CN105981584B - A kind of cultural method of richness calcium cicada fungus - Google Patents
A kind of cultural method of richness calcium cicada fungus Download PDFInfo
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- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 title claims abstract description 67
- 239000011575 calcium Substances 0.000 title claims abstract description 67
- 229910052791 calcium Inorganic materials 0.000 title claims abstract description 67
- 241000931705 Cicada Species 0.000 title claims abstract description 36
- 241000233866 Fungi Species 0.000 title claims abstract description 35
- 238000000034 method Methods 0.000 title claims abstract description 34
- 239000007787 solid Substances 0.000 claims abstract description 45
- FNAQSUUGMSOBHW-UHFFFAOYSA-H calcium citrate Chemical group [Ca+2].[Ca+2].[Ca+2].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O FNAQSUUGMSOBHW-UHFFFAOYSA-H 0.000 claims abstract description 39
- 239000001354 calcium citrate Substances 0.000 claims abstract description 39
- 235000013337 tricalcium citrate Nutrition 0.000 claims abstract description 39
- 159000000007 calcium salts Chemical class 0.000 claims abstract description 31
- 241001625026 Cordyceps cicadae Species 0.000 claims abstract description 18
- 239000012530 fluid Substances 0.000 claims abstract description 14
- 235000015097 nutrients Nutrition 0.000 claims abstract description 14
- 239000007788 liquid Substances 0.000 claims abstract description 8
- 230000003321 amplification Effects 0.000 claims abstract description 3
- 238000003199 nucleic acid amplification method Methods 0.000 claims abstract description 3
- 239000002609 medium Substances 0.000 claims description 66
- 239000001963 growth medium Substances 0.000 claims description 45
- 239000013078 crystal Substances 0.000 claims description 23
- 240000007594 Oryza sativa Species 0.000 claims description 10
- 235000007164 Oryza sativa Nutrition 0.000 claims description 10
- 241000209140 Triticum Species 0.000 claims description 10
- 235000021307 Triticum Nutrition 0.000 claims description 10
- 235000009566 rice Nutrition 0.000 claims description 10
- 244000068988 Glycine max Species 0.000 claims description 6
- 235000010469 Glycine max Nutrition 0.000 claims description 6
- 235000013339 cereals Nutrition 0.000 claims description 6
- 240000008620 Fagopyrum esculentum Species 0.000 claims description 5
- 235000009419 Fagopyrum esculentum Nutrition 0.000 claims description 5
- 240000005979 Hordeum vulgare Species 0.000 claims description 5
- 235000007340 Hordeum vulgare Nutrition 0.000 claims description 5
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- 229960005069 calcium Drugs 0.000 abstract description 65
- 239000001362 calcium malate Substances 0.000 abstract description 6
- 229940016114 calcium malate Drugs 0.000 abstract description 6
- QXDHJHQRJCJRAU-UHFFFAOYSA-N calcium;2-hydroxypropane-1,2,3-tricarboxylic acid Chemical compound [Ca].OC(=O)CC(O)(C(O)=O)CC(O)=O QXDHJHQRJCJRAU-UHFFFAOYSA-N 0.000 abstract description 6
- VSGNNIFQASZAOI-UHFFFAOYSA-L calcium acetate Chemical compound [Ca+2].CC([O-])=O.CC([O-])=O VSGNNIFQASZAOI-UHFFFAOYSA-L 0.000 abstract description 3
- 239000001639 calcium acetate Substances 0.000 abstract description 3
- 229960005147 calcium acetate Drugs 0.000 abstract description 3
- 235000011092 calcium acetate Nutrition 0.000 abstract description 3
- 239000004227 calcium gluconate Substances 0.000 abstract description 3
- 229960004494 calcium gluconate Drugs 0.000 abstract description 3
- 235000013927 calcium gluconate Nutrition 0.000 abstract description 3
- MKJXYGKVIBWPFZ-UHFFFAOYSA-L calcium lactate Chemical compound [Ca+2].CC(O)C([O-])=O.CC(O)C([O-])=O MKJXYGKVIBWPFZ-UHFFFAOYSA-L 0.000 abstract description 3
- 239000001527 calcium lactate Substances 0.000 abstract description 3
- 229960002401 calcium lactate Drugs 0.000 abstract description 3
