CN113317206A - Culture medium combination for tissue culture and rapid propagation of lilium martagon, tissue culture and rapid propagation method and application - Google Patents
Culture medium combination for tissue culture and rapid propagation of lilium martagon, tissue culture and rapid propagation method and application Download PDFInfo
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- CN113317206A CN113317206A CN202110812371.XA CN202110812371A CN113317206A CN 113317206 A CN113317206 A CN 113317206A CN 202110812371 A CN202110812371 A CN 202110812371A CN 113317206 A CN113317206 A CN 113317206A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/002—Culture media for tissue culture
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/10—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material
- A01G24/12—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material containing soil minerals
- A01G24/15—Calcined rock, e.g. perlite, vermiculite or clay aggregates
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/20—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material
- A01G24/22—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material containing plant material
- A01G24/23—Wood, e.g. wood chips or sawdust
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/20—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material
- A01G24/28—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material containing peat, moss or sphagnum
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
Abstract
The invention provides a culture medium combination for tissue culture and rapid propagation of lilium martagon, a tissue culture and rapid propagation method and application, and belongs to the technical field of plant tissue rapid propagation. According to the method, through optimization of each component of the culture medium, the induction culture medium and the differentiation culture medium are combined into one, the proliferation culture medium and the subculture culture medium are combined into one, and the strong seedling culture medium and the rooting culture medium are combined into one, so that the tissue culture steps of the lilium martagon are simplified, the rapid propagation of the lilium martagon is realized, the problems that the lilium martagon has sporadic field distribution, few population quantity and is endangered and extinct are solved, a foundation is laid for comprehensive protection, development and continuous utilization of the lilium martagon and germplasm resources preservation, and the research and development blank of the lilium martagon in biotechnology is filled.
Description
Technical Field
The invention belongs to the technical field of plant tissue rapid propagation, and particularly relates to a culture medium combination for tissue culture rapid propagation of lilium martagon, a tissue culture rapid propagation method and application.
Background
Lilium medogen is a new species of Lilium published in 1985 by Mr. Lijunyun, and the model specimen is a specimen collected from Xizang Xigeda Xiongjiu in 1980 by Chengwei Mr. Chengwei. At present, the lilium martagon is in a situation of being endangered and extincted, is a typical wild plant with a very small population, and has great significance in expanding propagation and protecting. At present, a tissue culture rapid propagation method aiming at the lilium martagon does not exist, so that a tissue culture rapid propagation method aiming at the lilium martagon is urgently needed.
Disclosure of Invention
In order to solve the problems, the invention provides a culture medium combination for tissue culture and rapid propagation of the lilium martagon, a tissue culture and rapid propagation method and application. The culture medium combination for tissue culture and rapid propagation of the lilium martagon can realize rapid propagation of the lilium martagon and lay a foundation for comprehensive protection, development and utilization of the lilium martagon.
In order to achieve the above purpose, the invention provides the following technical scheme:
the invention provides a culture medium combination for tissue culture and rapid propagation of lilium martagon, which comprises an induction and differentiation culture medium, a proliferation and subculture medium and a strong seedling and rooting culture medium;
the induction and differentiation culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: : 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the induction and differentiation culture medium is 5.8-6.4;
the proliferation and subculture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.5 mg/L6-BA, 0.5-1.0 mg/L NAA, 40-50 g/L sucrose and 4.5-5.5g/L agar; the pH value of the proliferation and subculture medium is 5.8-6.4;
the strong seedling and rooting culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the strong seedling and rooting culture medium is 5.8-6.4;
the concentrations of the trace elements in the modified MS medium were as follows: KI 1.66mg/L, H3BO3 12.40mg/L、MnSO4·4H2O 44.6mg/L、ZnSO4·7H2O 17.20mg/L、Na2MoO4·2H2O 0.50mg/L、CuSO4·5H2O0.05 mg/L and CoCl2·6H2O 0.05mg/L。
Preferably, the culture medium also comprises a transplanting culture medium, wherein the transplanting culture medium comprises perlite, humus and raw rosewood; the volume ratio of the perlite to the humus to the raw rosewood is (1: 2): (1-2): (1-2).
The invention provides the application of the culture medium combination in the technical scheme in tissue culture and rapid propagation of the lilium martagon.
The invention provides a tissue culture and rapid propagation method of lilium martagon, which comprises the following steps:
inducing and differentiating the sterilized explant in the inducing and differentiating culture medium in the technical scheme to grow adventitious buds;
carrying out proliferation and subculture on the adventitious bud in the proliferation and subculture medium in the technical scheme to obtain a rootless plantlet;
and (3) performing strong seedling and rooting culture on the non-rooted clump seedling in the strong seedling and rooting culture medium in the technical scheme to obtain the lilium martagon sterile seedling.
Preferably, the species of the explant comprises scales, leaves, petals, pistils or stamens of the lilium nigrum.
Preferably, the method for disinfecting the explant comprises the following steps: soaking the explants for 3-10 min by 10% by volume of Wanjin brand spray disinfectant, and washing the soaked explants for 3 times by using filtered water; sterilizing the washed explant with 75% alcohol for 10s, and washing with sterile water for 3 times; and (3) disinfecting the explant washed by the sterile water for 5-8 min by using 0.1% mercuric chloride solution, and washing 3-5 times by using the sterile water.
Preferably, the culture conditions of the induction and differentiation culture are as follows: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (10-12) L: (12-14) D, culturing for 45-60D;
the culture conditions of proliferation and subculture are as follows: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (10-12) L: (12-14) D, culturing for 45-60D;
the culture conditions of the strong seedling and rooting culture are as follows: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (5-6) L: (18-19) D, and culturing for 45-60 days.
Preferably, transplanting is further included after the sterile seedlings are obtained; the transplanting substrate is the transplanting culture medium in the technical scheme; the culture conditions after transplanting are as follows: the temperature is 23-25 ℃, the relative humidity of air is 40-50%, and the relative humidity of the substrate is 40-50%.
