CN102835317A - Method for producing selenized dendrobium huoshanense culture - Google Patents
Method for producing selenized dendrobium huoshanense culture Download PDFInfo
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- CN102835317A CN102835317A CN2012103667319A CN201210366731A CN102835317A CN 102835317 A CN102835317 A CN 102835317A CN 2012103667319 A CN2012103667319 A CN 2012103667319A CN 201210366731 A CN201210366731 A CN 201210366731A CN 102835317 A CN102835317 A CN 102835317A
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Abstract
The invention discloses a selenized dendrobium huoshanense culture and a method for producing the selenized dendrobium huoshanense culture. The method comprises the steps of: inducing to obtain an aseptic seedling of dendrobium huoshanense, breeding to obtain callus of selenium-resistant dendrobium huoshanense by collaboration of dark and low-salt medium, domesticating the selenized dendrobium huoshanense callus by the low-salt medium assisted by blue light, and producing the selenized dendrobium huoshanense culture by illumination of a fluorescent lamp. The mass fraction of organic selenium in dried product of the selenized dendrobium huoshanense culture produced by the method described by the invention is not less than 50mug/g. According to the method provided by the invention, high-quality dendrobium huoshanense substitute without hormone and pesticide residue can be produced at low cost and large scale via standardization; the selenized dendrobium huoshanense culture product has activity quality of the dendrobium huoshanense and the organic selenium, and can be applied to medicines and health-care products.
Description
Technical field
The invention belongs to field of plant tissue culture technique, be specifically related to a kind of production method of selenizing Dendrobidium huoshanness culture.
Background technology
Selenium is one of requisite trace element among the human life activity, has significantly to delay senility, improve physiological functions such as immunity of organisms, anticancer, anti-oxidant, radioresistance, anti-plant disease.China's most of areas famine selenium element, the earth surface major part is poor selenium water and soil, the Se content overwhelming majority can not satisfy the needs of human body to selenium in the human foods; Therefore; Develop rich selenium product, improve selenium intake, have great importance for the general level of the health that improves the people.
Dendrobidium huoshanness (
Dendrobium huoshanense) be the orchid family Dendrobium herbaceous plant, have the beneficial stomach that promotes the production of body fluid, moistening lung is quenched the thirst, voiceless sound makes eye bright effect, praised highly top grade in the stem of noble dendrobium by the successive dynasties book on Chinese herbal medicine, also receive the favor of people in modern deeply.But the growth of Dendrobidium huoshanness is bordering on harshness to environmental requirement, excessively gathers and the habitat change in addition, and wild Dendrobidium huoshanness resource is seriously exhausted.The exhaustion of resource has seriously restricted the development and use that Dendrobidium huoshanness medicine, health care are worth.
Plant tissue culture technique is the important component part of biotechnology, at the aspects such as preservation of the quick breeding of improved seeds, germ plasm resource by successful Application.People adopt method for tissue culture, form callus or intend protocorm or the bud of growing thickly through explant induction, have obtained Dendrobidium huoshanness sterile test tube seedling, can obtain stem of noble dendrobium regeneration plant after the cultivation.Yet because the particularity of growing environment and physiological trait, test-tube plantlet shifts out when planting in the culture vessel of aseptic and illumination, temperature, moisture stable; Normal photosynthetic physiology activity is difficult to be guaranteed; The growth retardation of test-tube plantlet, pathology or lethality are high, therefore; In the test-tube seedling transplanting domestication stage, need to add hormone to strengthen photoautotrophy ability, anti-adversity ability and the transplanting survival rate of test-tube plantlet; And, behind the test-tube seedling transplanting land for growing field crops, still be unable to do without and spray a large amount of agricultural chemicals with prevention and elimination of disease and pests.For this reason, the Dendrobidium huoshanness resource of field cultivation is also very rare, has only minute quantity to be transplanted in Huoshan the county magistrate towards the medicinal material field.It is thus clear that current urgent need solves the method for producing the Dendrobidium huoshanness of no hormone, non agricultural chemical residuum fast and efficiently, with sustainable use Dendrobidium huoshanness in imminent danger, the service human health.
