Breeding method of cinnamomum camphora and application of breeding method

Abstract

The invention provides a cinnamomum camphora breeding method and application thereof, and belongs to the technical field of plant cultivation method.The breeding method comprises the following steps that 1, terminal bud stems of current-year branches of cinnamomum camphora are selected as explants, and clustered seedlings are obtained through culture; soaking the clustered seedlings in a composite bactericide for 10-15 minutes, soaking the base parts of the clustered seedlings in a growth regulator solution for 10-15 minutes, and then inoculating the clustered seedlings into a rooting culture medium for rooting culture to obtain tissue culture seedlings; (2) soil with the pH value of 5-6 is selected to be loosened, sterilized, fertilized and watered; (3) cutting the tissue culture seedlings into soil; (4) disinfecting the tissue culture seedlings again after cuttage, and spraying a rooting powder nutrient solution on soil around the tissue culture seedlings; and (5) covering with a shading net, culturing for 35-40 days, removing the shading net, and continuing maintenance management for 50-60 days. The breeding method is high in rooting rate, average plant height and survival rate.

Description

Technical Field

[0001] The invention belongs to the technical field of plant cultivation methods, and particularly relates to a breeding method of borneol camphor and application thereof. Background Art

[0002] Camphor is a special camphor tree variety. Unlike other camphor trees, Camphor is rich in dextrorotatory borneol (borneol), which is one of the main sources of natural borneol, a very precious natural substance with a wide range of medicinal value and commercial uses. Camphor is a plant with extremely high value. Its discovery not only brings valuable natural resources to China, but also provides a new source of raw materials for industries such as medicine, spices and chemicals. In recent years, the scale of Camphor plantations has expanded dramatically, and the market demand for high-quality Camphor seedlings has continued to grow. However, the high transportation cost of Camphor seedlings, the inability to directly transport them to the mountains for planting, and the low survival rate of planting have restricted the development of the Camphor seedling industry.

[0003] The breeding methods of Camphor camphor mainly involve two methods: cutting seedling cultivation and tissue culture. Both methods have certain technical problems in actual operation. Technical problems of cutting seedling cultivation: (1) Unstable survival rate: Although the survival rate of cutting seedling cultivation can reach more than 80%, the actual survival rate may fluctuate due to environmental factors, operating techniques and other factors. The survival rate directly affects the quality and quantity of seedlings. (2) High operating technology requirements: Cutting seedling cultivation has certain requirements on the skills of the operator. Improper operation may cause the cuttings to be difficult to root or grow poorly. For example, the selection, treatment, disinfection and bed management of cuttings all require precise control. (3) Environmental conditions: The success of cutting seedling cultivation is greatly affected by environmental conditions, such as temperature, humidity, and light. For example, a specific temperature range and suitable humidity are required to promote rooting. (4) Pest and disease control: Cuttings are easily attacked by pests and diseases during the process, and effective prevention and control measures need to be taken. Pests and diseases will affect the survival rate of cuttings and the quality of seedlings. Technical problems with tissue culture: (1) High technical threshold: Tissue culture requires strict sterile conditions and professional laboratory equipment. The technical threshold is high and requires special training and technical support. (2) Complex operation: The process of tissue culture is relatively complicated, including disinfection, culture medium preparation, inoculation, culture and other steps. Each step needs to be strictly controlled to ensure the success rate of culture. (3) High cost: The equipment and consumables required for tissue culture are expensive. Coupled with the labor cost of professional technicians, the entire process is expensive.

[0004] The Chinese invention patent with the announcement number CN 108781957A discloses a method for factory-based seedling cultivation of borneol camphor seedlings, wherein the seedlings are transferred to 128-mesh plug trays or 105-mesh plug trays for planting for 1.5-2.5 months after seedling hardening, and then 50-mesh tree plug trays or nutrient bags are used to continue planting for 2.5-4 months, thus accelerating the seedling cultivation speed of borneol camphor seedlings, and also reducing the transportation cost of borneol camphor, making borneol camphor trees easy to carry when planted on the mountain. However, the planting survival rate of the borneol camphor seedlings of the method is not ideal, resulting in high seedling cultivation costs. The demand of the existing market cannot be met.

[0005] Therefore, a novel breeding method and application of borneol camphor are badly needed. Summary of the invention

[0006] The invention aims to provide a breeding method of borneol camphor and application thereof.

