CN110604049B - Wild-returning ecological planting method for dendrobium officinale - Google Patents
Wild-returning ecological planting method for dendrobium officinale Download PDFInfo
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Abstract
The invention discloses a wild-returning ecological planting method for dendrobium officinale, which comprises the steps of selecting mature wild or stone-imitated wild dendrobium officinale capsules; culturing seeds of capsules into dendrobium candidum protocorms, taking the dendrobium candidum protocorms, culturing in a protocorm proliferation culture medium, cutting off buds when the number of proliferated buds reaches 70-95%, transferring the cut buds to a rooting culture medium, and culturing for 25-30 days under the condition of specific temperature to prepare for starting domestication of seedlings; culturing the cultured seedlings in a greenhouse for 6-10 months to obtain dendrobium officinale plants; selecting a mountain landform area, and putting dendrobium officinale plants into gaps between stones or nailing roots into the stones; watering, cultivating for 3-5 years, and collecting. The dendrobium officinale obtained by the wild-returning ecological planting method disclosed by the invention almost completely simulates the ecological environment planting of the wild dendrobium officinale, the polysaccharide content of the obtained dendrobium officinale can reach 29.8%, and the dendrobium officinale has a better anti-tumor effect on the pharmacological effect and is far stronger than that of the dendrobium officinale planted in a greenhouse.
Description
Technical Field
The invention relates to the technical field of dendrobium officinale, in particular to a wild-returning ecological planting method of dendrobium officinale.
Background
The Dendrobium officinale is Orchidaceae plant (Dendrobium officinalekimura et Migo), which is recorded in pharmacopoeia and has the effects of benefiting stomach, promoting the production of body fluid, nourishing yin and clearing heat, and modern pharmacological research also shows that the dried stem has various effects of resisting tumor, enhancing immunity, resisting oxidation, reducing blood fat and the like. Dendrobium officinale is a traditional rare Chinese medicinal material, but due to the unique growth environment requirements and the mining in all generations, the wild resources of Dendrobium officinale are becoming rare and endangering extinction.
At present, the dendrobium officinale is mostly planted in an artificial greenhouse or attached to a tree, but the artificial greenhouse or attached tree is mostly fertilized, and is sprayed with pesticide or used with bactericide, so that economic benefit is emphasized too much, and the dendrobium officinale is harvested in 1-2 years, so that the problems of poor quality such as poor quality, low nutritional value and product added value and the like are solved. At present, domestic literatures and patent reports about ecological planting of dendrobium officinale are reported, or artificial means is still applied for fertilizing or bactericides are used, only the dendrobium officinale is transferred from a greenhouse to be planted on stones, and the dendrobium officinale is managed by a greenhouse planting method. For example, chinese patent CN 107624537 a discloses "a wild-simulated planting method of dendrobium officinale", which requires fertilization. Chinese patent CN 106305387A discloses a method for cultivating imitated wild dendrobium nobile by sticking stone, which uses a bactericide. Undoubtedly, the use of fertilizers and bactericides increases the content of harmful elements or heavy metals in dendrobium officinale.
With the improvement of living standard of people, the quality requirement of the dendrobium officinale will be higher and higher, so that the development of a natural pollution-free method for completely returning the dendrobium officinale to wild ecological planting with high nutritional ingredients and good pharmacological effects is an urgent problem to be solved.
The above background disclosure is only for the purpose of assisting understanding of the inventive concept and technical solutions of the present invention, and does not necessarily belong to the prior art of the present patent application, and should not be used for evaluating the novelty and inventive step of the present application in the case that there is no clear evidence that the above content is disclosed at the filing date of the present patent application.