Callus just comes down to have identical metabolism of plant and morphological development potential at the somatic embryo of Growth and Differentiation, can be used to the active substance that replaces crude drug production relevant.Publication number is that the patent of invention of CN1762207A adopts method for plant tissue culture; Get that dendrobium candidum seed asepsis test-tube plantlet is induced, protocorm is induced, protocorm is tamed; Transfer to and cultivate the seedling growth on the medium, add methyl, sodium selenite and other essential nutrients in the seedling growth medium to produce selenium-enriched officinal dendrobium stem.But the different stems of noble dendrobium are different because of physiological property and life condition separately, and its cultural method is also inequality, Dendrobidium huoshanness itself be exactly in the Dendrobium plant the most difficult worker cultivate kind.
Summary of the invention
In order to realize that low cost, standardization, large-scale production do not have the high-quality Dendrobidium huoshanness substitute of hormone, non agricultural chemical residuum, the present invention provides a kind of production method of selenizing Dendrobidium huoshanness culture.The callus moisture is high, it is tender and crisp to organize, and therefore, the present invention adopts plant tissue culture technique, the collaborative low salt culture medium that contains organic selenium of light control technique, production selenizing Dendrobidium huoshanness culture; Light is not merely photosynthesis of plant radiant energy is provided, also for plant provides Signal Regulation its growth course, and the pattern of growing of controlling plant.
Concrete operations step of the present invention is following:
A kind of production method of selenizing Dendrobidium huoshanness culture comprises that utilizing the high salt culture medium of solid to induce obtains the aseptic seedling of Dendrobidium huoshanness; The Dendrobidium huoshanness of anti-selenium callus is obtained in dark collaborative low salt culture medium seed selection; The auxiliary low salt culture medium domestication selenizing Dendrobidium huoshanness callus of blue light, fluorescent lamp illumination production selenizing Dendrobidium huoshanness culture; The organic selenium mass fraction is not less than 50 μ g/g in the dry selenizing Dendrobidium huoshanness culture.
The concrete production operation step of selenizing Dendrobidium huoshanness culture is following:
(1) inducing of the aseptic seedling of Dendrobidium huoshanness: get ripe Dendrobidium huoshanness fruit; The alcohol water blend rinsing that first use volume fraction is 70～75 mL/100mL 3～5 minutes uses mass fraction to soak 20～30 minutes as the aqueous sodium hypochlorite solution of 2g/100mL again, uses aseptic water washing then 3～5 times; Remove pericarp at last; Take out seed, seed evenly is sprinkling upon on the high salt culture medium of solid, under 20 ± 2 ℃ of temperature, intensity of illumination 2000 ± 200 luxs, 12 hours condition of illumination every day, cultivated 5～6 months; Collect the high aseptic seedling of 4～5cm, be the aseptic seedling of Dendrobidium huoshanness; The high salt culture medium of said solid is made up of the raw material of following weight or volume: sucrose 30000 mg, potassium nitrate 950 mg, ammonium nitrate 825 mg, calcium chloride 220 mg, magnesium sulfate 185 mg, potassium dihydrogen phosphate 85 mg, manganese sulphate 8.45 mg, zinc sulphate 4.3 mg, boric acid 3.1 mg, KI 0.415 mg, sodium molybdate 0.125 mg, copper sulphate 0.0125 mg, cobalt chloride 0.0125 mg, disodium ethylene diamine tetraacetate 18.625 mg, ferrous sulfate 13.925 mg, inositol 50 mg, glycine 1 mg, puridoxine hydrochloride 0.25 mg, nicotinic acid 0.25 mg, thiamine hydrochloride 0.05 mg, agar 5000～6000 mg, water 1 L, pH value 5.8;
(2) seed selection of the Dendrobidium huoshanness of anti-selenium callus: the stem-segment with node of getting long 0.5～1.0 centimetre the aseptic seedling of Dendrobidium huoshanness; Insert in the low salt culture medium of first solid; In 20 ± 2 ℃ of temperature, dark surrounds, cultivated 30～35 days, obtain the Dendrobidium huoshanness of anti-selenium callus;
(3) domestication of selenizing Dendrobidium huoshanness callus: get the Dendrobidium huoshanness of anti-selenium callus, change in the low salt culture medium of second solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2The blue light illumination condition of/s light quantity was cultivated 35～45 days down, collected eugonic emerald first callus; First callus is changed in the low salt culture medium of the 3rd solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald second callus; Second callus is changed in the low salt culture medium of the 4th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 3rd callus; The 3rd callus is changed in the low salt culture medium of the 5th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 4th callus; The 4th callus is changed in the low salt culture medium of the 6th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000～2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 35～45 days; Collect eugonic emerald callus, be selenizing Dendrobidium huoshanness callus provenance;
(4) production of selenizing Dendrobidium huoshanness culture: get selenizing Dendrobidium huoshanness callus provenance; Change in the low salt culture medium of sterilization the 7th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000～2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 45～50 days, get selenizing Dendrobidium huoshanness culture; The organic selenium mass fraction is not less than 50 μ g/g in the dry selenizing Dendrobidium huoshanness culture.