[0007] In order to achieve the above object, the present invention provides the following technical solutions: A breeding method for borneol camphor, the breeding method comprising the following steps: (1): Select the terminal bud stem segment of the current year's branch of Camphora borneol as the explant, sterilize it, and inoculate it into the primary culture medium to induce callus tissue; Inoculating the callus tissue into a proliferation and subculture medium, proliferating and subculturing, and obtaining clustered seedlings; The clump seedlings are soaked in a compound fungicide, then soaked in a growth regulator solution, and then inoculated in a rooting medium for cultivation to obtain tissue culture seedlings; The composite fungicide includes: carbendazim 0.5-0.9μg / mL, galangin 3.2-3.7μg / mL, aloe-emodin 1.4-1.7μg / mL, celangulin 0.4-0.6μg / mL, matrine 0.7-1.2μg / mL, prepared with water; Rooting medium composition: 1 / 2MS, sucrose 12-14g / L, agar 20-22g / L and NAA 0.6-0.9mg / L; (2): Loosen the soil, sterilize, fertilize and water to obtain treated soil; (3): Transplanting the tissue culture seedlings into the soil treated in step (2); (4): After cutting, disinfect the tissue culture seedlings again, spray rooting powder nutrient solution on the soil around the tissue culture seedlings, then water them, insert bamboo strips and cover with film to seal; (5): Cover with a shade net and fix the four sides, cultivate under light conditions, and continue maintenance and management after removing the shade net.

[0008] Further, the breeding method of borneol camphor comprises the following steps: (1) The terminal bud stem segments of the current year branches of Camphora borneol were selected as explants and disinfected; then they were inoculated into the primary culture medium to induce callus tissue; Inoculating the callus tissue into a proliferation subculture medium for proliferation subculture to obtain clustered seedlings; The clump seedlings are first soaked in a composite fungicide for 10-15 minutes, then soaked at the base in a growth regulator solution for 10-15 minutes, and then inoculated in a rooting medium for rooting culture to obtain tissue culture seedlings; The composite fungicide includes the following components: carbendazim 0.5-0.9 μg / mL, galangin 3.2-3.7 μg / mL, aloe-emodin 1.4-1.7 μg / mL, celangulin 0.4-0.6 μg / mL, matrine 0.7-1.2 μg / mL, prepared with water; The rooting medium consists of 1 / 2MS, sucrose 12-14 g / L, agar 20-22 g / L and NAA 0.6-0.9 mg / L; (2) Select soil with a pH value between 5 and 6, loosen the soil, sterilize, fertilize, and water to obtain treated soil; (3) On a sunny day at 13-17°C, transplant the rooted tissue culture seedlings into the soil treated in step (2), with the leaves aligned in the same direction and the buds placed one centimeter above the soil surface. The row and plant spacing is 6-8 centimeters. (4) After cutting, disinfect the tissue culture seedlings again, spray the soil around the tissue culture seedlings with rooting powder nutrient solution, spray 300-500mL of rooting powder nutrient solution on each tissue culture seedling on average, then water them, insert bamboo strips, cover with film to seal, and control the humidity at 70%-90%; (5) Cover with a shade net with a shading rate of 87-92% and fix the four sides. Cultivate under the condition of 7000-9000 lux for 35-40 days, remove the shade net; light 10000-12000 lux, control the growth temperature at 16-20℃, and continue maintenance and management for 50-60 days.

[0009] At present, in the market, carbendazim solution is preferred as a bactericide in the breeding process to prevent pathogens from invading from wounds, but in the breeding process of camphor camphor, it is found that the use effect is not ideal, affecting the survival rate. The present invention has a good bactericidal effect by preparing a composite bactericide by compounding carbendazim and multiple components, thereby improving the survival rate of camphor camphor breeding. The concentration of carbendazim used in the present invention can not only play a bactericidal effect but also not cause damage to the fringe strips, galangin has a strong antibacterial activity, and the bactericidal effect can be further enhanced; aloe-emodin helps to promote wound healing and reduce the damage of the fringe strips during the treatment process; celangulin has antibacterial activity and can inhibit the growth of various pathogenic microorganisms; matrine has a wide range of antibacterial and antiviral effects, and can further reduce the pathogens on the fringe strips. These components work together to produce a synergistic effect, that is, each component cooperates with each other, enhances the overall bactericidal effect, and then improves the survival rate of camphor camphor breeding.