Preferably, before transplanting, the method further comprises hardening the aseptic seedlings; the seedling exercising conditions are as follows: the light shading rate is 70-85%, the temperature is 23-25 ℃, the relative humidity is 40-50%, and the seedling hardening time is 7-10 d.
The invention provides the application of the method in the technical scheme in tissue culture and rapid propagation of the lilium martagon.
Has the advantages that:
the invention provides a culture medium combination for tissue culture and rapid propagation of lilium martagon, which comprises an induction and differentiation culture medium, a proliferation and subculture medium and a strong seedling and rooting culture medium; the induction and differentiation culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.1-1.0 mg/L of 6-BA, 0.1-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the induction and differentiation culture medium is 5.8-6.4; the proliferation and subculture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.1-2.0 mg/L6-BA, 0.5-1.0 mg/L NAA, 40-50 g/L sucrose and 4.5-5.5g/L agar; the pH value of the proliferation and subculture medium is 5.8-6.4; the strong seedling and rooting culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the strong seedling and rooting culture medium is 5.8-6.4. According to the method, through optimization of each component of the culture medium, the induction culture medium and the differentiation culture medium are combined into one, the proliferation culture medium and the subculture culture medium are combined into one, and the strong seedling culture medium and the rooting culture medium are combined into one, so that the tissue culture steps of the lilium martagon are simplified, the rapid propagation of the lilium martagon is realized, the problems that the lilium martagon has sporadic field distribution, few population quantity and is endangered and extinct are solved, a foundation is laid for comprehensive protection, development and continuous utilization of the lilium martagon and germplasm resources preservation, and the research and development blank of the lilium martagon in biotechnology is filled.
Furthermore, the invention provides a tissue culture and rapid propagation method of the lilium pedunculatum, which adopts the culture medium combination in the technical scheme to carry out tissue culture on the lilium pedunculatum. The method provided by the invention has high induced differentiation rate and rooting rate, the breeding period is only 45-60 days, the breeding rate of the lilium martagon is greatly improved, and technical support is provided for the protection and breeding, introduction and domestication, storage and large-scale production of the species.
Drawings
FIG. 1 shows the differentiation of adventitious buds of an indexinized lily bulb after 2 weeks of induction culture;
FIG. 2 shows the growth of adventitious buds of an indexinated lily bulb after 45 days of scale induction culture;
FIG. 3 shows the growth of adventitious buds of a plant of Leucopaxillus 50 days after induced culture;
FIG. 4 shows the growth of the adventitious bud of lilium notatum in the modified MS +1.0 mg/L6-BA +0.5mg/L NAA proliferation sub-passage for 50 d;
FIG. 5 shows the growth of the adventitious bud of lilium notatum after proliferation and subculture for 40d in modified MS +0.5 mg/L6-BA +0.5mg/L NAA;
FIG. 6 shows the growth of the adventitious bud of lilium notatum after proliferation and subculture for 50 days in modified MS +0.5 mg/L6-BA +0.5mg/L NAA;
FIG. 7 shows the growth of sound seedlings and rooting cultures of the lilium martagon (L.) Druce for 50 days;
FIG. 8 is the growth of bulblets after 60 days of incubation of the lilium martagon in rooting medium.
Detailed Description
The invention provides a culture medium combination for tissue culture and rapid propagation of lilium martagon, which comprises an induction and differentiation culture medium, a proliferation and subculture medium and a strong seedling and rooting culture medium;
the induction and differentiation culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the induction and differentiation culture medium is 5.8-6.4;
the proliferation and subculture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.5 mg/L6-BA, 0.5-1.0 mg/L NAA, 40-50 g/L sucrose and 4.5-5.5g/L agar; the pH value of the proliferation and subculture medium is 5.8-6.4;
the strong seedling and rooting culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of NAA, 40-50 g/L of cane sugar and 4.5-5.5g/L of agar; the pH value of the strong seedling and rooting culture medium is 5.8-6.4;
the concentrations of the trace elements in the modified MS medium were as follows: KI 1.66mg/L, H3BO3 12.40mg/L、MnSO4·4H2O 44.6mg/L、ZnSO4·7H2O 17.20mg/L、Na2MoO4·2H2O 0.50mg/L、CuSO4·5H2O0.05 mg/L and CoCl2·6H2O 0.05mg/L。
The induction and differentiation culture medium takes an improved MS culture medium as a basic culture medium. In the present invention, the modified MS medium preferably doubles the trace elements in the MS medium, i.e. the concentration of the trace elements in the modified MS medium of the present invention is preferably as follows: KI 1.66mg/L, H3BO3 12.40mg/L、MnSO4·4H2O 44.6mg/L、ZnSO4·7H2O 17.20mg/L、Na2MoO4·2H2O 0.50mg/L、CuSO4·5H2O0.05 mg/L and CoCl2·6H2O0.05 mg/L; the concentration content of the MS culture medium is continuously adopted by other components except trace elements, sucrose and agar in the improved MS culture medium. The improved MS culture medium is taken as a basic culture medium, which is beneficial to the growth of the lilium martagon, so that the cultivated lilium martagon leaves are stretched and not curled, and the occurrence of glass seedlings is prevented. The induction and differentiation culture medium also comprises 0.5-1.0 mg/L of 6-BA; the concentration of the 6-BA is further preferably 0.5-0.8 mg/L; still more preferably 0.5 mg/L. Induction and differentiation according to the inventionThe culture medium also comprises 0.5-1.0 mg/L NAA; the concentration of the NAA is further preferably 0.5-0.8 mg/L; still more preferably 0.5 mg/L. The induction and differentiation culture medium specific hormone combination can realize the one-step induction of adventitious buds by the indemnated lily explant, the induction culture medium and the differentiation culture medium are combined into a whole, the operation steps are simplified, the adventitious bud induction effect is good, the adventitious bud induction rate can reach 100%, and the average individual number of the induced adventitious buds is 3-5.