Said first solid hangs down the low salt culture medium of salt culture medium, second solid, the low salt culture medium of the 3rd solid, the low salt culture medium of the 4th solid, the low salt culture medium of the 5th solid, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and forms by the raw material of following weight or volume: sucrose 20000 mg, potassium nitrate 80 mg, citric acid 2.0 mg, nitrate of lime 287.0 mg, magnesium sulfate 738.0 mg, sodium sulphate 53.0 mg, potassium chloride 65.0 mg, sodium dihydrogen phosphate 19.1 mg, thiamine hydrochloride 0.1 mg, pyridoxine hydrochloride 0.1 mg, hydrochloric acid 0.5 mg, glycine 3.0 mg, manganese sulphate 6.6 mg, zinc sulphate 2.7 mg, KI 0.75 mg, boric acid 0.5 mg, agar 5000～6000 mg, water 1 L, pH value 5.8.
Said first solid hangs down in salt culture medium, the low salt culture medium of second solid and all also contains the Se-enriched yeast that mass fraction is 0.1g/L; Go back the Se-enriched yeast that mass fraction is 0.2g/L in the low salt culture medium of the 3rd solid; The low salt culture medium of the 4th solid also mass fraction is the Se-enriched yeast of 0.5g/L, and the 5th solid hangs down in salt culture medium, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and all also contains the Se-enriched yeast that mass fraction is 1.0g/L.
The production bacterial strain of said Se-enriched yeast be the preservation of Chinese common micro-organisms culture presevation administrative center saccharomyces cerevisiae (
Saccharomyces cerevisiae), preserving number CGMCC 0539; Se-enriched yeast is cultivated according to the production method of the patent No. 01100600.5 and is obtained, and is faint yellow or fallow particle or powder, and total selenium is 670 ~ 730 μ g/g, and wherein inorganic selenium must not be crossed 15.0% of total selenium amount.
Compared with prior art, the inventive method has following advantage:
(1) the present invention has realized that low cost, standardization, scale ground produce the high-quality Dendrobidium huoshanness substitute of no hormone, non agricultural chemical residuum, have not only protected Dendrobidium huoshanness germ plasm resource in imminent danger, and for sustainable use Dendrobidium huoshanness resource new way are provided;
(2) the present invention unites the low salt culture medium that contains organic selenium through the light control technique, and the seed selection Dendrobidium huoshanness of anti-selenium callus adopts fluorescent lamp illumination associating organic selenium production selenizing Dendrobidium huoshanness culture, efficient simple, controllable process again;
(3) the selenizing Dendrobidium huoshanness culture produced of the present invention has the double action of organic selenium and Dendrobidium huoshanness concurrently, and product quality is high, improves the Dendrobidium huoshanness industrial competitiveness, increases the Dendrobidium huoshanness economic benefit.
Embodiment
Through embodiment the present invention is done explanation further below.
