CN108739053A - A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn - Google Patents
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn Download PDFInfo
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- CN108739053A CN108739053A CN201810648019.5A CN201810648019A CN108739053A CN 108739053 A CN108739053 A CN 108739053A CN 201810648019 A CN201810648019 A CN 201810648019A CN 108739053 A CN108739053 A CN 108739053A
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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
- A01G18/00—Cultivation of mushrooms
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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
- A01G18/00—Cultivation of mushrooms
- A01G18/20—Culture media, e.g. compost
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Abstract
The invention discloses a kind of methods of long-term preservation high cordycepin Cordyceps militaris spawn, preparation including culture medium, and contain 0.5~1 part of maleic hydrazide, 30~50 parts of American aloe extract, 5~10 parts of sodium carboxymethylcellulose, 3~5 parts of triethanolamine, 50~80 parts of water, 400~500 parts of PDA culture medium in the culture medium；Spawn incubation；Culture dehydration, fungi preservation.The present invention prepares culture, preservation of the special culture media for Cordyceps militaris spawn, maleic hydrazide, American aloe extract in culture medium are used cooperatively the growth that can inhibit Cordyceps militaris spawn, the service life is preserved to extend it, sodium carboxymethylcellulose can be good at being retained, so that the Cordyceps militaris spawn of preservation is chronically at optimum state, further extends storage life.The present invention is easy to operate, at low cost, can be good at preserving Cordyceps militaris spawn, and by the Cordyceps militaris spawn that the method for the present invention preserves, storage life can reach 4 years or more, significantly larger than conventional method.
Description
Technical field
The invention belongs to fungi preservation technical fields, and in particular to a kind of side of long-term preservation high cordycepin Cordyceps militaris spawn
Method.
Background technology
It is true to belong to Ascomycota, Hypocreales, cordyceps sinensis Cordycepps, Cordyceps for Cordyceps militaris also known as Cordceps militaris, northern Chinese caterpillar Fungus etc.
Bacterium is a kind of edible medicinal cordyceps sinensis.Riched Cordyceps Militaris containing cordycepin, cordycepic acid and Pentostatin etc. have anticancer, antibacterial, it is antifatigue and
The multiple biological activities ingredient such as immune is improved, medical value is similar to cordyceps sinensis, some ingredients are even better than traditional winter
The worm summer grass, because due to be widely used as cordyceps sinensis substitute be applied to medicinal and edible mushroom field.As people produce cordyceps sinensis
The value recognizations such as the medicinal and health care of product deepen continuously, and the demand to Cordyceps militaris resource and product also increasingly increases.
In recent years, since extensive manual excavates, cause Cordyceps militaris wild resource seriously deficient；And in artificial large-scale production
In the process, also often occur causing spawn degeneration and the phenomena such as serious that makes a variation because strain genetic background is unclear, cause fructification
The problems such as deformity, yield reduction and quality decline, restrict the development and upgrading of cordyceps sinensis industry.Microbiological Culture Collection is mainly
By delaying the metabolism etc. of strain, the strain in Subculture is avoided to make a variation and pollution, in addition it is dead the problems such as
Occur, ensures that strain has growth characteristics that are normal and stablizing and vigor.
Main problem present in Cordyceps militaris spawn preservation at present is：The preservation period is shorter (generally 3~6 months), bacterium
Kind vigor is low, and (with preservation time lengthening, Ability to form fruitbody, which reduces, even can not form fructification, involution form bacterium easily occur
Strain).Therefore, it is necessary to develop a kind of method of new long-term preservation high cordycepin Cordyceps militaris spawn, to extend strain guarantor
The period is hidden, spawn activity is improved.
Invention content
The present invention provides a kind of methods of long-term preservation high cordycepin Cordyceps militaris spawn, solve pupa worm in the prior art
The careless culture presevation period is shorter, the low problem of spawn activity.