Disclosure of Invention
Aiming at the problems, the invention provides a wild-returning ecological planting method for dendrobium officinale. The dendrobium officinale product planted by the method almost completely simulates the ecological environment planting of wild dendrobium officinale, does not apply chemical fertilizer, spray pesticide and degerming agent, is low in cost, natural, pollution-free, green and healthy, and is easy to popularize on a large scale.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
a wild-returning ecological planting method for dendrobium officinale comprises the following steps:
(1) seed selection: selecting mature wild or stone-like wild dendrobium officinale capsules;
(2) culturing: taking dendrobium officinale capsules, carrying out tissue culture to obtain protocorms, culturing in a protocorm proliferation culture medium, cutting buds which grow to 1-2cm in height and have 2-3 leaves when the number of proliferated buds reaches 70% -95%, transferring the cut buds to a rooting culture medium, culturing for 25-30 days under a specific temperature condition, and preparing for domesticating seedlings after 2-3 white roots with the length of 1cm appear at the base parts; the protocorm proliferation culture medium: pH =5.8, MS +10% -30% potato +1% -5% sucrose +0.5% -1.2% agar; the rooting medium comprises the following components in parts by weight: pH =5.4-5.6, MS +8% -20% banana +2.0mg/LNAA +2% -8% sucrose +0.4% -1.2% agar;
(3) domesticating seedlings: culturing the cultured seedlings in a greenhouse for 6-10 months to obtain dendrobium officinale plants;
(4) planting: selecting a mountain landform area with the altitude of 50-600m, the relative humidity of 55-70% and the annual rainfall of 1000-1500mm, dividing the dendrobium officinale plants into a cluster according to 3-5 plants, and then putting the dendrobium officinale plants into gaps between stones or nailing roots into the stones; watering, cultivating for 3-5 years, and collecting.
Further, the stone is one or a combination of several of rubble, sandstone, volcanic, granite or limestone. Preferably conglomerate or sandstone.
Further, the specific temperature of the step (2) is 20-35 ℃ day temperature and 15-30 ℃ night temperature.
Further, watering in the step (4) is to water for 1-2 times every day within 5-7 days after cultivation; after 5-7 days, 1-2 days, pouring 1 time of water.
Further, the spacing distance between the two clusters of dendrobium officinale plants is 5-10 cm.
Further, the formula of the protocorm proliferation culture medium is as follows: pH =5.8, MS +20% potato +3% sucrose +0.8% agar.
Further, the formula of the rooting medium is as follows: pH =5.4-5.6, MS +10% banana +2.0mg/LNAA +3% sucrose +0.8% agar.
Further, the specific method for obtaining the protocorm by tissue culture comprises the following steps: selecting mature capsules, performing disinfection and sterilization treatment on the mature capsules on a sterile super clean bench, soaking the mature capsules for 20-30s by using 70% alcohol, then washing the mature capsules for 2-3 times by using sterile water, cutting a 2-3cm mouth at the port of the disinfected capsule by using a scalpel, then hanging the capsule with the opening facing downwards from the mouth of an original bulb induction culture medium, lightly tapping to uniformly spread seeds in the capsule in the original bulb induction culture medium, sowing 3-5 capsules for one capsule, covering a bottle cap after sowing, placing the capsule in a germination culture chamber, performing dark culture for one week, and then transferring the capsule into a normal illumination culture chamber for culture for 45-55 days to obtain the original bulb.
Further, the formula of the protocorm induction culture medium is as follows: pH =5.8, MS +1.0mg-3.0mg/L6-BA +2-4% sucrose +0.4% -1.2% agar.
Compared with the prior art, the invention has the advantages and beneficial effects that:
1. the dendrobium officinale product obtained by the wild-returning ecological planting method almost completely simulates the ecological environment planting of wild dendrobium officinale, does not apply chemical fertilizer, spray pesticide and degerming agent, has low cost, is natural and pollution-free, is green and healthy, and is easy to popularize on a large scale.
2. The quality of the dendrobium officinale product obtained by returning to wild ecological planting is close to wild, and compared with the dendrobium officinale product planted in a greenhouse and other modes, the dendrobium officinale product has the advantages of obvious rust color of stems and leaves, transparent stems and stalks, good quality and the like.
3. Compared with the dendrobium officinale planted in a greenhouse or other manners, the dendrobium officinale product obtained by the planting method has the defects of high moisture content, high fiber dregs and the like, and has the characteristics of high polysaccharide content, fragrant taste, tooth adhesion when being chewed and less fiber.
