, adding 0.03-0.6 mg / L can promote the induced differentiation of cluster seedlings. The test on the induced growth of axillary buds of new chrysanthemum rhizomes, the induction rate of adding N treatment for 10 days reached 60%, and the induction rate of not adding N treatment was only 20%.

3. The method for accelerating chrysanthemum reproduction using liquid culture medium according to claim 1, characterized in that: Under the premise of appropriate N and KT concentrations and ratios, different chrysanthemum reproduction and growth rates can be obtained by adjusting the concentration of the culture medium. The culture medium is mainly composed of MS+6-BA2.0mg / L+ and NAA0.1mg / L, and the concentration should be between 0.03-4mg / L to maintain reproduction coordination.

4. The method for accelerating chrysanthemum reproduction using liquid culture medium according to claim 1, characterized in that: The method for disinfecting and sterilizing the explant comprises the following steps: washing the explant with running water, washing powder solution, ethanol solution and sodium hypochlorite solution respectively, and then washing with sterile water.

5. The method for accelerating chrysanthemum reproduction using liquid culture medium according to claim 1, characterized in that: Seedlings were produced by inducing buds from chrysanthemum stems. The leaves began to swell 5 days after inoculation, and the axillary buds of the chrysanthemum stems began to germinate 3 to 5 days after inoculation of the stem segments with axillary buds. As the days went by, adventitious buds were induced from the base of the chrysanthemum stems. Compared with the method of inducing adventitious buds from chrysanthemum stems, the adventitious buds induced from the stem segments with chrysanthemum stems produced fewer buds, but the bud formation time was shorter.

6. The method for accelerating chrysanthemum reproduction using liquid culture medium according to claim 1, characterized in that: Chrysanthemum stems and stem segments with axillary buds were used for induction experiments, and it was determined that the stem segments with chrysanthemum stems had the best effect in inducing adventitious buds. The induced chrysanthemum seedlings were subcultured and rooted, and it was determined that the suitable subculture medium was MS+6-BA2.0mg / L+NAA0.1mg / L, and the suitable rooting medium was 1 / 2MS+NAA0.5mg / L. After hardening the seedlings, they were transplanted into the culture medium, and the light intensity, temperature and humidity, and the spraying of nutrient solution were controlled.

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