The present invention provides a kind of methods of long-term preservation high cordycepin Cordyceps militaris spawn, include the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：0.5~1 part of maleic hydrazide, 30~50 parts of American aloe extract, carboxylic
5~10 parts of sodium carboxymethylcellulose pyce, 3~5 parts of triethanolamine, 50~80 parts of water, 400~500 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains maleic hydrazide
Triethanolamine solution；
Step 1.3, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains water-setting
Glue；
Step 1.4, by three second of maleic hydrazide in the American aloe extract weighed in step 1.1, PDA culture medium, step 1.2
Hydrogel in alkanolamine solution and step 1.3 is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, Cordyceps militaris spawn is inoculated on the Cordyceps militaris spawn slant medium obtained in step 1.4,
First 3~5d of light culture, then carries out 10~15d of optical culture, and culture terminates to obtain Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, it is original 60~70% Cordyceps militaris spawn culture to be dehydrated to volume, obtains dehydration pupa
Cordyceps species culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C.
Preferably, the preparation method of the American aloe extract is as follows：
It takes dry American aloe to pulverize, the second for being equivalent to 10 times of American aloe powder weight is then added into American aloe powder
Alcoholic solution, the ultrasonic extraction 60min at 50 DEG C, is filtered after extraction, collects filtrate, concentrating the filtrate to relative density is
1.2~1.3 to get to the American aloe extract.
Preferably, every liter of PDA culture medium is composed of the following components：Potato 200g, glucose 20g, agar 20g,
Surplus is water.
Preferably, in the step 2, the cultivation temperature of light culture is 23~25 DEG C.
Preferably, in the step 2, the cultivation temperature of optical culture is 23~25 DEG C, and intensity of illumination is 100~200lux.
Preferably, in the step 3, Cordyceps militaris spawn culture is dehydrated in -35~-20 DEG C of sterile cryo drying machine
It is original 50~60% to volume.
Compared with prior art, the beneficial effects of the present invention are：
The present invention prepares culture, preservation of the special culture media for Cordyceps militaris spawn, maleic hydrazide, American aloe in culture medium
Extract is used cooperatively the growth that can inhibit Cordyceps militaris spawn, preserves the service life to extend it, sodium carboxymethylcellulose can
Water conservation well, makes the Cordyceps militaris spawn of preservation be chronically at optimum state, improves bacterial activity, while also making cordyceps
Cordycepin content is substantially unaffected in kind, is taken out after culture presevation, can grow normal mycelium rapidly, further
Extend storage life.
The present invention is easy to operate, at low cost, can be good at preserving Cordyceps militaris spawn, the pupa preserved by the method for the present invention
Cordyceps species, storage life can reach 4 years or more, significantly larger than conventional method.
Specific implementation mode
In order to enable those skilled in the art to more fully understand, technical scheme of the present invention is practiced, with reference to specific
The invention will be further described for embodiment, but illustrated embodiment is not as a limitation of the invention.
Experimental method described in following each embodiments is unless otherwise specified conventional method；The reagent and material,
Unless otherwise specified, it can be commercially available on the market.
Embodiment 1
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：0.5 part of maleic hydrazide, 50 parts of American aloe extract, carboxymethyl are fine
Plain 10 parts of the sodium of dimension, 5 parts of triethanolamine, 80 parts of water, 400 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains maleic hydrazide
Triethanolamine solution；
Step 1.3, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains water-setting
Glue；
Step 1.4, by three second of maleic hydrazide in the American aloe extract weighed in step 1.1, PDA culture medium, step 1.2
Hydrogel in alkanolamine solution and step 1.3 is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, the Cordyceps militaris spawn that cordycepin content is 1.12g/100g is inoculated into step 1.4 and is obtained
Cordyceps militaris spawn slant medium on, first at 23~25 DEG C light culture 3d, be then in 23~25 DEG C, intensity of illumination
Optical culture 15d under conditions of 200lux, culture terminate to obtain Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture is dehydrated in -20 DEG C of sterile cryo drying machine to volume and is
Originally 60%, obtain dehydration Cordyceps militaris spawn culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time is 4 years,
After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is that 1.08g/100g does not become substantially compared with before preservation
Change；
After preserving 4 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA trainings are transplanted under aseptic condition
It supports on base, can be sprouted after 2 days and grow normal mycelium.