The induction and differentiation culture medium also comprises 30-50 g/L of sucrose; the concentration of the sucrose is further preferably 40-50 g/L; further preferably 45-50 g/L; still more preferably 45 g/L. The sucrose with specific concentration can provide a carbon source required by the lilium martagon, promote good plant growth, promote the growth of the bulblet and improve the percentage of the bulblet. The induction and differentiation culture medium also comprises 4.5-5.5g/L agar; the concentration of the agar is more preferably 5-5.5 g/L; more preferably 5g/L, agar in the present invention mainly acts as a coagulant.
The proliferation and subculture medium takes an improved MS culture medium as a basic culture medium. In the present invention, the modified MS medium is preferably the same as the modified MS medium used in the induction and differentiation medium described in the above technical means. The proliferation and subculture medium also comprises 0.5-1.5 mg/L of 6-BA; the concentration of the 6-BA is further preferably 0.5-1.0 mg/L; still more preferably 0.5 mg/L. The proliferation and subculture medium also comprises 0.5-1.0 mg/L NAA; the concentration of the NAA is further preferably 0.5-0.8 mg/L; still more preferably 0.5 mg/L. The specific hormone combination of the proliferation and the subculture medium can realize the integration of the proliferation and the subculture of the lilium martagon, the operation steps are simplified, the leaves of the obtained adventitious bud plants are extended, and the number of the adventitious buds is 5-6. The proliferation and subculture medium also comprises 30-50 g/L of sucrose; the concentration of the sucrose is further preferably 40-50 g/L; further preferably 45-50 g/L; still more preferably 45 g/L. The specific sucrose concentration of the invention is beneficial to promoting the growth of the bulblet, and the whole plant grows well. The proliferation and subculture medium also comprises 4.5-5.5g/L agar; the concentration of the agar is more preferably 5-5.5 g/L; more preferably 5g/L, agar in the present invention mainly acts as a coagulant.
The strong seedling and rooting culture medium takes an improved MS culture medium as a basic culture medium. In the present invention, the modified MS medium is preferably the same as the modified MS medium used in the induction and differentiation medium described in the above technical means. The strong seedling and rooting culture medium also comprises 0.5-1.0 mg/L of 6-BA; the concentration of the 6-BA is further preferably 0.5-0.8 mg/L; still more preferably 0.5 mg/L. The strong seedling and rooting culture medium also comprises 0.5-1.0 mg/L NAA; the concentration of the NAA is further preferably 0.8-1.0 mg/L; still more preferably 1.0 mg/L. The combination of strong seedlings and specific hormones in a rooting culture medium can realize the combination of strong seedlings and rooting culture of the lilium martagon, simplifies the operation steps, and has good rooting and strong seedling effects, the rooting rate can reach 100 percent, and the multiplication multiple of bulblets can reach 7 times. The strong seedling and rooting culture medium also comprises 30-50 g/L of sucrose; the concentration of the sucrose is further preferably 40-50 g/L; more preferably 45 to 50 g/L. The invention is 45g/L of sucrose. The strong seedling and rooting culture medium also comprises 4.5-5.5g/L agar; the concentration of the agar is further preferably 4.8-5.2 g/L; more preferably 5g/L, agar in the present invention mainly acts as a coagulant.
The culture medium composition for tissue culture and rapid propagation of the lilium martagon also preferably comprises a transplanting culture medium. In the present invention, the transplanting medium preferably includes perlite, humus soil and raw rosewood. In the invention, the volume ratio of the perlite to the humus to the raw rosewood is preferably (1-2): (1-2): (1-2); more preferably 1: 2: 1. according to the invention, the proportion of the perlite, the humus soil and the raw red soil is specified, so that the porosity of the matrix and the air permeability of the root are increased, and sufficient nutrition is provided for the root; meanwhile, laterite is soil deep in the soil layer, the bacteria carrying amount is less, the growth of aseptic seedlings is facilitated, and the transplanting survival rate of the lilium martagon can reach 100% by using the transplanting culture medium.
The culture medium combination for tissue culture and rapid propagation of the lilium martagon provided by the invention combines an induction culture medium and a differentiation culture medium into one, combines a proliferation culture medium and a subculture culture medium into one, combines a strong seedling culture medium and a rooting culture medium into one, simplifies the operation steps of tissue culture of the lilium martagon, is easy to grow seedlings, has high effective propagation rate, realizes rapid propagation of the lilium martagon, solves the problems of sporadic field distribution, rare population and imminent extinction of the lilium martagon, lays a foundation for comprehensive protection, development and continuous utilization of the lilium martagon and germplasm resource preservation of the lilium martagon, and fills the blank of research and development of the lilium martagon on biotechnology.
The invention provides the application of the culture medium combination in the technical scheme in tissue culture and rapid propagation of the lilium martagon. The tissue rapid propagation culture of the lilium martagon is carried out by adopting the culture medium combination in the technical scheme, so that the propagation quantity and the growth rate of the lilium martagon are greatly improved, and technical support is provided for the protection and propagation, introduction and domestication, preservation and large-scale production of the species.
The invention provides a tissue culture and rapid propagation method of lilium martagon, which comprises the following steps:
inducing and differentiating the sterilized explant in the inducing and differentiating culture medium in the technical scheme to grow adventitious buds;
carrying out proliferation and subculture on the adventitious bud in the proliferation and subculture medium in the technical scheme to obtain a rootless plantlet;
and (3) performing strong seedling and rooting culture on the non-rooted clump seedling in the strong seedling and rooting culture medium in the technical scheme to obtain the lilium martagon sterile seedling.