Embodiment 1:
A kind of concrete production operation step of selenizing Dendrobidium huoshanness culture is following:
Inducing of step 1, the aseptic seedling of Dendrobidium huoshanness: get ripe Dendrobidium huoshanness fruit; Uses earlier volume fraction to be the alcohol water blend rinsing of 70mL/100mL 5 minutes, use mass fraction again, use aseptic water washing then 3 times as the aqueous sodium hypochlorite solution immersion of 2g/100mL 30 minutes; Remove pericarp at last; Take out seed, seed evenly is sprinkling upon on the high salt culture medium of solid, under 20 ± 2 ℃ of temperature, intensity of illumination 2000 ± 200 luxs, 12 hours condition of illumination every day, cultivated 6 months; Collect the high aseptic seedling of 4～5cm, be the aseptic seedling of Dendrobidium huoshanness;
The high salt culture medium of said solid is made up of the raw material of following weight or volume: sucrose 30000 mg, potassium nitrate 950 mg, ammonium nitrate 825 mg, calcium chloride 220 mg, magnesium sulfate 185 mg, potassium dihydrogen phosphate 85 mg, manganese sulphate 8.45 mg, zinc sulphate 4.3 mg, boric acid 3.1 mg, KI 0.415 mg, sodium molybdate 0.125 mg, copper sulphate 0.0125 mg, cobalt chloride 0.0125 mg, disodium ethylene diamine tetraacetate 18.625 mg, ferrous sulfate 13.925 mg, inositol 50 mg, glycine 1 mg, puridoxine hydrochloride 0.25 mg, nicotinic acid 0.25 mg, thiamine hydrochloride 0.05 mg, agar 5000 mg, water 1 L, pH value 5.8;
The seed selection of step 2, the Dendrobidium huoshanness of anti-selenium callus: the stem-segment with node of getting long 0.5-1.0 centimetre the aseptic seedling of Dendrobidium huoshanness; Insert in the low salt culture medium of first solid; In 20 ± 2 ℃ of temperature, dark surrounds, cultivated 30 days, obtain the Dendrobidium huoshanness of anti-selenium callus;
The domestication of step 3, selenizing Dendrobidium huoshanness callus: get the Dendrobidium huoshanness of anti-selenium callus, change in the low salt culture medium of second solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2The blue light illumination condition of/s light quantity was cultivated 35 days down, collected eugonic emerald first callus; First callus is changed in the low salt culture medium of the 3rd solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald second callus; Second callus is changed in the low salt culture medium of the 4th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 3rd callus; The 3rd callus is changed in the low salt culture medium of the 5th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 4th callus; The 4th callus is changed in the low salt culture medium of the 6th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 35 days; Collect eugonic emerald callus, be selenizing Dendrobidium huoshanness callus provenance;
The production of step 4, selenizing Dendrobidium huoshanness culture: get selenizing Dendrobidium huoshanness callus provenance; Change in the low salt culture medium of sterilization the 7th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 50 days, get selenizing Dendrobidium huoshanness culture; Organic selenium content is 50.67 μ g/g in the dried finished of said selenizing Dendrobidium huoshanness culture.
Said first solid hangs down the low salt culture medium of salt culture medium, second solid, the low salt culture medium of the 3rd solid, the low salt culture medium of the 4th solid, the low salt culture medium of the 5th solid, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and forms by the raw material of following weight or volume: sucrose 20000 mg, potassium nitrate 80 mg, citric acid 2.0 mg, nitrate of lime 287.0 mg, magnesium sulfate 738.0 mg, sodium sulphate 53.0 mg, potassium chloride 65.0 mg, sodium dihydrogen phosphate 19.1 mg, thiamine hydrochloride 0.1 mg, pyridoxine hydrochloride 0.1 mg, hydrochloric acid 0.5 mg, glycine 3.0 mg, manganese sulphate 6.6 mg, zinc sulphate 2.7 mg, KI 0.75 mg, boric acid 0.5 mg, agar 5000 mg, water 1 L, pH value 5.8.