[0010] Furthermore, the preparation method of the rooting powder nutrient solution is: (1) Mix burdock root, astragalus root and eucalyptus leaves in a mass ratio of 1: (0.5-0.7): (1.2-1.4), crush to less than 80-100 mesh, and obtain a mixture; add 10-13 mass parts of water to 1 mass part of the mixture to obtain a mixed solution; continue to add Lactobacillus brevis to carry out anaerobic fermentation at a fermentation temperature of 32-35° C. and a fermentation time of 15-20 hours; sterilize to obtain a sterilized product; continue to add Lactobacillus plantarum to carry out anaerobic fermentation at a fermentation temperature of 32-35° C. and a fermentation time of 10-12 hours; after sterilization, filter through a 0.22 μm filter membrane to obtain a filtrate; concentrate and dry the filtrate to obtain a baking powder; (2) Mix potassium α-naphthylacetate (Shanghai Yuanye Biotechnology Co., Ltd.), potassium 3-indolebutyrate (Shanghai Yuanye Biotechnology Co., Ltd.) and fermentation powder to obtain a mixed powder. Dilute the mixed powder and water at a ratio of 400-500 mg:1 L to obtain a rooting powder nutrient solution.

[0011] Furthermore, the concentration of Lactobacillus brevis in the mixed solution is 10 6 -10 7 CFU / mL.

[0012] Furthermore, the concentration of Lactobacillus plantarum in the sterilized material is 10 7 -10 8 CFU / mL.

[0013] Furthermore, the mass ratio of potassium α-naphthylacetate and potassium 3-indolebutyrate to baking powder is (0.2-0.5):1:(1.4-1.7).

[0014] The rooting powder nutrient solution of the present invention is added with potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder at the same time, which can improve the rooting rate of borneol camphor breeding. Potassium α-naphthylacetate can promote the growth of the root system of borneol camphor, and is particularly effective in promoting the formation of lateral roots and adventitious roots. An appropriate concentration can significantly improve the rooting rate, and too high or too low a concentration may inhibit rooting. Potassium 3-indolebutyrate is also an auxin-type plant growth regulator, which is particularly effective in promoting rooting and helps to improve the success rate of rooting of cuttings. The active components in the baking powder can stimulate the growth of the plant root system and improve the rooting rate. The synergistic effect of these components can promote the rapid formation and growth of the root system, thereby improving the success rate of cuttings. And when these three components are compounded at a specific ratio, the average rooting number of breeding can be improved.

[0015] Furthermore, the growth regulator solution includes the following components: 6-BA 6.5-8.6 mg / L, NAA 1.2-3.5 mg / L, IBA 1.3-2.6 mg / L, IAA 2.4-3.5 mg / L, and plant extract 2.4-4.6 mg / L, and is prepared with water.

[0016] 6-Benzylaminopurine (6-BA), naphthylacetic acid (NAA), indolebutyric acid (IBA) and indoleacetic acid (IAA) are all commercially available products.

[0017] Furthermore, the preparation method of the plant extract is: (1) Mixing Platycladus orientalis leaves, agar-agar and astragalus, and crushing them to a particle size of less than 80-100 meshes to obtain a mixed material; (2) The mixed material is subjected to supercritical extraction at 40-45°C, 30-35MPa, CO 2 Extraction was carried out for 100-120 min at a flow rate of 17-20 L / h to obtain the first fraction, and then the first fraction was extracted at 25-30 °C, 25-30 MPa, CO 2 Extraction for 60-80 min at a flow rate of 17-20 L / h to obtain a second fraction; (3) The first fraction and the second fraction are mixed to obtain a plant extract.

[0018] Furthermore, the mass ratio of Platycladus orientalis leaves, Amanita charantia and Astragalus membranaceus is 1:(1.3-1.5):(0.3-0.7).

[0019] The present invention can improve the average plant height of borneol camphor by using a growth regulator liquid. By reasonably adding 6-BA, NAA, IBA, IAA and plant extracts to the growth regulator liquid, the height of the plant obtained in the breeding process can be effectively improved. These components act on the growth and development of the plant through multiple mechanisms such as promoting cell division, cell elongation, bud formation and root growth, thereby achieving the purpose of increasing the plant height.