4. According to the dendrobium officinale product obtained by returning to wild ecological planting, the content of polysaccharide in the dendrobium officinale can reach 29.8%, and the dendrobium officinale product has a better anti-tumor effect on the pharmacological effect and is far stronger than 14.9% of the dendrobium officinale planted in a greenhouse.
Drawings
FIG. 1 shows the annual morphology of Dendrobium officinale planted in greenhouse and returning to wild ecological planting.
FIG. 2 shows the three-year-old morphology comparison between the wild-returning ecological planting and the greenhouse planting of Dendrobium officinale.
FIG. 3 shows the overall ecological environment of the wild-returning ecological planting and greenhouse planting of Dendrobium officinale.
FIG. 4 shows the comparison of the blooming states of the dendrobium officinale planted in the greenhouse in the wild-returning ecological planting mode.
Wherein, A in the attached drawing is the returning wild ecological planting of the invention, and B in the attached drawing is greenhouse planting.
DETAILED DESCRIPTION OF EMBODIMENT (S) OF INVENTION
The present invention will be described in further detail with reference to specific embodiments. It should be emphasized that the following description is merely exemplary in nature and is not intended to limit the scope of the invention or its application.
Example 1
A wild-returning ecological planting method for dendrobium officinale comprises the following steps:
(1) seed selection: selecting mature wild Zhejiang dendrobium officinale seeds;
(2) culturing: and (3) sterilizing the capsule selected in the step (1) in a sterile super clean bench, soaking the capsule in 70% alcohol for 30s, and then flushing the capsule with sterile water for 3 times. Cutting a 2cm mouth at the disinfected capsule pod port by using a scalpel, then hanging the capsule with the opening downwards at the mouth of a protocorm induction culture medium, wherein the formula of the protocorm induction culture medium comprises the following components: pH =5.8, MS +1.0mg/L6-BA +2% sucrose +0.4% agar. Lightly beating to make the seeds in the capsule uniformly sprinkle in the protocorm induction culture medium. 5 bottles of capsule seeds. After sowing, the seedlings are covered with bottle caps and placed in a germination culture room for dark culture for one week, and then the seedlings are transferred to a normal illumination culture room for culture for 50 days. Thus obtaining the protocorm. Culturing in an protocorm proliferation culture medium, cutting off buds which grow to 1-2cm in height and have 2-3 leaves when the number of the proliferated buds reaches 70%, transferring the cut buds to a rooting culture medium, culturing at 27 ℃ in the daytime and 23 ℃ at night for 30 days, and allowing 2-3 white roots with the length of 1cm to appear at the base part to prepare for domesticating seedlings; the protocorm proliferation culture medium: pH =5.8, MS +10% potato +1% sucrose +0.5% agar; the rooting medium comprises the following components in parts by weight: pH =5.4, MS +8% banana +2.0mg/LNAA +2% sucrose +0.4% agar;
(3) domesticating seedlings: culturing the cultured seedlings in a greenhouse for 6 months to obtain dendrobium officinale plants;
(4) planting: selecting a mountain landform area with the altitude of 300m, the relative humidity of 55% and the annual rainfall of 1000mm, dividing dendrobium officinale plants into a group according to 3-5 plants, then placing the dendrobium officinale plants into gaps between volcanic landform stones or nailing roots into the stones, wherein the spacing distance between the two groups of dendrobium officinale plants is 8 cm; watering for 1 time every day within 7 days after cultivation; after 7 days, watering 1 time for 2 days, cultivating for 3 years, and harvesting.