Said first solid hangs down in salt culture medium, the low salt culture medium of second solid and all also contains the Se-enriched yeast that mass fraction is 0.1g/L; Go back the Se-enriched yeast that mass fraction is 0.2g/L in the low salt culture medium of the 3rd solid; The low salt culture medium of the 4th solid also mass fraction is the Se-enriched yeast of 0.5g/L, and the 5th solid hangs down in salt culture medium, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and all also contains the Se-enriched yeast that mass fraction is 1.0g/L.The production bacterial strain of Se-enriched yeast be the preservation of Chinese common micro-organisms culture presevation administrative center saccharomyces cerevisiae (
Saccharomyces cerevisiae), preserving number CGMCC 0539.
Table 1
Table 1 is the quality standard of Se-enriched yeast, and Se-enriched yeast is cultivated according to the production method of the patent No. 01100600.5 and obtained.
Embodiment 2:
A kind of concrete production operation step of selenizing Dendrobidium huoshanness culture is following:
Inducing of step 1, the aseptic seedling of Dendrobidium huoshanness: get ripe Dendrobidium huoshanness fruit; Uses earlier volume fraction to be the alcohol water blend rinsing of 75mL/100mL 3 minutes, use mass fraction again, use aseptic water washing then 5 times as the aqueous sodium hypochlorite solution immersion of 2g/100mL 20 minutes; Remove pericarp at last; Take out seed, seed evenly is sprinkling upon on the high salt culture medium of solid, under 20 ± 2 ℃ of temperature, intensity of illumination 2000 ± 200 luxs, 12 hours condition of illumination every day, cultivated 6 months; Collect the high aseptic seedling of 4～5cm, be the aseptic seedling of Dendrobidium huoshanness;
The high salt culture medium agar of said solid is 5500 mg, and the weight or volume that other raw materials are formed is with embodiment 1;
The seed selection of step 2, the Dendrobidium huoshanness of anti-selenium callus: the stem-segment with node of getting long 0.5-1.0 centimetre the aseptic seedling of Dendrobidium huoshanness; Insert in the low salt culture medium of first solid; In 20 ± 2 ℃ of temperature, dark surrounds, cultivated 33 days, obtain the Dendrobidium huoshanness of anti-selenium callus;
The domestication of step 3, selenizing Dendrobidium huoshanness callus: get the Dendrobidium huoshanness of anti-selenium callus, change in the low salt culture medium of second solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2The blue light illumination condition of/s light quantity was cultivated 45 days down, collected eugonic emerald first callus; First callus is changed in the low salt culture medium of the 3rd solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald second callus; Second callus is changed in the low salt culture medium of the 4th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 3rd callus; The 3rd callus is changed in the low salt culture medium of the 5th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 4th callus; The 4th callus is changed in the low salt culture medium of the 6th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2200 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 45 days; Collect eugonic emerald callus, be selenizing Dendrobidium huoshanness callus provenance;
The production of step 4, selenizing Dendrobidium huoshanness culture: get selenizing Dendrobidium huoshanness callus provenance; Change in the low salt culture medium of sterilization the 7th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2200 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 50 days, get selenizing Dendrobidium huoshanness culture; Organic selenium content is 60.32 μ g/g in the dried finished of said selenizing Dendrobidium huoshanness culture.
The agar that said first solid hangs down salt culture medium, the low salt culture medium of second solid, the low salt culture medium of the 3rd solid, the low salt culture medium of the 4th solid, the low salt culture medium of the 5th solid, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid is 5000 mg, and the weight or volume that other raw materials are formed is with embodiment 1.