[0020] The invention also provides application of the borneol camphor tree breeding method.

[0021] Compared with the prior art, the advantages and beneficial effects of the present invention are: 1. The composite fungicide prepared by compounding carbendazim and multiple components has good bactericidal effect and improves the survival rate of borneol camphor breeding.

[0022] 2. Adding potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder to the rooting powder nutrient solution of the present invention can improve the rooting rate of borneol camphor breeding. When these three components are compounded in a specific ratio, the average rooting number of breeding can be improved.

[0023] 3. The present invention can increase the average plant height of borneol camphor by using a growth regulator solution. DETAILED DESCRIPTION

[0024] 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 them. 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.

[0025] Some of the raw materials used in the present invention were purchased from the following manufacturers: Carbendazim was purchased from Hubei Qiniu Chemical Technology Co., Ltd.; Galangin, Sichuan Jingcuibao Biotechnology Co., Ltd.; Aloe-emodin, Shanghai Yuanye Biotechnology Co., Ltd.; Celangulin, Shanghai Yuanye Biotechnology Co., Ltd.; Matrine, Shanghai Yuanye Biotechnology Co., Ltd.; Potassium α-naphthyl acetate, Shanghai Yuanye Biotechnology Co., Ltd.; Potassium 3-indolebutyrate, Shanghai Yuanye Biotechnology Co., Ltd.; Lactobacillus brevis, accession number: SHBCCD14346, was purchased from Shanghai Microbiological Collection Center.

[0026] Lactobacillus plantarum, accession number: CCTCCAB2010210, was purchased from China Center for Type Culture Collection.

[0027] The composition of the primary culture medium was MS medium, sucrose 25 g / L, agar 25 g / L, 6-BA 1.0 mg / L, and NAA 0.8 mg / L.

[0028] The composition of the proliferation subculture medium is MS medium, sucrose 20g / L, agar 25g / L, 6-BA 2mg / L, and NAA 2mg / L.

[0029] Example 1 This embodiment provides a breeding method for borneol camphor, the breeding method comprising the following steps: (1) selecting the terminal bud stem segment of the current year branch of Camphor camphor as an explant and performing a disinfection treatment; inoculating the explant into a primary culture medium to induce callus tissue; inoculating the callus tissue into a proliferation subculture medium for proliferation subculture to obtain clump seedlings; soaking the clump seedlings in a composite fungicide for 12 minutes, then soaking the base in a growth regulator solution for 12 minutes, and then inoculating the seedlings in a rooting culture medium for rooting culture to obtain tissue culture seedlings; The rooting medium is 1 / 2MS, sucrose 13g / L, agar 21g / L, NAA 0.8mg / L; (2) Select soil with a pH value of 5.5, loosen the soil, sterilize, fertilize, and water to obtain treated soil; (3) On a sunny day at 15°C, transplant the rooted tissue culture seedlings into the soil treated in step (2), with the leaves aligned in the same direction and the buds placed one centimeter above the soil surface. The row and plant spacing is 7 centimeters. (4) After cutting, disinfect the tissue culture seedlings again, spray the soil around the tissue culture seedlings with rooting powder nutrient solution, spray 400 mL per tissue culture seedling on average, then water them, insert bamboo strips and cover them with white film to seal them, and ensure that the humidity is controlled at 80%; (5) Cover with a shade net with a shading rate of 90%, fix the four sides, and provide a light of 8000 lux. Cultivate for 36 days, remove the shade net, provide a light of 11000 lux, control the growth temperature at 18°C, and continue maintenance and management for 55 days.

[0030] The composite fungicide includes the following components: 0.7 μg / mL of carbendazim, 3.4 μg / mL of galangin, 1.5 μg / mL of aloe-emodin, 0.5 μg / mL of celangulin, and 0.9 μg / mL of matrine, and is prepared with water.