The invention preferably sterilizes the explant and then induces and differentiates the explant. In the present invention, the species of the explant preferably includes scales, leaves, petals, pistils or stamens of lilium nigrum; further preferably scales, leaves, pistils or anther-removed stamens; even more preferably scales. In the present invention, the method for sterilizing the explant preferably comprises the steps of: soaking the explant in 10% Wanjin brand spray disinfectant; the soaking time is preferably 3-10 min; further preferably 5-10 min; more preferably 8 min. After soaking, the invention preferably washes the soaked explants with filtered water 3 times. After washing, the invention preferably disinfects washed explants for 10s with 75% alcohol and washes with sterile water for 3 times. After the sterile water washing, the explant after the sterile water washing is sterilized by 0.1% mercuric chloride solution for 5-8 min, preferably 6-7 min, and more preferably 7 min. After the sterilizing by using 0.1% mercuric chloride solution, the invention is preferably washed by using sterile water for 3-5 times; more preferably 4 to 5 times, and still more preferably 5 times. In the present invention, when the explant is a flake, it is also preferable to wash off the soil on the flake with tap water before soaking with 10% Wanjin brand spray disinfectant. When the explant is a leaf, petal, pistil, or anther-removed stamen, the step of washing off the soil is preferably omitted. The explant sterilization method has the aseptic rate of 70%.
After the explant is sterilized, the sterilized explant is induced and differentiated in the induction and differentiation culture medium in the technical scheme to grow adventitious buds. The explant of the invention is preferably inoculated flat in a culture medium. When the explant is a scale, the inoculation amount of the scale is preferably 2-4 scales/100 cm2(ii) a Further preferably 2 flakes/100 cm2(ii) a The scales are preferably inoculated in the culture medium with the concave surfaces facing upwards. When the explant is a leaf or petal, preferably cutting the leaf or petal into small sections with the size of 1-2 cm, and then inoculating; the preferred inoculation amount of the leaves or the petals is 4-5 small sections/100 cm2(ii) a Further preferably 4 pieces/100 cm2. When the explant is pistil or anther-removed stamen, the pistil or anther-removed stamen is preferably transversely cut into 2-3 cm segments, and the inoculation amount is preferably 2-5 segments/100 cm2(ii) a Further preferably 2-3 small sections/100 cm2. In the present invention, the temperature for the induction and differentiation culture is preferably 23 to 25 ℃, and more preferably 25 ℃. In the invention, the illumination intensity of the induction and differentiation culture is preferably 1500-2000 LUX; further preferably 1600-; more preferably 1800 LUX. In the present inventionIn the light, the illumination period is preferably (10-12) L: (12-14) D; more preferably (11-12) L: (12-13) D; more preferably L12: 12D. In the invention, the culture time of the induction and differentiation culture is preferably 45-60 d; further preferably 50 to 60 days; more preferably 50 d. Under the specific induction and differentiation culture conditions, the induction and differentiation adventitious bud of the lilium martagon has good growth vigor.
After induction and differentiation culture, the invention carries out proliferation and subculture on the adventitious bud in the proliferation and subculture medium in the technical scheme to obtain the rootless plantlet. In the present invention, the temperature for propagation and subculture is preferably 23 to 25 ℃, and more preferably 24 to 25 ℃. In the invention, the illumination intensity of the proliferation and subculture is preferably 1500-2000 LUX; further preferably 1600 to 1800 LUX; more preferably 1800 LUX. In the present invention, the photoperiod of the proliferation and subculture is preferably (10 to 12) L: (12-14) D; more preferably (11-12) L: (12-13) D; still more preferably 12L: 12D. In the invention, the culture time of the proliferation and subculture is preferably 45-60 d; further preferably 50 to 60 days; more preferably 50 d. Under the specific conditions of proliferation and subculture, the lily without roots grows well.
After proliferation and subculture, the invention performs strong seedling and rooting culture on the rootless clump seedlings in the strong seedling and rooting culture medium in the technical scheme to obtain the aseptic seedlings of the lilium martagon. In the present invention, the non-rooted plantlet is preferably divided into individual plants, and then strong seedling and rooting culture are performed. In the invention, the temperature of the strong seedlings and the rooting culture is preferably 23-25 ℃, and more preferably 24-25 ℃. In the invention, the illumination intensity of the strong seedling and rooting culture is preferably 1500-2000 LUX; further preferably 1600 to 1800 LUX; more preferably 1800 LUX. In the invention, the sound seedling and rooting are preferably (5-7) L: (17-19) D; more preferably (5-6) L: (18-19) D; more preferably 6L: 18D. In the invention, the culture time of the strong seedling and rooting culture is preferably 45-60 d; further preferably 50 to 60 days; more preferably 50 d. Under the specific conditions of strong seedling and rooting culture, the rooting rate of the lilium martagon is 100%, the percentage of bulblets is as high as 55%, and the lilium martagon grows well.
After strong seedling and rooting culture, the invention preferably performs seedling hardening on the obtained aseptic seedling. The seedling exercising of the invention is preferably to move the aseptic seedlings to a greenhouse for exercising. In the invention, the shading rate of the greenhouse is preferably 70-85%; further preferably 75-80%; still more preferably 75%. In the invention, the temperature of the greenhouse is preferably 23-25 ℃; more preferably 24 deg.c. In the invention, the relative air humidity of the greenhouse is preferably 40-50%; further preferably 40% to 45%; still more preferably 45%. In the invention, the seedling exercising time is preferably 7-10 days; further preferably 7-8 d; even more preferably 8 d. Hardening the seedlings after obtaining the aseptic seedlings, and being beneficial to improving the survival rate after transplanting.
After hardening off, the invention preferably transplants the aseptic seedlings after hardening off. In the present invention, the substrate for transplantation is preferably the transplantation medium described in the above technical solution. The transplanting medium is preferably prepared by spraying 800-1000 times of carbendazim, mixing with soil, sealing and sterilizing with a plastic film for 7 days, adjusting the pH to 6.2-6.4, and then transplanting. In the invention, the culture temperature after transplanting is preferably 23-25 ℃, and more preferably 24-25 ℃. In the invention, the humidity of the matrix after transplanting is preferably 40-50%; further preferably 40% to 45%; still more preferably 45%. The special transplanting conditions of the invention ensure the high survival rate of the transplanted Italian lily rooted seedlings, which can reach 100%.