Embodiment 2
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：0.8 part of maleic hydrazide, 40 parts of American aloe extract, carboxymethyl are fine
Plain 8 parts of the sodium of dimension, 4 parts of triethanolamine, 60 parts of water, 450 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains maleic hydrazide
Triethanolamine solution；
Step 1.3, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains water-setting
Glue；
Step 1.4, by three second of maleic hydrazide in the American aloe extract weighed in step 1.1, PDA culture medium, step 1.2
Hydrogel in alkanolamine solution and step 1.3 is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, the Cordyceps militaris spawn that cordycepin content is 1.26g/100g is inoculated into step 1.4 and is obtained
Cordyceps militaris spawn slant medium on, first at 23~25 DEG C light culture 5d, be then in 23~25 DEG C, intensity of illumination
Optical culture 10d under conditions of 150lux, culture terminate to obtain Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture is dehydrated in -25 DEG C of sterile cryo drying machine to volume and is
Originally 55%, obtain dehydration Cordyceps militaris spawn culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time 4.5
Year；After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is that 1.28g/100g does not have substantially compared with before preservation
It changes；
After preserving 4.5 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA is transplanted under aseptic condition
It can be sprouted on culture medium, after 2 days and grow normal mycelium.
Embodiment 3
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：1 part of maleic hydrazide, 30 parts of American aloe extract, carboxymethyl cellulose
5 parts of plain sodium, 3 parts of triethanolamine, 50 parts of water, 500 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains maleic hydrazide
Triethanolamine solution；
Step 1.3, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains water-setting
Glue；
Step 1.4, by three second of maleic hydrazide in the American aloe extract weighed in step 1.1, PDA culture medium, step 1.2
Hydrogel in alkanolamine solution and step 1.3 is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, the Cordyceps militaris spawn that cordycepin content is 1.15g/100g is inoculated into step 1.4 and is obtained
Cordyceps militaris spawn slant medium on, first at 23~25 DEG C light culture 4d, be then in 23~25 DEG C, intensity of illumination
Optical culture 12d under conditions of 100lux, culture terminate to obtain Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture is dehydrated in -35 DEG C of sterile cryo drying machine to volume and is
Originally 50%, obtain dehydration Cordyceps militaris spawn culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time is 4 years；
After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is that 1.18g/100g does not become substantially compared with before preservation
Change；
After preserving 4 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA trainings are transplanted under aseptic condition
It supports on base, can be sprouted after 2 days and grow normal mycelium.
It should be noted that the preparation method of American aloe extract is as follows：
It takes dry American aloe to pulverize, the second for being equivalent to 10 times of American aloe powder weight is then added into American aloe powder
Alcoholic solution, the ultrasonic extraction 60min at 50 DEG C, is filtered after extraction, collects filtrate, concentrating the filtrate to relative density is
1.2~1.3 to get to the American aloe extract.
Explanation is needed further exist for, every liter of PDA culture medium is composed of the following components：Potato 200g, glucose 20g,
Agar 20g, surplus are water.
In order to further illustrate effect, the present invention is also provided with comparative example, specific as follows.
Comparative example 1
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, Spawn incubation
Aseptically, Cordyceps militaris spawn is inoculated on PDA slant mediums, first the light culture at 23~25 DEG C
3d, then 23~25 DEG C, intensity of illumination be 200lux under conditions of optical culture 15d, culture terminate obtain Cordyceps militaris spawn training
Support object；
Step 2, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture that cordycepin content is 1.12g/100g is sterile at -20 DEG C
Dehydration to volume is 60% originally in freeze drier, obtains dehydration Cordyceps militaris spawn culture；
Step 3, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time 2.5
Year；After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is 0.96g/100g, compared with before preservation, variation is not
Greatly；
After preserving 2.5 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA is transplanted under aseptic condition
It can just be sprouted on culture medium, after 5 days and grow normal mycelium.