Example 2
A wild-returning ecological planting method for dendrobium officinale comprises the following steps:
(1) seed selection: selecting mature Guangxi imitating wild dendrobium officinale capsules on Shi;
(2) culturing: taking the capsule selected in (1), sterilizing in a sterile super clean bench, soaking in 70% ethanol for 20s, and rinsing with sterile water for 3 times. Cutting a 3cm mouth at the disinfected capsule pod port by using a scalpel, then hanging the capsule with the opening downwards at the mouth of a protocorm induction culture medium, wherein the formula of the protocorm induction culture medium comprises the following components: pH =5.8, MS +3.0mg/L6-BA +4% sucrose +1.2% agar. Lightly beating to make the seeds in the capsule uniformly sprinkle in the protocorm induction culture medium. One capsule is sowed in 3 bottles. After sowing, the seedlings are covered with bottle caps and placed in a germination culture room for dark culture for one week, and then the seedlings are transferred to a normal illumination culture room for culture for 55 days. Thus obtaining the protocorm. Culturing in an protocorm proliferation culture medium, cutting off buds which grow to 1-2cm in height and have 2-3 leaves when the number of the proliferated buds reaches 80%, transferring the cut buds to a rooting culture medium, culturing at 28 ℃ in the daytime and 25 ℃ at night for 25 days, and allowing 2-3 white roots with the length of 1cm to appear at the base part to prepare for domesticating seedlings; the protocorm proliferation culture medium: pH =5.8, MS +30% potato +5% sucrose +1.2% agar; the rooting medium comprises the following components in parts by weight: pH =5.6, MS +20% banana +2.0mg/LNAA +8% sucrose +1.2% agar;
(3) domesticating seedlings: culturing the cultured seedlings in a greenhouse for 8 months to obtain dendrobium officinale plants;
(4) planting: selecting a mountain landform area with the altitude of 600m, the relative humidity of 65% and the annual rainfall of 1500mm, dividing dendrobium officinale plants into a group according to 3-5 plants, then nailing the roots into the stone or gaps between limestone landform stones, wherein the spacing distance between the two groups of dendrobium officinale plants is 10 cm; watering 2 times a day within 6 days after cultivation; after 6 days, 1 day of watering 1 time, cultivating for 4 years, and harvesting.
Example 3
A wild-returning ecological planting method for dendrobium officinale comprises the following steps:
(1) seed selection: selecting mature wild Danxia dendrobium officinale capsules;
(2) culturing: selecting the capsule selected in (1), sterilizing in a sterile super clean bench, soaking in 70% ethanol for 30s, and rinsing with sterile water for 3 times. Cutting a 3cm mouth at the disinfected capsule pod port by using a scalpel, then hanging the capsule with the opening downwards at the mouth of a protocorm induction culture medium, wherein the formula of the protocorm induction culture medium comprises the following components: pH =5.8, MS +2.0mg/L6-BA +3% sucrose +0.8% agar. Lightly beating to make the seeds in the capsule uniformly sprinkle in the protocorm induction culture medium. 4 bottles of capsules are sown. After sowing, the seedlings are covered with bottle caps and placed in a germination culture room for dark culture for one week, and then the seedlings are transferred to a normal illumination culture room for culture for 50 days. Thus obtaining the protocorm. Culturing in an protocorm proliferation culture medium, cutting off buds which grow to 1-2cm in height and have 2-3 leaves when the number of the proliferated buds reaches 95%, transferring the cut buds to a rooting culture medium, culturing at 28 ℃ in the daytime and 25 ℃ at night for 25 days, and allowing 2-3 white roots with the length of 1cm to appear at the base part to prepare for domesticating seedlings; the protocorm proliferation culture medium: pH =5.8, MS +20% potato +3% sucrose +0.8% agar; the rooting medium comprises the following components in parts by weight: pH =5.4, MS +10% banana +2.0mg/LNAA +3% sucrose +0.8% agar;
(3) domesticating seedlings: culturing the cultured seedlings in a greenhouse for 10 months to obtain dendrobium officinale plants;
(4) planting: selecting a mountain landform area with the altitude of 500m, the relative humidity of 70% and the annual rainfall of 1200mm, dividing dendrobium officinale plants into a cluster according to 3-5 plants, then placing the cluster into a gap between limestone landform stones or nailing the roots into the stones, wherein the spacing distance between the two clusters of dendrobium officinale plants is 7 cm; watering for 2 times a day within 7 days after cultivation; after 7 days, 1 day of watering 1 time, cultivating for 5 years, and harvesting.
The comparison of the one-year and three-year growth forms of dendrobium officinale in the embodiment of the invention with the wild-returning ecological planting map (A) and the greenhouse planting map (B) is shown in the attached figures 1 and 2. The figure shows that the growth vigor of the dendrobium officinale planted by the method is close to that of wild ecological planting, and the dendrobium officinale planted by the method is obviously different from that planted by a greenhouse.