[0031] The preparation method of rooting powder nutrient solution is: (1) Burdock root, astragalus root and eucalyptus leaves in a mass ratio of 1:0.6:1.3 were mixed and crushed to less than 100 mesh to obtain a mixture. 12 parts by mass of water were added to 1 part by mass of the mixture to obtain a mixed solution. Lactobacillus brevis was added to the mixture. The concentration of Lactobacillus brevis in the mixed solution was 10 6CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 33°C and a fermentation time of 17h. After sterilization, a sterilized product was obtained, and Lactobacillus plantarum was added. The concentration of Lactobacillus plantarum in the sterilized product was 10 8 CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 33°C and a fermentation time of 11 h. After sterilization, the filtrate was filtered through a 0.22 μm filter membrane to obtain the filtrate, which was concentrated and dried to obtain the baking powder; (2) Potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder are mixed in a mass ratio of 0.4:1:1.5 to obtain a mixed powder, and the mixed powder is diluted with water in a ratio of 450 mg:1 L to obtain a rooting powder nutrient solution.

[0032] The growth regulator liquid includes the following components: 7.5 mg / L 6-BA, 2.4 mg / L NAA, 2.2 mg / L IBA, 3.1 mg / L IAA, and 4.0 mg / L plant extract, and is prepared with water.

[0033] The preparation method of the plant extract is: (1) Mix Platycladus orientalis leaves, Gelidium amanshurica, and Astragalus membranaceus in a mass ratio of 1:1.4:0.5, and grind them into particles with a size of less than 100 meshes to obtain a mixture; (2) The mixed material is subjected to supercritical extraction at 42°C, 32MPa, CO 2 The first fraction was obtained by extraction at a flow rate of 18 L / h for 110 min. The first fraction was then concentrated under the conditions of 27 °C, 28 MPa, and CO 2 Extraction was performed for 70 min at a flow rate of 18 L / h to obtain a second fraction; (3) The first fraction and the second fraction are mixed to obtain a plant extract.

[0034] Example 2 This embodiment provides a breeding method for borneol camphor, the breeding method comprising the following steps: (1) selecting the terminal bud stem segment of the current year branch of Camphor camphor as an explant and performing a disinfection treatment; inoculating the explant into a primary culture medium to induce callus tissue; inoculating the callus tissue into a proliferation subculture medium for proliferation subculture to obtain clump seedlings; soaking the clump seedlings in a composite fungicide for 10 minutes, then soaking the base in a growth regulator solution for 10 minutes, and then inoculating the seedlings into a rooting culture medium for rooting culture to obtain tissue culture seedlings; The rooting medium is 1 / 2MS, sucrose 12g / L, agar 20g / L, NAA 0.6mg / L; (2) Select soil with a pH value of 5, loosen the soil, sterilize, fertilize, and water to obtain treated soil; (3) On a sunny day at 13°C, transplant the rooted tissue culture seedlings into the soil treated in step (2), with the leaves aligned in the same direction and the buds placed one centimeter above the soil surface. The row and plant spacing is 6 centimeters. (4) After cutting, disinfect the tissue culture seedlings again, spray the rooting powder nutrient solution on the soil around the tissue culture seedlings, spray 300 mL per tissue culture seedling on average, then water them, insert bamboo strips and cover them with white film to seal them, and ensure that the humidity is controlled at 70%; (5) Cover with a shade net with a shading rate of 87%, fix the four sides, and provide light of 7000 lux. Cultivate for 35 days, remove the shade net, provide light of 10000 lux, control the growth temperature at 16°C, and continue maintenance and management for 50 days.

[0035] The composite fungicide includes the following components: 0.5 μg / mL of carbendazim, 3.2 μg / mL of galangin, 1.4 μg / mL of aloe-emodin, 0.4 μg / mL of celangulin, and 0.7 μg / mL of matrine, and is prepared with water.

[0036] The preparation method of rooting powder nutrient solution is: (1) Burdock root, astragalus root and eucalyptus leaves in a mass ratio of 1:0.5:1.2 were mixed and crushed to less than 80 meshes to obtain a mixture. 10 parts by mass of water were added to 1 part by mass of the mixture to obtain a mixed solution. Lactobacillus brevis was added to the mixture. The concentration of Lactobacillus brevis in the mixed solution was 10 6 CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 32°C and a fermentation time of 15 h. After sterilization, a sterilized product was obtained, and Lactobacillus plantarum was added. The concentration of Lactobacillus plantarum in the sterilized product was 10 7 CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 32°C and a fermentation time of 10 h. After sterilization, the filtrate was filtered through a 0.22 μm filter membrane to obtain the filtrate, which was concentrated and dried to obtain the baking powder; (2) Potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder are mixed in a mass ratio of 0.2:1:1.4 to obtain a mixed powder, and the mixed powder is diluted with water in a ratio of 400 mg:1 L to obtain a rooting powder nutrient solution.