The tissue culture and rapid propagation method of the lilium martagon provided by the invention has the advantages that the induced differentiation rate and the rooting rate are high, the propagation period is only 45-60 days, the propagation rate of the lilium martagon is greatly improved, and the technical support is provided for the protection and propagation, introduction and domestication, storage and large-scale production of the species.
The invention provides application of the method in the technical scheme in the rapid propagation of the tissue of the lilium martagon.
For further illustration of the present invention, the culture medium combination, the tissue culture rapid propagation method and the application of the present invention are described in detail below with reference to the examples, but they should not be construed as limiting the scope of the present invention.
Example 1 Induction and differentiation media screening
Taking scales on the blackout lily bulbs as explants, washing off soil on the scales with tap water, soaking for 8min with 10% Wanjin brand spray disinfectant, washing for 3 times with filtered water, taking the scales on a super clean bench, disinfecting for 10s with 75% alcohol, washing for 3 times with sterile water, disinfecting for 7min with 0.1% mercuric chloride solution, and washing for 3-5 times with sterile water to obtain disinfected explants for later use.
The sterilized explants were inoculated into the following 5 induction and differentiation media, 1 flake was placed flat per bottle of media, with the concave side facing upward. The 5 kinds of induction and differentiation culture media are respectively: 1) improved MS culture medium (trace element added by 1 time) +0.1 mg/L6-BA +0.1mg/L NAA; 2) improved MS culture medium (trace element added by 1 time) +0.2 mg/L6-BA +0.2mg/L NAA; 3) improved MS culture medium (trace element added by 1 time) +0.5 mg/L6-BA +0.5mg/L NAA; 4) improved MS culture medium (trace elements added by 1 time) +0.8 mg/L6-BA +0.8mg/L NAA; 5) improved MS culture medium (trace element plus 1 time) +1.0 mg/L6-BA +1.0mg/L NAA. The above-mentioned induction and differentiation culture medium also contains 40g/L of cane sugar, 5g/L of agar and pH 6.0. The trace elements in the improved MS culture medium are: KI 1.66mg/L, H3BO3 12.40mg/L、MnSO4·4H2O 44.6mg/L、ZnSO4·7H2O 17.20mg/L、Na2MoO4·2H2O 0.50mg/L、CuSO4·5H2O0.05 mg/L and CoCl2·6H2O0.05 mg/L. The conditions for induction and differentiation culture are as follows: light intensity is 1800LUX, temperature is 25 ℃, and illumination period is 10L: 24D, the culture period is 50D; after two weeks of induction, adventitious buds began to grow out, and the condition of inducing adventitious buds for 50 days was counted. The statistical results are shown in Table 1.
TABLE 1 Induction and differentiation Medium screening
Remarking: in the table "-" indicates no callus; "+" indicates callus less than 0.1 cm; "+ +" indicates callus over 0.1 cm.
The results in Table 1 show that the induction rate of adventitious buds of the lilium notatum is 100% when the lilium notatum adventitious buds are induced on an improved MS + 0.5-1.0 mg/L6-BA + 0.5-1.0 mg/L NAA culture medium, and the average individual number of the induced adventitious buds is 3-5; wherein the induction effect of improved MS +0.5 mg/L6-BA +0.5mg/L NAA is the best, and the induction conditions are shown in figures 1-3.
Example 2 proliferation and subculture Medium screening
The modified MS (same as the modified MS culture medium in example 1) culture medium is used as a basic culture medium for hormone screening of the proliferation and subculture medium, 5 gradients of 0.1mg/L, 0.5mg/L, 1.0mg/L, 1.5mg/L and 2.0mg/L are set for cytokinin 6-BA, and 2 gradients of 0.5mg/L and 1.0mg/L are set for auxin NAA, and the uniform design method is adopted to screen the appropriate proliferation and subculture medium. The multiplication and subculture medium also contains 45g/L of sucrose, 5g/L of agar and pH 6.0. The conditions for proliferation and subculture were: light intensity is 1800LUX, temperature is 25 ℃, and illumination period is 10L: 24D, the culture period is 50D. The statistical results are shown in Table 2.
TABLE 2 proliferation and subculture medium screening
Remarking: "-" indicates no callus; "+" indicates callus less than 0.1 cm; "+ +" indicates callus over 0.1 cm.
As is clear from the results shown in Table 2, when the proliferation and subculture medium is an improved MS medium (trace elements added by 1 time) + 0.5-1.5 mg/L6-BA + 0.5-1.0 mg/L NAA (naphthylacetic acid), the number of adventitious buds is 5-6; among them, the improved MS +0.5 mg/L6-BA +0.5mg/L NAA had the best effect in cultivation, leaves were extended, and adventitious buds were plump, as shown in FIGS. 4 to 6.
Example 3 Strong shoot and rooting Medium screening
An improved MS (same as the improved MS culture medium in example 1) culture medium is used as a basic culture medium for hormone screening of the strong seedling and rooting culture medium, 2 gradients of 0.5mg/L and 1.0mg/L of cytokinin 6-BA and 2 gradients of 0.5mg/L and 1.0mg/L of auxin NAA are set, and the proper strong seedling and rooting culture medium is screened. The strong seedlings and the rooting culture medium also contain 45g/L of sucrose, 5g/L of agar and pH 6.0. The conditions of strong seedling and rooting culture are as follows: the light intensity is 1500-: 19D, the culture period is 50D. The induction of adventitious buds for 50 days was counted. The statistical results are shown in Table 3.
TABLE 3 Strong seedling and rooting Medium screening
Remarking: "-" indicates no callus; "+" indicates callus less than 0.1 cm; "+ +" indicates callus over 0.1 cm.