- 235000011086 calcium lactate Nutrition 0.000 abstract description 3
- NEEHYRZPVYRGPP-UHFFFAOYSA-L calcium;2,3,4,5,6-pentahydroxyhexanoate Chemical compound [Ca+2].OCC(O)C(O)C(O)C(O)C([O-])=O.OCC(O)C(O)C(O)C(O)C([O-])=O NEEHYRZPVYRGPP-UHFFFAOYSA-L 0.000 abstract description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 12
- HFNBEHZHDROQEC-UHFFFAOYSA-H tricalcium;2-hydroxypropane-1,2,3-tricarboxylate;trihydrate Chemical compound O.O.O.[Ca+2].[Ca+2].[Ca+2].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O HFNBEHZHDROQEC-UHFFFAOYSA-H 0.000 description 11
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- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
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- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 2
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- ZCCIPPOKBCJFDN-UHFFFAOYSA-N calcium nitrate Chemical compound [Ca+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O ZCCIPPOKBCJFDN-UHFFFAOYSA-N 0.000 description 2
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- 235000015099 wheat brans Nutrition 0.000 description 2
- IPDWABJNXLNLRA-UHFFFAOYSA-N 2,3-dihydroxybutanedioic acid;2-hydroxypropane-1,2,3-tricarboxylic acid Chemical compound OC(=O)C(O)C(O)C(O)=O.OC(=O)CC(O)(C(O)=O)CC(O)=O IPDWABJNXLNLRA-UHFFFAOYSA-N 0.000 description 1
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- 239000002126 C01EB10 - Adenosine Substances 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 241000272201 Columbiformes Species 0.000 description 1
- KQLDDLUWUFBQHP-UHFFFAOYSA-N Cordycepin Natural products C1=NC=2C(N)=NC=NC=2N1C1OCC(CO)C1O KQLDDLUWUFBQHP-UHFFFAOYSA-N 0.000 description 1
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- OFEZSBMBBKLLBJ-BAJZRUMYSA-N cordycepin Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)C[C@H]1O OFEZSBMBBKLLBJ-BAJZRUMYSA-N 0.000 description 1
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Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The present invention relates to a kind of cultural methods of rich calcium cicada fungus, and this method comprises the following steps: step 1, Cordyceps cicadae strain expands culture in liquid medium；Step 2, the strain after step 1 amplification is inoculated into solid medium and carries out solid culture；Step 3, coremium is harvested；It is characterized in that, calcium salt is added in step 1 fluid nutrient medium and/or step 2 solid medium；The calcium salt is calcium citrate, citric acid-calcium malate, calcium acetate, calcium gluconate, any one or a few in calcium lactate.
Description
Technical field
The present invention relates to a kind of cultural methods of cicada fungus, and in particular to a kind of cultural method of richness calcium cicada fungus.
Background technique
Cicada fungus (Isaria cicadae Miquel) belongs to the Ascomycota (Ascomycot) of mycota (FUNGI), disk
Bacterium subphylum (Pezizomycotina), excrement shell Gammaproteobacteria (Sordariomycetes), Hypocreales (Hypocreales), cordyceps sinensis section
(Cordycipitaceae), Isaria category (Isaria) is the rare Chinese medicine in China, is to colonize in cicada (being commonly called as " cicada ")
On a kind of Cordyceps Militaris.Its medicinal existing more than 1000 years history, is traditional one of the rare medicinal herbs in China, has various medicines
With value.Main component has adenosine, Cordyceps sinensis polysaccharide, cordycepic acid (mannitol), cordycepin, uracil, sterol, alkaloid, dimension life
Element, inorganic salts, mineral matter element etc..Cicada fungus extensive effect with unique such as improves renal function, antitumor, adjusts and is immunized
System, strengthening by means of tonics improve cell ability, antifatigue, blood pressure lowering, blood glucose, and antipyretic, analgesia improves sleep, improve lipid generation
It thanks, promotes hematopoiesis, antimycotic, anti-aging etc..