Embodiment 3:
A kind of concrete production operation step of selenizing Dendrobidium huoshanness culture is following:
Inducing of step 1, the aseptic seedling of Dendrobidium huoshanness: get ripe Dendrobidium huoshanness fruit; Uses earlier volume fraction to be the alcohol water blend rinsing of 75mL/100mL 4 minutes, use mass fraction again, use aseptic water washing then 4 times as the aqueous sodium hypochlorite solution immersion of 2g/100mL 25 minutes; Remove pericarp at last; Take out seed, seed evenly is sprinkling upon on the high salt culture medium of solid, under 20 ± 2 ℃ of temperature, intensity of illumination 2000 ± 200 luxs, 12 hours condition of illumination every day, cultivated 5 months; Collect the high aseptic seedling of 4～5cm, be the aseptic seedling of Dendrobidium huoshanness;
The high salt culture medium agar of said solid is 6000 mg, and the weight or volume that other raw materials are formed is with embodiment 1;
The seed selection of step 2, the Dendrobidium huoshanness of anti-selenium callus: the stem-segment with node of getting long 0.5-1.0 centimetre the aseptic seedling of Dendrobidium huoshanness; Insert in the low salt culture medium of first solid; In 20 ± 2 ℃ of temperature, dark surrounds, cultivated 35 days, obtain the Dendrobidium huoshanness of anti-selenium callus;
The domestication of step 3, selenizing Dendrobidium huoshanness callus: get the Dendrobidium huoshanness of anti-selenium callus, change in the low salt culture medium of second solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2The blue light illumination condition of/s light quantity was cultivated 40 days down, collected eugonic emerald first callus; First callus is changed in the low salt culture medium of the 3rd solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 40 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald second callus; Second callus is changed in the low salt culture medium of the 4th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 40 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 3rd callus; The 3rd callus is changed in the low salt culture medium of the 5th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 40 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 4th callus; The 4th callus is changed in the low salt culture medium of the 6th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 40 days; Collect eugonic emerald callus, be selenizing Dendrobidium huoshanness callus provenance;
The production of step 4, selenizing Dendrobidium huoshanness culture: get selenizing Dendrobidium huoshanness callus provenance; Change in the low salt culture medium of sterilization the 7th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 45 days, get selenizing Dendrobidium huoshanness culture; Organic selenium content is 58.76 μ g/g in the dried finished of said selenizing Dendrobidium huoshanness culture.
The agar that said first solid hangs down salt culture medium, the low salt culture medium of second solid, the low salt culture medium of the 3rd solid, the low salt culture medium of the 4th solid, the low salt culture medium of the 5th solid, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid is 6000 mg, and the weight or volume that other raw materials are formed is with embodiment 1.
Claims (3)
1. the production method of a selenizing Dendrobidium huoshanness culture; It is characterized in that: comprise that utilizing the high salt culture medium of solid to induce obtains the aseptic seedling of Dendrobidium huoshanness; The Dendrobidium huoshanness of anti-selenium callus is obtained in dark collaborative low salt culture medium seed selection; The auxiliary low salt culture medium domestication selenizing Dendrobidium huoshanness callus of blue light, fluorescent lamp illumination production selenizing Dendrobidium huoshanness culture; The organic selenium mass fraction is not less than 50 μ g/g in the dry selenizing Dendrobidium huoshanness culture.
2. the production method of a selenizing Dendrobidium huoshanness culture is characterized in that the concrete production operation step of said selenizing Dendrobidium huoshanness culture is following:
(1) inducing of the aseptic seedling of Dendrobidium huoshanness: get ripe Dendrobidium huoshanness fruit; The alcohol water blend rinsing that first use volume fraction is 70～75 mL/100mL 3～5 minutes uses mass fraction to soak 20～30 minutes as the aqueous sodium hypochlorite solution of 2g/100mL again, uses aseptic water washing then 3～5 times; Remove pericarp at last; Take out seed, seed evenly is sprinkling upon on the high salt culture medium of solid, under 20 ± 2 ℃ of temperature, intensity of illumination 2000 ± 200 luxs, 12 hours condition of illumination every day, cultivated 5～6 months; Collect the high aseptic seedling of 4～5cm, be the aseptic seedling of Dendrobidium huoshanness; The high salt culture medium of said solid is made up of the raw material of following weight or volume: sucrose 30000 mg, potassium nitrate 950 mg, ammonium nitrate 825 mg, calcium chloride 220 mg, magnesium sulfate 185 mg, potassium dihydrogen phosphate 85 mg, manganese sulphate 8.45 mg, zinc sulphate 4.3 mg, boric acid 3.1 mg, KI 0.415 mg, sodium molybdate 0.125 mg, copper sulphate 0.0125 mg, cobalt chloride 0.0125 mg, disodium ethylene diamine tetraacetate 18.625 mg, ferrous sulfate 13.925 mg, inositol 50 mg, glycine 1 mg, puridoxine hydrochloride 0.25 mg, nicotinic acid 0.25 mg, thiamine hydrochloride 0.05 mg, agar 5000～6000 mg, water 1 L, pH value 5.8;
(2) seed selection of the Dendrobidium huoshanness of anti-selenium callus: the stem-segment with node of getting long 0.5～1.0 centimetre the aseptic seedling of Dendrobidium huoshanness; Insert in the low salt culture medium of first solid; In 20 ± 2 ℃ of temperature, dark surrounds, cultivated 30～35 days, obtain the Dendrobidium huoshanness of anti-selenium callus;
(3) domestication of selenizing Dendrobidium huoshanness callus: get the Dendrobidium huoshanness of anti-selenium callus, change in the low salt culture medium of second solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2The blue light illumination condition of/s light quantity was cultivated 35～45 days down, collected eugonic emerald first callus; First callus is changed in the low salt culture medium of the 3rd solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald second callus; Second callus is changed in the low salt culture medium of the 4th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 3rd callus; The 3rd callus is changed in the low salt culture medium of the 5th solid, at 20 ± 2 ℃ of temperature, 60 μ mol/m
2Cultivated 35～45 days under 12 hours the condition of blue light illumination every day of/s light quantity, collect eugonic emerald the 4th callus; The 4th callus is changed in the low salt culture medium of the 6th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000～2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 35～45 days; Collect eugonic emerald callus, be selenizing Dendrobidium huoshanness callus provenance;
(4) production of selenizing Dendrobidium huoshanness culture: get selenizing Dendrobidium huoshanness callus provenance; Change in the low salt culture medium of sterilization the 7th solid; Under 20 ± 2 ℃ of temperature, intensity of illumination 2000～2500 luxs, 12 hours condition of fluorescent lamp illumination every day, cultivated 45～50 days, get selenizing Dendrobidium huoshanness culture; The organic selenium mass fraction is not less than 50 μ g/g in the dry selenizing Dendrobidium huoshanness culture;
Said first solid hangs down the low salt culture medium of salt culture medium, second solid, the low salt culture medium of the 3rd solid, the low salt culture medium of the 4th solid, the low salt culture medium of the 5th solid, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and forms by the raw material of following weight or volume: sucrose 20000 mg, potassium nitrate 80 mg, citric acid 2.0 mg, nitrate of lime 287.0 mg, magnesium sulfate 738.0 mg, sodium sulphate 53.0 mg, potassium chloride 65.0 mg, sodium dihydrogen phosphate 19.1 mg, thiamine hydrochloride 0.1 mg, pyridoxine hydrochloride 0.1 mg, hydrochloric acid 0.5 mg, glycine 3.0 mg, manganese sulphate 6.6 mg, zinc sulphate 2.7 mg, KI 0.75 mg, boric acid 0.5 mg, agar 5000～6000 mg, water 1 L, pH value 5.8;
Said first solid hangs down in salt culture medium, the low salt culture medium of second solid and all also contains the Se-enriched yeast that mass fraction is 0.1g/L; Go back the Se-enriched yeast that mass fraction is 0.2g/L in the low salt culture medium of the 3rd solid; The low salt culture medium of the 4th solid also mass fraction is the Se-enriched yeast of 0.5g/L, and the 5th solid hangs down in salt culture medium, the low salt culture medium of the 6th solid, the low salt culture medium of the 7th solid and all also contains the Se-enriched yeast that mass fraction is 1.0g/L.