From the results in table 3, it can be seen that when the strong seedling and rooting medium is an improved MS medium (trace elements added by 1 time) + 0.5-1.0 mg/L6-BA + 0.5-1.0 mg/L NAA (naphthylacetic acid), the multiplication of bulblet is 4-7, and the rooting rate reaches 100%; wherein, the strong seedling and rooting effect of the improved MS +0.5 mg/L6-BA +1.0mg/L NAA is the best, as shown in figure 7 and figure 8.
Example 4 transplantation Medium screening
The rooting aseptic seedlings of the lilium martagon which are obtained by improving the MS +0.5 mg/L6-BA +1.0mg/L NAA culture medium in the embodiment 3 are transferred to a greenhouse with the shading rate of 75-85% for hardening seedlings one week in advance. Transplanting the rooted aseptic seedling after hardening seedling into different transplanting culture media, and screening the proper transplanting culture media. The experimental transplanting culture media are respectively as follows: raw red soil, perlite, a humus soil/raw red soil mixed matrix and a perlite/humus soil/raw red soil mixed matrix, wherein the volume ratio of humus soil to raw red soil in the humus soil/raw red soil mixed matrix is 1: 1, the volume ratio of the perlite to the humus soil to the raw laterite in the perlite/humus soil/raw laterite mixed matrix is as follows: 1: 2: 1. spraying 800-1000 times of carbendazim on the transplanting culture medium, mixing with soil, sealing and disinfecting with a plastic film for 7 days, adjusting the pH value to 6.2-6.4, and then transplanting. And spraying water and covering a plastic film in time after transplanting, wherein the temperature of the greenhouse is 24-25 ℃, and the humidity of the matrix is 45%. The number of transplants per treatment was 20, and the experiment was repeated 3 times. And after 30d of transplanting, counting the transplanting survival rate of each treatment. The results are shown in Table 4.
TABLE 4 selection of transplanting Medium
From the results in table 4, it can be seen that the mixed matrix of perlite, humus and raw red soil has the highest transplanting survival rate, and the survival rate after 30d transplanting reaches 100%.
Example 5 tissue culture Using leaves as explants
Explant disinfection: taking the sepiolite lily leaves as explants, soaking the sepiolite lily leaves in 10% Wanjin brand spray disinfectant for 5-10 min, washing the sepiolite lily leaves for 3 times by filtering water, taking the sepiolite lily leaves on a super clean bench, disinfecting the sepiolite lily leaves for 10s by using 75% alcohol, washing the sepiolite lily leaves for 3 times by using sterile water, disinfecting the sepiolite lily leaves for 5-8 min by using 0.1% mercuric chloride solution, and washing the sepiolite leaves for 3-5 times by using sterile water to obtain disinfected leaves for later use.
Cutting the disinfected leaves into small sections with the size of 1-2 cm, and inoculating the small sections to an induction and differentiation culture medium, wherein the induction and differentiation culture medium is as follows: modified MS +0.5 mg/L6-BA +0.5mg/L NAA. The conditions for induction and differentiation culture were the same as in example 1. After 2 weeks, adventitious buds were induced, the rate of induction of adventitious buds was 25%, and the average number of adventitious buds induced was 2.
Inoculating the induced adventitious bud into a proliferation and subculture medium for proliferation and subculture. Wherein the proliferation and subculture medium is: modified MS +0.5 mg/L6-BA +0.5mg/L NAA. The conditions for proliferation and subculture were the same as in example 2. After 50 days of culture, rootless clump seedlings are obtained, the leaves of the rootless clump seedlings are stretched, and the number of adventitious buds is 6.
Dividing the rootless seedlings generated after subculture into single plants, transferring the single plants to a strong seedling and rooting culture medium for strong seedling and rooting culture, wherein the strong seedling and rooting culture medium comprises: MS +0.5 mg/L6-BA +1.0mg/L NAA. The conditions for strong seedling and rooting culture were the same as in example 3. And culturing for 50 days to obtain the sterile complete small plant seedlings of the lilium martagon, wherein the number of leaves is 2-4, and the rooting rate is 100%.
And moving the obtained aseptic seedlings of the lilium martagon to a greenhouse with a shading rate of 75-85% for hardening seedlings one week in advance. Transplanting the rooted aseptic seedlings after hardening into a transplanting culture medium, wherein the transplanting matrix is a perlite/humus soil/raw laterite mixed matrix, and the volume ratio of the perlite to the humus soil to the raw laterite is as follows: 1: 2: 1. spraying 800-1000 times of carbendazim on the transplanting culture medium, mixing with soil, sealing and disinfecting with a plastic film for 7 days, adjusting the pH value to 6.2-6.4, and then transplanting. And spraying water and covering a plastic film in time after transplanting, wherein the temperature of the greenhouse is 24-25 ℃, and the humidity of the matrix is 45%. After 30 days of transplanting, the transplanting survival rate is 100 percent.
Example 6 tissue culture Using pistils as explants
Explant disinfection: taking the pistil of the lilium martagon (which is a denudation lily) as an explant, soaking the pistil in 10% Wanjin spray disinfectant for 5-10 min, filtering and washing for 3 times, taking the pistil on a super clean bench, disinfecting the pistil for 10s by using 75% alcohol, washing for 3 times by using sterile water, disinfecting the pistil for 5-8 min by using 0.1% mercuric chloride solution, and washing for 3-5 times by using sterile water to obtain the disinfected pistil for later use.
Cutting the disinfected pistils into small sections of 2-3 cm, and inoculating the small sections to an induction and differentiation culture medium, wherein the induction and differentiation culture medium comprises: modified MS +0.5 mg/L6-BA +0.5mg/L NAA. The conditions for induction and differentiation culture were the same as in example 1. After 2 weeks, adventitious buds were induced, the rate of induction of adventitious buds was 20%, and the average number of adventitious buds induced was 1.5.
Inoculating the induced adventitious bud into a proliferation and subculture medium for proliferation and subculture. Wherein the proliferation and subculture medium is: modified MS +0.5 mg/L6-BA +0.5mg/L NAA. The conditions for proliferation and subculture were the same as in example 2. After 50 days of culture, rootless clump seedlings are obtained, the leaves of the rootless clump seedlings are stretched, and the number of adventitious buds is 6.