Calcium is basic element needed by human, and calcium mainly exists in the form of bone, tooth in human body, 99% in human body
Calcareous there are movement and the masticatory abilities of in bone tooth, supporting human body.Other 1% is present in blood and histoorgan,
Referred to as blood calcium.Calcium deficiency easily leads to children's anorexia, partial eclipse, is not easy to fall asleep, easily wake up with a start, easy catching a cold, hair sparse, intellectual development
It is slow, learn to walk late, teething evening or teethe irregular, paroxysmal abdominal pain diarrhea, X or O-shaped leg, pigeon breast；Calcium deficiency also results in teenager
Energy is not concentrated, fatiguability, is had a pain in the back, immunity is low, decayed tooth or hypoplasia of tooth；Pregnant and lactant women knots weary
Power, arthralgia, dizziness, anaemia and antenatal hypertension syndrome, oedema and galactosis are insufficient；Calcium deficiency causes the elderly senile
Skin disease is itched, and heel pain, lumbar vertebrae cervical vertebra ache, tooth mobility falls off, hence it is evident that bow-backed, height reduces, appetite stimulator, digestion
Road ulcer, dreaminess, insomnia, agitation, irritability etc..It is shown according to " Chinese residents nutrition and health " survey report in 2002, China
People's calcium deficiency situation is still very serious, and the daily intaking amount of resident's calcium is 391 milligrams, just corresponds to the 41% of recommended intake.Cause
It is very necessary that this is replenished the calcium by food.
Prior art culture richness calcium microorganism is mostly added in the medium by calcium original of the inorganic acid salt of calcium, and such as " 3 kinds edible
The research of bacterium mycelium richness calcium " (Bu Qingmei etc., Food Science, 2009), " research of Poria mycelium enrichment calcium " (Wang Wei's rosy clouds
Deng edible mushroom, 2007), " technique and its detection of deep glossy ganoderma fermenting biological concentration calcium " (Miao Jing sesame, Xuzhou Engineering Institute
Journal, in October, 2005) it individually discloses with CaCl2、Ca(NO3)2、CaCO3, CaO be that calcium source carries out rich calcium training to microorganism
It supports.The prior art is not disclosed in the technology that calcium salt culture richness calcium cicada fungus is added in cicada fungus culture medium.
Summary of the invention
The purpose of the present invention is to provide a kind of cultural methods of rich calcium cicada fungus.
It is another object of the present invention to provide a kind of for cultivating the culture medium of cicada fungus.
The purpose of the present invention is what is be achieved by the following scheme:
A kind of cultural method of richness calcium cicada fungus, this method comprises the following steps:
Step 1, Cordyceps cicadae strain expands culture in liquid medium；
Step 2, the strain after step 1 amplification is inoculated into solid medium and carries out solid culture；
Step 3, coremium is harvested；
It is characterized in that, calcium salt is added in step 1 fluid nutrient medium and/or step 2 solid medium.
Further, the calcium salt is the acylate of calcium；Further, the calcium salt is calcium citrate, citric acid-apple
Sour calcium, calcium acetate, calcium gluconate, any one or a few in calcium lactate；Further, the calcium salt be calcium citrate or
Citric acid-calcium malate；Further, the calcium citrate is indissoluble crystal form calcium citrate, calcium citrate trihydrate or high molten crystalline substance
Any one or a few in type calcium citrate；
Further, the adding manner of calcium salt is calcium salt to be added in solid medium or in solid medium and Liquid Culture
Calcium salt is added in base；
Further, it is ultrasonically treated after the step 1 and/or 2 addition calcium salts；Further, sonication treatment time is
10-30min。
Further, during step 2 solid culture, shading culture, cultivation temperature 22-24 are used in the mycelial growth stage
℃；It originates to harvest stages from coremium using illumination cultivation, cultivation temperature is 20-22 DEG C；Relative humidity is in incubation
60-70%；Further, guarantee that intensity of illumination is 100-200Lx when coremium originates.
For the purpose of Liquid Culture process described in step 1 of the present invention is expanded by strain, conventional strain propagation method can be used,
Combination including any one or a few method in inclined-plane culture, shaking flask culture, seed tank culture, fermentation tank culture；It is used
Culture medium is the fluid nutrient medium of this field routine, such as PSA culture medium, PDA culture medium, PDY culture medium, PSB culture medium, yeast
Leach powder-compound amino acid-sucrose culture medium, yeast extract powder-white granulated sugar-soy bean protein hydrolysate culture medium, wheat bran liquor-white
Granulated sugar culture medium etc.；Further preferably PSB culture medium.