[0037] The growth regulator liquid comprises the following components: 6.5 mg / L 6-BA, 1.2 mg / L NAA, 1.3 mg / L IBA, 2.4 mg / L IAA and 2.4 mg / L plant extract, and is prepared with water.

[0038] The preparation method of the plant extract is: (1) Mix Platycladus orientalis leaves, Gelidium amansii, and Astragalus membranaceus in a mass ratio of 1:1.3:0.3, and grind them into particles with a size of less than 80 meshes to obtain a mixture; (2) The mixed material is subjected to supercritical extraction at 40°C, 30MPa, CO 2 The first fraction was obtained by extraction at a flow rate of 17 L / h for 100 min. The first fraction was then extracted at 25 °C, 25 MPa, and CO 2 The second fraction was obtained by extraction at a flow rate of 17 L / h for 60 min. (3) The first fraction and the second fraction are mixed to obtain a plant extract.

[0039] Example 3 This embodiment provides a breeding method for borneol camphor, the breeding method comprising the following steps: (1) selecting the terminal bud stem segment of the current year branch of Camphor camphor as an explant and performing a disinfection treatment; inoculating the explant into a primary culture medium to induce callus tissue; inoculating the callus tissue into a proliferation subculture medium for proliferation subculture to obtain a clump seedling; soaking the clump seedling in a composite fungicide for 15 minutes, then soaking the base of the clump seedling in a growth regulator solution for 15 minutes, and then inoculating the clump seedling in a rooting culture medium for rooting culture to obtain a tissue culture seedling; The rooting medium is 1 / 2MS, sucrose 14g / L, agar 22g / L, NAA 0.9mg / L; (2) Select soil with a pH value of 6, loosen the soil, sterilize, fertilize, and water to obtain treated soil; (3) On a sunny day at 17°C, transplant the rooted tissue culture seedlings into the soil treated in step (2), with the leaves aligned in the same direction and the buds placed one centimeter above the soil surface. The row and plant spacing is 8 centimeters. (4) After cutting, disinfect the tissue culture seedlings again, spray the rooting powder nutrient solution on the soil around the tissue culture seedlings, spray 500mL per tissue culture seedling on average, then water them, insert bamboo strips and cover them with white film to seal them, and ensure that the humidity is controlled at 90%; (5) Cover with a shade net with a shading rate of 92%, fix the four sides, and provide light at 9000 lux. After culturing for 40 days, remove the shade net, provide light at 12000 lux, control the growth temperature at 20°C, and continue maintenance and management for 60 days.

[0040] The composite fungicide includes the following components: 0.9 μg / mL of carbendazim, 3.7 μg / mL of galangin, 1.7 μg / mL of aloe-emodin, 0.6 μg / mL of celangulin, and 1.2 μg / mL of matrine, and is prepared with water.

[0041] The preparation method of rooting powder nutrient solution is: (1) Burdock root, astragalus root and eucalyptus leaves in a mass ratio of 1:0.5:1.2 were mixed and crushed to less than 100 mesh to obtain a mixture. 13 parts by mass of water were added to 1 part by mass of the mixture to obtain a mixed solution. Lactobacillus brevis was added to the mixture. The concentration of Lactobacillus brevis in the mixed solution was 10 7 CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 35°C and a fermentation time of 20 h. After sterilization, a sterilized product was obtained, and Lactobacillus plantarum was added. The concentration of Lactobacillus plantarum in the sterilized product was 10 8 CFU / mL, anaerobic fermentation was carried out at a fermentation temperature of 35°C and a fermentation time of 12 h. After sterilization, the filtrate was filtered through a 0.22 μm filter membrane to obtain the filtrate, which was concentrated and dried to obtain the baking powder; (2) Potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder are mixed in a mass ratio of 0.5:1:1.7 to obtain a mixed powder, and the mixed powder is diluted with water in a ratio of 500 mg:1 L to obtain a rooting powder nutrient solution.