Comparative example 2
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：10 parts of sodium carboxymethylcellulose, 80 parts of water, PDA culture medium 400
Part；
Step 1.2, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains water-setting
Glue；
Step 1.3, the PDA culture medium weighed in step 1.1 is uniformly mixed with the hydrogel in step 1.2, obtains pupa
Cordyceps species culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, Cordyceps militaris spawn is inoculated on the Cordyceps militaris spawn slant medium obtained in step 1.3,
First at 23~25 DEG C light culture 3d, then 23~25 DEG C, intensity of illumination be 200lux under conditions of optical culture 15d, culture
End obtains Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture that cordycepin content is 1.12g/100g is sterile at -20 DEG C
Dehydration to volume is 60% originally in freeze drier, obtains dehydration Cordyceps militaris spawn culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time 2.8
Year；After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is that 1.01g/100g does not have substantially compared with before preservation
It changes；
After preserving 2.8 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA is transplanted under aseptic condition
It can just be sprouted on culture medium, after 5 days and grow normal mycelium.
Comparative example 3
A kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, includes the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：0.5 part of maleic hydrazide, 50 parts of American aloe extract, triethanolamine 5
Part, 400 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains maleic hydrazide
Triethanolamine solution；
Step 1.3, by maleic hydrazide in the American aloe extract, PDA culture medium and the step 1.2 that are weighed in step 1.1
Triethanolamine solution is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn inclined-plane culture
Base；
Step 2, Spawn incubation
Aseptically, Cordyceps militaris spawn is inoculated on the Cordyceps militaris spawn slant medium obtained in step 1.3,
First at 23~25 DEG C light culture 3d, then 23~25 DEG C, intensity of illumination be 200lux under conditions of optical culture 15d, culture
End obtains Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, Cordyceps militaris spawn culture that cordycepin content is 1.12g/100g is sterile at -20 DEG C
Dehydration to volume is 60% originally in freeze drier, obtains dehydration Cordyceps militaris spawn culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration
Cordyceps militaris spawn culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C, the holding time 3.5
Year；After preservation, detect that the content of cordycepin in Cordyceps militaris spawn is that 1.13g/100g does not have substantially compared with before preservation
It changes；
After preserving 2.5 years, the dehydration Cordyceps militaris spawn culture that step 4 is preserved takes out, and PDA is transplanted under aseptic condition
It can just be sprouted on culture medium, after 4 days and grow normal mycelium.
From the correction data of Examples 1 to 3 and comparative example 1~3 it is found that Cordyceps militaris spawn can preserve in Examples 1 to 3
4 years or more, considerably longer than comparative example 1~3, through analysis, the reason is as follows that：
Conventional PDA culture medium culture Cordyceps militaris spawn is sampled in comparative example 1, i.e., is given birth to without any inhibition strain in culture medium
Long ingredient, therefore, holding time only have 2.5 years.
Culture medium has lacked maleic hydrazide and American aloe extract compared with Example 1, in formula in comparative example 2, and maleic hydrazide is
A kind of conventional growth inhibitor can be good at inhibiting plant growth, not notable to the effect for inhibiting Cordyceps militaris spawn growth,
But itself and American aloe extract can be good at the growth for delaying Cordyceps militaris spawn, to extend its preservation when being used cooperatively
Phase.
Culture medium compared with Example 1, has lacked sodium carboxymethylcellulose, sodium carboxymethylcellulose in comparative example 3 in formula
It is a kind of water-retaining agent of superior performance, the Cordyceps militaris spawn of preservation can be made to be in optimum state for a long time, avoids dehydration fast
It spends fast and goes bad.
In summary, the special culture media that the present invention prepares is used for the culture of Cordyceps militaris spawn, preserves, the blueness in culture medium
Fresh element, American aloe extract are used cooperatively the growth that can inhibit Cordyceps militaris spawn, preserve the service life to extend it, carboxymethyl is fine
The plain sodium of dimension can be good at being retained, and the Cordyceps militaris spawn of preservation is made to be chronically at optimum state, improve bacterial activity, and strain is protected
It is taken out after Tibetan, normal mycelium can be grown rapidly, further extend storage life.