The comparison of the overall ecological environment of the wild-returning ecological planting and the greenhouse planting of the dendrobium officinale in the embodiment is shown in fig. 3 (a) and 3 (B), and the comparison of the flowering forms of the wild-returning ecological planting and the greenhouse planting of the dendrobium officinale is shown in fig. 4 (a) and 4 (B). The dendrobium officinale planted by the method is closer to wild ecology, and the dendrobium officinale planted by the method has more flowers.
Determination of active ingredients
Polysaccharide content determination was performed on the wild-back ecologically planted dendrobium officinale planted in example 3 and the corresponding greenhouse-planted dendrobium officinale (the polysaccharide content here is the ratio of the extracted dendrobium officinale polysaccharide to the dendrobium officinale powder, not the polysaccharide content determined by the phenol-sulfuric acid method used in the Chinese pharmacopoeia) and the anticancer activity thereof was determined.
(1) Determination of polysaccharide content
Collecting fresh whole plant of herba Dendrobii, cleaning with clear water, cutting into small pieces with file, drying in oven at 60 deg.C for 48 hr until it is completely dried, pulverizing, and sieving with No. 5 sieve.
Respectively weighing 50 g of dendrobium candidum powder planted in the wild-returning ecological environment of Shaoguan Danxia and the greenhouse, adding 1000 mL of petroleum ether, performing reflux extraction for 1 h, filtering, adding 1000 mL of 80% ethanol into filter residue, performing reflux extraction for 1 h, filtering, adding 1500 mL of pure water into the filter residue, performing reflux extraction for 3 times, combining filtrates, concentrating in a rotary evaporator to a proper amount, adding 4 times of volume of absolute ethyl alcohol, standing overnight, filtering, and freeze-drying the filter residue to respectively obtain 15.86g of dendrobium candidum polysaccharide planted in the wild-returning ecological environment, and 13.60g of dendrobium candidum polysaccharide planted in the greenhouse, wherein the mass of the dendrobium candidum polysaccharide accounts for 31.74% and 27.20% of the dendrobium candidum powder.
(2) Research on anti-tumor activity of crude dendrobium officinale polysaccharide
Preparing 4 mg/mL solution of Dendrobium officinale polysaccharide with PBS (phosphate buffered saline), filtering with a sterile 0.22 μm microporous membrane to obtain a dosing mother solution, and preparing for subsequent cell experiments; HeLa cells were cultured in MEM complete medium, passaged, and the experiment was performed when the cells entered the exponential growth phase, the cell concentration was adjusted to 5X 104 cells/mL, inoculated into a 96-well plate, culturing 100 μ L per well for 24 hr, administering with incomplete culture medium to obtain mother liquor with gradient concentration of 25, 50, 100, 200, 400 μ g/mL, administering for 24 hr, adding MTT (3- (4, 5-dimethylthiazole-2) -2, 5-diphenyl tetrazolium bromide) solution, incubating for 4 h, removing culture medium in the pores by suction, adding DMSO (dimethyl sulfoxide), and (3) measuring the absorbance at 490 nm in an enzyme-labeling instrument, and evaluating the activity of the dendrobium officinale polysaccharide planted in the wild ecological mode and the greenhouse mode by taking the inhibition rate as an index.
The result shows that the anticancer activity of the dendrobium candidum polysaccharide planted in the wild ecological environment is obviously higher than that of the dendrobium candidum polysaccharide planted in the greenhouse. Specifically, after 24 hours of administration, the corresponding inhibition rate of the administration concentration of 25-400 mug/mL is 5.0%, 6.6%, 9.8%, 21.7% and 29.8%, the effect has dose dependence, and the inhibition rate of the wild-simulated ecological planting dendrobium officinale crude polysaccharide with the concentration of 400 mug/mL has significance on the HeLa cell of the cervical cancer. The inhibition rates of the crude dendrobium officinale polysaccharide administration concentrations planted in the greenhouse are 8.5%, 9.8%, 9.3%, 5.8% and 14.9% from 25-400 mug/mL, the inhibition rates are not dependent on dosage, and the inhibition rate of the highest concentration is only 14.9%, which is not significant.