Dividing the rootless seedlings generated after subculture into single plants, transferring the single plants to a strong seedling and rooting culture medium for strong seedling and rooting culture, wherein the strong seedling and rooting culture medium comprises: MS +0.5 mg/L6-BA +1.0mg/L NAA. The conditions for strong seedling and rooting culture were the same as in example 3. And culturing for 50 days to obtain the sterile complete small plant seedlings of the lilium martagon, wherein the number of leaves is 2-4, and the rooting rate is 100%.
Transferring the obtained aseptic seedlings of the lilium martagon to a greenhouse with a shading rate of 75-85% for hardening seedlings one week before transplanting. Transplanting the rooted aseptic seedlings after hardening into a transplanting culture medium, wherein the transplanting matrix is a perlite/humus soil/raw laterite mixed matrix, and the volume ratio of the perlite to the humus soil to the raw laterite is as follows: 1: 2: 1. spraying 800-1000 times of carbendazim on the transplanting culture medium, mixing with soil, sealing and disinfecting with a plastic film for 7 days, adjusting the pH value to 6.2-6.4, and then transplanting. And spraying water and covering a plastic film in time after transplanting, wherein the temperature of the greenhouse is 24-25 ℃, and the humidity of the matrix is 45%. After 30 days of transplanting, the transplanting survival rate is 100 percent.
Example 7 Effect of species of basal Medium on the growth of lilium martagon
The influence of the medium based on the modified MS medium and the MS medium on the growth of the lilium martagon was examined. Wherein, the other components of the induction and differentiation culture medium are 0.5mg/L of 6-BA, 0.5mg/L of NAA, 45g/L of sucrose and 5g/L of agar; the other components of the proliferation and subculture medium are 0.5 mg/L6-BA +0.5mg/L NAA +40g/L sucrose +5g/L agar; the other components of the strong seedling and rooting culture medium are 0.5mg/L of 6-BA, 1.0mg/L of NAA, 40g/L of cane sugar and 5g/L of agar. The culture conditions in each stage were the same as those in the culture stages corresponding to examples 1 to 3. After 50 days of culture, the growth of the lilium notatum was examined. The results are shown in Table 1.
TABLE 5 Effect of trace elements in the Medium on the growth of Sophia Lily
The results in table 5 show that the leaves of the aseptic seedlings of the lilium martagon obtained by using the MS culture medium as the basic culture medium are curled, the incidence rate of the glass seedlings is 6-7%, and the leaves of the glass seedlings are water-immersed, translucent or transparent, which affects the growth of the offspring; the improved MS culture medium is taken as a basic culture medium, which is beneficial to the growth of the lilium martagon, so that the cultivated lilium martagon leaves are stretched, and the occurrence of 'glass seedlings' is prevented.
Example 8 Effect of sucrose in Medium on the growth of Lilium Oxycomum
The effect on the growth of the lilium notatum when the concentration of sucrose in the culture medium is 30g/L, 35g/L, 40g/L, 45g/L and 50g/L respectively. The induction and differentiation culture medium is modified MS +0.5 mg/L6-BA +0.5mg/L NAA +5g/L agar. The proliferation and subculture medium is modified MS +0.5 mg/L6-BA +0.5mg/L NAA +5g/L agar; the strong seedling and rooting culture medium is modified MS +0.5 mg/L6-BA +1.0mg/L NAA +5g/L agar. The culture conditions in each stage were the same as those in the culture stages corresponding to examples 1 to 3. And after strong seedling and rooting culture for 60 days, counting the number and size of bulbs at the base part of the lilium martagon and the growth condition of seedlings. The results are shown in Table 6.
TABLE 6 Effect of sucrose in the Medium on the growth of Sophia oblata
Remarking: the bulblet percentage is the percentage of the number of the seedlings which are strong and generate bulblets after rooting for 50-60 days to the number of the inoculated seedlings.
The results in Table 6 show that the concentration of sucrose is 40-50 g/L, the pinocytose lily plantlets are good, and the percentage of bulblet is 25-55%; the concentration of sucrose is 45-50g/L, the small plant of the lilium martagon is good, and the percentage of bulblet is 50-55%; the concentration of sucrose is 45g/L, the pinocystan lily plantlet is good, and the percentage of bulblet is 55 percent the highest.
The results of the above embodiments show that the culture medium combination for tissue culture and rapid propagation of lilium martagon provided by the invention simplifies the operation steps of tissue culture of lilium martagon, has high induction rate and rooting rate, is easy to grow seedlings, effectively improves the propagation efficiency of lilium martagon, lays a foundation for comprehensive protection, development and continuous utilization of lilium martagon and germplasm resource preservation of lilium martagon, and fills up the blank of research and development of lilium martagon on biotechnology.
Although the present invention has been described in detail with reference to the above embodiments, it is only a part of the embodiments of the present invention, not all of the embodiments, and other embodiments can be obtained without inventive step according to the embodiments, and the embodiments are within the scope of the present invention.
Claims (10)
1. A culture medium combination for tissue culture and rapid propagation of the lilium martagon is characterized by comprising an induction and differentiation culture medium, a proliferation and subculture medium and a strong seedling and rooting culture medium;
the induction and differentiation culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of LNAA, 40-50 g/L of sucrose and 4.5-5.5g/L of agar; the pH value of the induction and differentiation culture medium is 5.8-6.4;
the proliferation and subculture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.5 mg/L6-BA, 0.5-1.0 mg/LNAA, 40-50 g/L sucrose and 4.5-5.5g/L agar; the pH value of the proliferation and subculture medium is 5.8-6.4;
the strong seedling and rooting culture medium takes an improved MS culture medium as a basic culture medium, and also comprises the following components in concentration: 0.5-1.0 mg/L of 6-BA, 0.5-1.0 mg/L of LNAA, 40-50 g/L of sucrose and 4.5-5.5g/L of agar; the pH value of the strong seedling and rooting culture medium is 5.8-6.4;
the concentrations of the trace elements in the modified MS medium were as follows: KI 1.66mg/L, H3BO312.40mg/L、MnSO4·4H2O 44.6mg/L、ZnSO4·7H2O 17.20mg/L、Na2MoO4·2H2O 0.50mg/L、CuSO4·5H2O0.05 mg/L and CoCl2·6H2O 0.05mg/L。
2. The culture medium combination of claim 1, further comprising a transplant culture medium comprising perlite, humus, and raw rosewood; the volume ratio of the perlite to the humus to the raw rosewood is (1: 2): (1-2): (1-2).