[0042] The growth regulator liquid comprises the following components: 8.6 mg / L 6-BA, 3.5 mg / L NAA, 2.6 mg / L IBA, 3.5 mg / L IAA and 4.6 mg / L plant extract, and is prepared with water.

[0043] The preparation method of the plant extract is: (1) Mix Platycladus orientalis leaves, Gelidium amanitae, and Astragalus membranaceus in a mass ratio of 1:1.5:0.7, and grind them into particles with a size of less than 100 meshes to obtain a mixture; (2) The mixed material is subjected to supercritical extraction at 45°C, 35MPa, CO 2 The first fraction was obtained by extraction at a flow rate of 20 L / h for 120 min. The first fraction was then concentrated to 30 °C, 30 MPa, and CO 2 Extraction was performed for 80 min at a flow rate of 20 L / h to obtain a second fraction; (3) The first fraction and the second fraction are mixed to obtain a plant extract.

[0044] Comparative Example 1 The difference between this comparative example and Example 1 is that the composite fungicide is replaced by a carbendazim aqueous solution with a concentration of 7 μg / mL.

[0045] Comparative Example 2 The difference between this comparative example and Example 1 is that the composite fungicide includes the following components: 3.4 μg / mL of carbendazim, 0.7 μg / mL of galangin, 0.5 μg / mL of aloe-emodin, 2.0 μg / mL of celangulin, and 0.4 μg / mL of matrine, and is prepared with water.

[0046] Comparative Example 3 The difference between this comparative example and Example 1 is that the preparation method of the rooting powder nutrient solution is: mixing potassium α-naphthylacetate and potassium 3-indolebutyrate in a mass ratio of 0.4:1 to obtain a mixed powder, and diluting the mixed powder and water in a ratio of 450 mg:1L to obtain a rooting powder nutrient solution.

[0047] Comparative Example 4 The difference between this comparative example and Example 1 is that potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder are mixed in a mass ratio of 1:1:1.

[0048] Comparative Example 5 The difference between this comparative example and Example 1 is that the growth regulator solution includes the following components: 6-BA 2.4 mg / L, NAA 9.5 mg / L, IBA 3.1 mg / L, IAA 2.2 mg / L, and plant extract 2.0 mg / L, and is prepared with water.

[0049] Comparative Example 6 The difference between this comparative example and Example 1 is that the growth regulator solution includes the following components: 6-BA 7.5 mg / L, NAA 4.4 mg / L, IBA 4.2 mg / L, IAA 3.1 mg / L, and is prepared with water.

[0050] Comparative Example 7 The difference between this comparative example and Example 1 is that burdock root, astragalus root and eucalyptus leaves are mixed in a mass ratio of 1:1:1.

[0051] Comparative Example 8 The difference between this comparative example and Example 1 is that Platycladus orientalis leaves, Gelidium amansii and Astragalus membranaceus are mixed in a mass ratio of 1:1:1.

[0052] Performance Testing Cultivation was performed according to the breeding methods of Examples 1-3 and Comparative Examples 1-8, 100 strains were cultivated in each group, and the growth of each treated borneol camphor tree was investigated. The results are shown in Table 1 below; Average root number = total number of roots of all plants / number of plants.

[0053] Table 1 Performance test results

[0054] From the above performance test results, it can be seen that the breeding effects of Examples 1-3 are good, especially the comprehensive performance of Example 1 is the most outstanding, which is mainly because the culture conditions such as the growth regulator, rooting powder nutrient solution and composite fungicide used produce a synergistic effect.

[0055] However, the comparative examples did not adopt the necessary technical solutions, which resulted in the corresponding performance tests being significantly worse than those of the embodiments. The composition of the composite fungicide in comparative examples 1-2 was changed, and the survival rate of borneol camphor breeding decreased. The composition of the rooting powder nutrient solution in comparative examples 3-4 and 7 was changed, and the rooting rate of borneol camphor breeding decreased. The composition of the growth regulator solution in comparative examples 5-6 and 8 was changed, and the average plant height of borneol camphor decreased. The above experimental results further demonstrate the importance of the technical solution defined in the present invention for its technical effect.