The dendrobium officinale product obtained by the wild-returning ecological planting method almost completely simulates the ecological environment planting of wild dendrobium officinale, the polysaccharide content of the obtained dendrobium officinale product can reach 29.8%, and the dendrobium officinale product has a better anti-tumor effect on the pharmacological effect and is far stronger than the dendrobium officinale polysaccharide planted in a greenhouse.
The foregoing is a more detailed description of the invention in connection with specific/preferred embodiments and is not intended to limit the practice of the invention to those descriptions. It will be apparent to those skilled in the art that various substitutions and modifications can be made to the described embodiments without departing from the spirit of the invention, and such substitutions and modifications are to be considered as within the scope of the invention.
Claims (4)
1. A wild-returning ecological planting method for dendrobium officinale is characterized in that: the method comprises the following steps:
(1) seed selection: selecting mature wild or stone-like wild dendrobium officinale capsules;
(2) culturing: taking dendrobium officinale capsules, carrying out tissue culture to obtain protocorms, culturing in a protocorm proliferation culture medium, cutting buds which grow to 1-2cm in height and have 2-3 leaves when the number of proliferated buds reaches 70% -95%, transferring the cut buds to a rooting culture medium, culturing for 25-30 days under a specific temperature condition, and allowing 2-3 white roots with the length of 1cm to appear at the base part to prepare for domesticating seedlings; the protocorm proliferation culture medium has pH =5.8, and MS +10% -30% of potato +1% -5% of sucrose +0.5% -1.2% of agar; the rooting medium comprises the following components in parts by weight: pH =5.4-5.6, MS +8% -20% banana +2.0mg/LNAA +2% -8% sucrose +0.4% -1.2% agar;
(3) domesticating seedlings: culturing the cultured seedlings in a greenhouse for 6-10 months to obtain dendrobium officinale plants;
(4) planting: selecting a mountain landform area with the altitude of 50-600m, the relative humidity of 55-70% and the annual rainfall of 1000-1500mm, dividing the dendrobium officinale plants into a cluster according to 3-5 plants, and then putting the dendrobium officinale plants into gaps between stones or nailing roots into the stones; watering, cultivating for 3-5 years, and collecting;
the stone is one or a combination of a plurality of red conglomerates, sandstones, volcanic rocks, granites or limestone;
the specific temperature of the step (2) is 20-35 ℃ in day and 15-30 ℃ in night;
the specific method for obtaining the protocorm by tissue culture comprises the following steps: selecting mature capsules, performing disinfection and sterilization treatment on the mature capsules on a sterile super clean bench, soaking the mature capsules for 20-30s by using 70% alcohol, then washing the mature capsules for 2-3 times by using sterile water, cutting a 2-3cm mouth at the port of the disinfected capsule by using a scalpel, then hanging the capsule with the opening facing downwards from the mouth of an original bulb induction culture medium, lightly tapping to uniformly spread seeds in the capsule in the original bulb induction culture medium, sowing 3-5 capsules for one capsule, covering a bottle cap after sowing, placing the capsule in a germination culture chamber, performing dark culture for one week, and then transferring the capsule into a normal illumination culture chamber for culture for 45-55 days to obtain the original bulb.
2. The wild-returning ecological planting method for dendrobium officinale according to claim 1, which is characterized in that: watering in the step (4) is to water for 1-2 times every day within 5-7 days after cultivation; after 5-7 days, 1-2 days, pouring 1 time of water.
3. The wild-returning ecological planting method for dendrobium officinale according to claim 1, which is characterized in that: the spacing distance between the two clusters of dendrobium officinale plants is 5-10 cm.
4. The wild-returning ecological planting method for dendrobium officinale according to claim 1, which is characterized in that: the formula of the protocorm induction culture medium comprises the following components: pH =5.8, MS +1.0mg-3.0mg/L6-BA +2-4% sucrose +0.4% -1.2% agar.
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| CN115005075B (en) * | 2022-07-15 | 2023-03-10 | 贵州省亚热带作物研究所 | Planting method for dendrobium nobile by nailing on stone surface in karst region |
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