It should be noted that involved in description of the invention when numberical range, it is thus understood that two of each numberical range
Any one numerical value can be selected between endpoint and two endpoints, since the step method of use is identical as Examples 1 to 3,
It repeats in order to prevent, description of the invention preferred embodiment, once a person skilled in the art knows basic wounds
The property made concept, then additional changes and modifications may be made to these embodiments.So the following claims are intended to be interpreted as includes
Preferred embodiment and all change and modification for falling into the scope of the invention.
Obviously, various changes and modifications can be made to the invention without departing from essence of the invention by those skilled in the art
God and range.In this way, if these modifications and changes of the present invention belongs to the range of the claims in the present invention and its equivalent technologies
Within, then the present invention is also intended to include these modifications and variations.
Claims (6)
1. a kind of method of long-term preservation high cordycepin Cordyceps militaris spawn, which is characterized in that include the following steps：
Step 1, the preparation of culture medium
Step 1.1, following each component is weighed by weight：0.5~1 part of maleic hydrazide, 30~50 parts of American aloe extract, carboxymethyl
5~10 parts of sodium cellulosate, 3~5 parts of triethanolamine, 50~80 parts of water, 400~500 parts of PDA culture medium；
Step 1.2, the maleic hydrazide weighed in step 1.1 is dissolved in the triethanolamine that step 1.1 weighs, obtains the three of maleic hydrazide
Ethanolamine solutions；
Step 1.3, the sodium carboxymethylcellulose weighed in step 1.1 is dissolved in the water that step 1.1 weighs, obtains hydrogel；
Step 1.4, by the triethanolamine of maleic hydrazide in the American aloe extract weighed in step 1.1, PDA culture medium, step 1.2
Hydrogel in solution and step 1.3 is uniformly mixed, and obtains Cordyceps militaris spawn culture medium；
It will be sub-packed in test tube while hot after Cordyceps militaris spawn culture medium high-temperature sterilization, be frozen into Cordyceps militaris spawn slant medium；
Step 2, Spawn incubation
Aseptically, Cordyceps militaris spawn is inoculated on the Cordyceps militaris spawn slant medium obtained in step 1.4, it is first dark
3~5d is cultivated, then 10~15d of optical culture, culture terminates to obtain Cordyceps militaris spawn culture；
Step 3, culture is dehydrated
Aseptically, it is original 50~60% Cordyceps militaris spawn culture to be dehydrated to volume, obtains dehydration Cordyceps militaris
Microbial strain culture；
Step 4, fungi preservation
The paraffin oil to have sterilized is injected into the test tube equipped with dehydration Cordyceps militaris spawn culture, and paraffin oil is made to flood dehydration pupa worm
Careless microbial strain culture 1cm or more, then be placed on the paraffin sealing to have sterilized and preserved at 4 DEG C.
2. the method for long-term preservation high cordycepin Cordyceps militaris spawn according to claim 1, which is characterized in that the dragon tongue
The preparation method of blue extract is as follows：
Take dry American aloe to pulverize, then into American aloe powder be added be equivalent to 10 times of American aloe powder weight ethyl alcohol it is molten
Liquid, the ultrasonic extraction 60min at 50 DEG C, is filtered after extraction, collect filtrate, concentrate the filtrate to relative density be 1.2~
1.3 to get to the American aloe extract.
3. the method for long-term preservation high cordycepin Cordyceps militaris spawn according to claim 1, which is characterized in that described in every liter
PDA culture medium is composed of the following components：Potato 200g, glucose 20g, agar 20g, surplus are water.
4. the method for long-term preservation high cordycepin Cordyceps militaris spawn according to claim 1, which is characterized in that the step
In 2, the cultivation temperature of light culture is 23~25 DEG C.
5. the method for long-term preservation high cordycepin Cordyceps militaris spawn according to claim 1, which is characterized in that the step
In 2, the cultivation temperature of optical culture is 23~25 DEG C, and intensity of illumination is 100~200lux.
6. the method for long-term preservation high cordycepin Cordyceps militaris spawn according to claim 1, which is characterized in that the step
In 3, it is original 50~60% that Cordyceps militaris spawn culture, which is dehydrated in -35~-20 DEG C of sterile cryo drying machine to volume,.
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