Solid medium described in step 2 of the present invention is this field Conventional solid culture medium, and such as grain culture medium can be selected from
Wheat, corn, rice, millet, soybean powder, buckwheat, barley, oat, brown rice, any one or a few in polished rice；Crops
Straw culture medium, such as any one or a few in straw, cornstalk, cotton stalk, beans bar, sesame straw；Crops leather shell culture medium,
Such as wheat bran, soybean skin, cotton seed hulls；Commodity trees branch culture medium etc.；Wherein preferred grain culture medium.
It is a kind of for cultivating the culture medium of cicada fungus, it is characterised in that calcium salt is added in the medium.
Further, the calcium salt is the acylate of calcium；Further, the calcium salt is calcium citrate, citric acid-apple
Sour calcium, calcium acetate, calcium gluconate, any one or a few in calcium lactate；Further, the calcium salt be calcium citrate or
Citric acid-calcium malate；Further, the calcium citrate is indissoluble crystal form calcium citrate, calcium citrate trihydrate or high molten crystalline substance
Any one or a few in type calcium citrate.
Further, it is ultrasonically treated after calcium salt being added in culture medium；Further, sonication treatment time 10-
30min。
Cicada fungus (Isaria cicadae Miquel) used is dispersed species in the present invention, is widely distributed in China Qinling Mountains-Huaihe River
18 provinces, city, areas on the south river.Subtropical zone and torrid areas on the south China the Changjiang river, and it is mostly high in low lying areas and Yunnan-Tibet
The river valleys areas such as former Jinsha jiang River, Nujiang, the Lancang River and the Yarlung Zangbo River.As long as, being pressed the annual 6-8 month in its season of growth
It can all be adopted according to the acquisition method of general Chinese medicine, be strain known in the state of the art, and acquisition can be bought from commercial channels.
Paecilomyces cicadae bacterial strain disclosed in the preferred CN102851353A of the present invention [Paecilomyces cicadae (Miq.) Samson],
It registers and protects at China Committee for Culture Collection of Microorganisms's common micro-organisms center (abbreviation CGMCC) on November 18th, 2009
Hiding, deposit number are CGMCC No.3453.
The present invention is studies have shown that organic calcium salt is added in solid medium and/or fluid nutrient medium to be promoted
The growth of cicada fungus, and the calcium content of its coremium can be significantly improved.
The influence that 1 organic calcium of experimental example grows cicada fungus
1, to the influence of cicada fungus dry weight
Organic calcium: calcium citrate (indissoluble crystal form) and citric acid-calcium malate
Experimental method: cultivating Cordyceps cicadae strain by 1 method of embodiment, and inclined-plane and fluid nutrient medium are PSB culture medium,
Solid medium is wheat broth, and is separately added into different amounts of calcium citrate (indissoluble crystal form) and citric acid-tartaric acid by table 1
Calcium is not added in control group solid medium in calcium.6 groups of repetitions are arranged in control group, and 8 groups of repetitions are arranged in each experimental group.After culture,
Coremium and mycelium gross dry weight are measured respectively, the results are shown in Table 1.
Influence of 1 organic calcium of table to cicada fungus dry weight
It is above-mentioned the experimental results showed that, organic calcium is added in the medium can significantly improve the dry weight of cicada fungus.
2, go out careless influence to cicada fungus
Organic calcium: calcium citrate trihydrate and calcium citrate (high soluble crystal)
Experimental method: cultivating Cordyceps cicadae strain by 1 method of embodiment, and inclined-plane and fluid nutrient medium are PSB culture medium,
Solid medium is wheat broth, and is separately added into different amounts of calcium citrate trihydrate and high soluble crystal calcium citrate by table 2,
Calcium is not added in control group solid medium.6 groups of repetitions are arranged in control group, and 8 groups of repetitions are arranged in each experimental group.After culture, point
Chu not be careless, it the results are shown in Table 2.
The influence that 2 organic calcium of table grows cicada fungus
It is above-mentioned the experimental results showed that, in the medium be added organic calcium can be obviously promoted cicada fungus go out carelessness.
To sum up, this experimental results showed that, organic calcium be added in the medium have to the growth of cicada fungus and apparent promote to make
With.