3. The use of the culture medium combination of claim 1 or 2 in tissue culture and rapid propagation of lilium martagon.
4. A tissue culture and rapid propagation method of lilium martagon, which is characterized by comprising the following steps:
inducing and differentiating the sterilized explant in the inducing and differentiating culture medium of claim 1 or 2 to grow adventitious buds;
proliferating and subculturing the adventitious bud in the proliferation and subculturing medium according to claim 1 or 2 to obtain a rootless plantlet;
and (3) performing strong seedling and rooting culture on the rootless clump seedlings in the strong seedling and rooting culture medium of claim 1 or 2 to obtain the aseptic seedlings of the lilium martagon.
5. The method of claim 4, wherein the species of the explant comprises scales, leaves, petals, pistils or stamens of lilium nigrum.
6. The method of claim 4, wherein the method of sterilizing the explant comprises the steps of: soaking the explants for 3-10 min by 10% by volume of Wanjin brand spray disinfectant, and washing the soaked explants for 3 times by using filtered water; sterilizing the washed explant with 75% alcohol for 10s, and washing with sterile water for 3 times; and (3) disinfecting the explant washed by the sterile water for 5-8 min by using 0.1% mercuric chloride solution, and washing 3-5 times by using the sterile water.
7. The method of claim 4, wherein the inducing and differentiating culture is performed under the following culture conditions: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (10-12) L: (12-14) D, culturing for 45-60D;
the culture conditions of proliferation and subculture are as follows: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (10-12) L: (12-14) D, culturing for 45-60D;
the culture conditions of the strong seedling and rooting culture are as follows: the temperature is 23-25 ℃, the illumination intensity is 1500-2000LUX, and the illumination period (5-6) L: (18-19) D, and culturing for 45-60 days.
8. The method of claim 4, further comprising transplanting after obtaining sterile seedlings; the transplanted substrate is the transplanting culture medium of claim 2; the culture conditions after transplanting are as follows: the temperature is 23-25 ℃, the relative humidity of air is 40-50%, and the relative humidity of the substrate is 40-50%.
9. The method of claim 8, further comprising acclimatizing the sterile plantlets prior to said transplanting; the seedling exercising conditions are as follows: the light shading rate is 70-85%, the temperature is 23-25 ℃, the relative humidity is 40-50%, and the seedling hardening time is 7-10 d.
10. Use of the method of any one of claims 4 to 9 in tissue culture and rapid propagation of lilium martagon.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5403736A (en) * | 1987-03-06 | 1995-04-04 | Mitsui Petrochemical Industries, Ltd. | Method of inducing formation of adventitious bud |
| CN102210266A (en) * | 2011-03-24 | 2011-10-12 | 内蒙古和信园蒙草抗旱绿化股份有限公司 | Culture medium for culturing lilium pumilum tissues |
| CN104604676A (en) * | 2014-12-30 | 2015-05-13 | 内蒙古和信园蒙草抗旱绿化股份有限公司 | Culture media for tissue culture of red lily |
| CN104920228A (en) * | 2015-07-17 | 2015-09-23 | 中国科学院昆明植物研究所 | Lilium duchartrei franch tissue culture rapid propagation and separation preservation method |
| CN110195077A (en) * | 2019-06-14 | 2019-09-03 | 长江师范学院 | A method of building Lilium lancifo1ium Thunb efficient genetic trasformation system |
| CN112616663A (en) * | 2020-12-18 | 2021-04-09 | 湖北顗泉种业生物技术有限公司 | Method for greatly shortening planting period of lilium davidii var davidii and rapidly propagating seedlings |
-
2021
- 2021-07-19 CN CN202110812371.XA patent/CN113317206B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5403736A (en) * | 1987-03-06 | 1995-04-04 | Mitsui Petrochemical Industries, Ltd. | Method of inducing formation of adventitious bud |
| CN102210266A (en) * | 2011-03-24 | 2011-10-12 | 内蒙古和信园蒙草抗旱绿化股份有限公司 | Culture medium for culturing lilium pumilum tissues |
| CN104604676A (en) * | 2014-12-30 | 2015-05-13 | 内蒙古和信园蒙草抗旱绿化股份有限公司 | Culture media for tissue culture of red lily |
| CN104920228A (en) * | 2015-07-17 | 2015-09-23 | 中国科学院昆明植物研究所 | Lilium duchartrei franch tissue culture rapid propagation and separation preservation method |
| CN110195077A (en) * | 2019-06-14 | 2019-09-03 | 长江师范学院 | A method of building Lilium lancifo1ium Thunb efficient genetic trasformation system |
| CN112616663A (en) * | 2020-12-18 | 2021-04-09 | 湖北顗泉种业生物技术有限公司 | Method for greatly shortening planting period of lilium davidii var davidii and rapidly propagating seedlings |
Non-Patent Citations (1)
| Title |
|---|
| 周欢等: ""百合科植物组织培养的研究进展"", 《湖北农业科学》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116391622A (en) * | 2023-04-19 | 2023-07-07 | 南京农业大学 | Lily tissue culture rapid propagation method and application thereof |
| CN116391622B (en) * | 2023-04-19 | 2024-04-30 | 南京农业大学 | Lily tissue culture rapid propagation method and application thereof |
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