[0056] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A breeding method for borneol camphor, characterized in that, The breeding method comprises the following steps: (1): Select the terminal bud stem segment of the current year's branch of Camphora borneol as an explant, sterilize it; inoculate it into a primary culture medium to induce callus tissue; inoculate the callus tissue into a proliferation subculture medium, proliferate and subculture it to obtain cluster seedlings; soak the cluster seedlings in a compound fungicide, then soak them in a growth regulator solution, and then inoculate them into a rooting culture medium for cultivation to obtain tissue culture seedlings; The composite fungicide includes: carbendazim 0.5-0.9μg / mL, galangin 3.2-3.7μg / mL, aloe-emodin 1.4-1.7μg / mL, celangulin 0.4-0.6μg / mL, matrine 0.7-1.2μg / mL, prepared with water; The rooting medium consists of: 1 / 2MS, sucrose 12-14 g / L, agar 20-22 g / L and NAA 0.6-0.9 mg / L; (2): Loosen the soil, sterilize, fertilize and water to obtain treated soil; (3): Transplanting the tissue culture seedlings into the soil treated in step (2); (4): After cutting, disinfect the tissue culture seedlings again, spray rooting powder nutrient solution on the soil around the tissue culture seedlings, then water them, insert bamboo strips and cover with film to seal; (5): Cover with a shade net and fix the four sides, cultivate under light conditions, and continue maintenance and management after removing the shade net.

2. The breeding method of borneol camphor according to claim 1, characterized in that, The preparation method of the rooting powder nutrient solution is: (1) Mix burdock root, astragalus root and eucalyptus leaves in a mass ratio of 1: (0.5-0.7): (1.2-1.4), crush to less than 80-100 mesh, and obtain a mixture; add 10-13 mass parts of water to 1 mass part of the mixture to obtain a mixed solution; continue to add Lactobacillus brevis to carry out anaerobic fermentation at a fermentation temperature of 32-35° C. and a fermentation time of 15-20 hours; sterilize to obtain a sterilized product; continue to add Lactobacillus plantarum to carry out anaerobic fermentation at a fermentation temperature of 32-35° C. and a fermentation time of 10-12 hours; after sterilization, filter through a 0.22 μm filter membrane to obtain a filtrate; concentrate and dry the filtrate to obtain a baking powder; (2) Mix potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder to obtain a mixed powder, and dilute the mixed powder with water at a ratio of 400-500 mg:1 L to obtain a rooting powder nutrient solution.

3. The breeding method of borneol camphor according to claim 2, characterized in that, The concentration of Lactobacillus brevis in the mixed solution was 10 6 -10 7 CFU / mL.

4. The breeding method of borneol camphor according to claim 2, characterized in that, The concentration of Lactobacillus plantarum in the sterilized material is 10 7 -10 8 CFU / mL.

5. The breeding method of borneol camphor according to claim 2, characterized in that, The mass ratio of potassium α-naphthylacetate, potassium 3-indolebutyrate and baking powder is (0.2-0.5):1:(1.4-1.7).

6. The method for breeding borneol camphor according to claim 1, characterized in that: The growth regulator liquid comprises the following components: 6.5-8.6 mg / L 6-BA, 1.2-3.5 mg / L NAA, 1.3-2.6 mg / L IBA, 2.4-3.5 mg / L IAA, and 2.4-4.6 mg / L plant extract, and is prepared with water.

7. The method for breeding borneol camphor according to claim 6, characterized in that: The preparation method of the plant extract is: (1) Mixing Platycladus orientalis leaves, Gelidium amansii, and Astragalus membranaceus, and crushing them to a particle size of less than 80-100 meshes to obtain a mixed material; (2) subjecting the mixed material to supercritical extraction, firstly extracting at 40-45°C, 30-35MPa, and a CO2 flow rate of 17-20L / h for 100-120min to obtain a first fraction, and then further extracting at 25-30°C, 25-30MPa, and a CO2 flow rate of 17-20L / h for 60-80min to obtain a second fraction; (3) The first fraction and the second fraction are mixed to obtain a plant extract.

8. The method for breeding borneol camphor according to claim 7, characterized in that: The mass ratio of Platycladus orientalis leaves, Gelidium agar-agar and Astragalus membranaceus is 1:(1.3-1.5):(0.3-0.7).

9. The method for breeding borneol camphor according to claim 7, characterized in that: Select tissue culture seedlings that have white spots after being cut for pruning.

10. Use of the borneol camphor breeding method according to any one of claims 1 to 9.

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