3. the production method of a kind of selenizing Dendrobidium huoshanness culture according to claim 2 is characterized in that: the production bacterial strain of said Se-enriched yeast be the preservation of Chinese common micro-organisms culture presevation administrative center saccharomyces cerevisiae (
Saccharomyces cerevisiae), preserving number CGMCC 0539; Se-enriched yeast is cultivated according to the production method of the patent No. 01100600.5 and is obtained, and is faint yellow or fallow particle or powder, and total selenium is 670 ~ 730 μ g/g, and wherein inorganic selenium must not be crossed 15.0% of total selenium amount.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103583364A (en) * | 2013-11-14 | 2014-02-19 | 合肥学院 | Quick hormone-free cultivation method of dendrobium huoshanense seedlings |
| CN103609812A (en) * | 2013-10-22 | 2014-03-05 | 界首市兆龙食品有限公司 | Coconut toffee containing selenized dendrobe |
| CN104996297A (en) * | 2015-06-30 | 2015-10-28 | 傅明尧 | Dendrobium officinale multielement medium |
| CN105010138A (en) * | 2015-06-29 | 2015-11-04 | 广西健宝石斛有限责任公司 | Tissue culture propagation method of Musa balbisiana Colla |
| CN105941079A (en) * | 2016-05-10 | 2016-09-21 | 湖北省硒泰源农业科技有限公司 | Green high-yield organic selenium rich apple plantation facility and production method |
| CN108719068A (en) * | 2018-06-08 | 2018-11-02 | 界首市彭阁家庭农场 | A kind of implantation methods of selenium-rich peanuts |
| CN110839899A (en) * | 2019-11-15 | 2020-02-28 | 扬州大学 | Method for synthesizing selenose by photocatalytic selenium transfer |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1762207A (en) * | 2005-11-11 | 2006-04-26 | 浙江省天然药用植物研究中心 | Method for cultivating selenium-enriched officinal dendrobium stem |
| CN101852368A (en) * | 2010-01-28 | 2010-10-06 | 杭州汉徽光电科技有限公司 | LED blended-light lamp used for tissue culture of orchids |
-
2012
- 2012-09-28 CN CN 201210366731 patent/CN102835317B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1762207A (en) * | 2005-11-11 | 2006-04-26 | 浙江省天然药用植物研究中心 | Method for cultivating selenium-enriched officinal dendrobium stem |
| CN101852368A (en) * | 2010-01-28 | 2010-10-06 | 杭州汉徽光电科技有限公司 | LED blended-light lamp used for tissue culture of orchids |
Non-Patent Citations (1)
| Title |
|---|
| 徐国华等: "硒对铁皮石斛拟原球茎生长及抗氧化系统的影响", 《南京师大学报(自然科学版)》 * |
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|---|---|---|---|---|
| CN103609812A (en) * | 2013-10-22 | 2014-03-05 | 界首市兆龙食品有限公司 | Coconut toffee containing selenized dendrobe |
| CN103609812B (en) * | 2013-10-22 | 2015-04-22 | 界首市兆龙食品有限公司 | Coconut toffee containing selenized dendrobe |
| CN103583364A (en) * | 2013-11-14 | 2014-02-19 | 合肥学院 | Quick hormone-free cultivation method of dendrobium huoshanense seedlings |
| CN103583364B (en) * | 2013-11-14 | 2015-09-23 | 合肥学院 | A kind of Dendrobidium huoshanness seedling is without hormone fast culture process |
| CN105010138A (en) * | 2015-06-29 | 2015-11-04 | 广西健宝石斛有限责任公司 | Tissue culture propagation method of Musa balbisiana Colla |
| CN105010138B (en) * | 2015-06-29 | 2017-05-24 | 广西健宝石斛有限责任公司 | Tissue culture propagation method of Musa balbisiana Colla |
| CN104996297A (en) * | 2015-06-30 | 2015-10-28 | 傅明尧 | Dendrobium officinale multielement medium |
| CN104996297B (en) * | 2015-06-30 | 2017-06-06 | 傅明尧 | A kind of polynary culture medium of dendrobium candidum |
| CN105941079A (en) * | 2016-05-10 | 2016-09-21 | 湖北省硒泰源农业科技有限公司 | Green high-yield organic selenium rich apple plantation facility and production method |
| CN105941079B (en) * | 2016-05-10 | 2019-03-19 | 湖北省硒泰源农业科技有限公司 | A kind of planting facility of green high yield apple rich in organic selenium |
| CN108719068A (en) * | 2018-06-08 | 2018-11-02 | 界首市彭阁家庭农场 | A kind of implantation methods of selenium-rich peanuts |
| CN110839899A (en) * | 2019-11-15 | 2020-02-28 | 扬州大学 | Method for synthesizing selenose by photocatalytic selenium transfer |
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