Influence of 2 organic calcium of experimental example to cicada fungus calcium content
Organic calcium: calcium citrate trihydrate and calcium citrate (high soluble crystal)
Experimental method: cultivating Cordyceps cicadae strain by 1 method of embodiment, and inclined-plane and fluid nutrient medium are PSB culture medium,
Solid medium is wheat broth, and different amounts of organic calcium is added by table 3, and calcium is not added in control group solid medium.It is right
According to group 6 groups of repetitions of setting, 8 groups of repetitions are arranged in each experimental group.After culture, the calcium content of coremium is measured respectively, is as a result seen
Table 3.
The measurement of calcium content: referring to the measurement of standard GB/T/T 5009.92-2003 Calcium In Food, using Atomic absorption
Spectrophotometry.
The influence of 3 calcium citrate trihydrate of table and high soluble crystal calcium citrate to cicada fungus calcium content
The experimental results showed that calcium citrate trihydrate and high soluble crystal calcium citrate is added in cicada fungus is manually cultivated,
The content for improving the calcium in coremium in various degree, to improve original product quality.Especially add in the medium
1.25% high soluble crystal calcium citrate can make the calcium content in coremium increase by 5.4 times.
Influence of the 3 organic calcium adding manner of experimental example to cicada fungus calcium content
1, using high soluble crystal calcium citrate as calcium salt, investigate its in the medium different adding manner to cicada fungus calcium content
Influence.Cultural method is the same as embodiment 2.
Mode one: high soluble crystal calcium citrate is added in solid medium, is added without calcium salt in fluid nutrient medium；
Mode two: high soluble crystal calcium citrate is added in liquid medium, is added without calcium salt in solid medium；
Mode three: high soluble crystal calcium citrate is added in liquid medium within and solid medium；
Control group: high soluble crystal calcium citrate is added without in liquid medium within and solid medium.
Experimental result is shown in Table 4.
Influence of the 4 organic calcium adding manner of table to cicada fungus calcium content
It is above-mentioned the experimental results showed that, calcium is added in solid medium or fluid nutrient medium respectively in three kinds of adding manners
Salt and calcium salt is added in liquid and solid medium simultaneously, the calcium content in coremium can be made to dramatically increase；And three
Kind mode is compared, three better effect of mode one and mode.
2, it using high soluble crystal calcium citrate as calcium salt, selects that calcium is added in liquid medium within and solid medium simultaneously
The processing mode of salt investigates the influence being ultrasonically treated after calcium salt is added in solid medium to cicada fungus calcium content.Culture
Method is the same as embodiment 1.
Mode one: directly Cordyceps cicadae strain is cultivated after calcium salt is added in solid medium
Mode two: ultrasonic treatment 15min again cultivates Cordyceps cicadae strain after calcium salt is added in solid medium
Experimental result is shown in Table 5.
Table 5 is ultrasonically treated the influence to calcium content in cicada fungus
Above-mentioned experimental result is shown, under same culture medium condition, after ultrasonic treatment, can significantly improve coremium
In calcium content, calcium content is not sonicated 3 times or so.
Specific embodiment:
1 Cordyceps cicadae strain cultural method of embodiment
Inclined-plane culture: Cordyceps cicadae strain (embodiment 10) is inoculated in slant tube, then the slant tube that strain will be inoculated with
22 DEG C of incubators are put into, incubation time is 7 days, covers with test tube to mycelia；
Liquid Culture: 200m1 fluid nutrient medium, the high pressure sterilization 30 under 0.11Mpa pressure are packed into 500m1 triangular flask
Minute, it is cooled to room temperature, the strain of 1 slant tube is inoculated into 500m1 triangular flask culture medium, constant temperature oscillation is placed in
Incubator is cultivated under the conditions of 22 ± 1 DEG C of temperature, 150 revs/min, and incubation time is 3 days；
Solid culture: the strain obtained by 25% inoculum concentration inoculation seed culture, the container after inoculation are put into culturing room
Culture, culture room temperature are 22-24 DEG C, relative air humidity 65%-70%；It first has to culturing room's shading making bacteria-producing room base
This dark.When mycelium covers with charge level (indoor culture 3~5 days), it is transferred to illumination cultivation.It inspects periodically after inoculation, picks in time
Except the culture vessel for growing potential difference, infecting miscellaneous bacteria.It hereafter is induced synthesis coremium period, keeping culture room temperature is 20-22
DEG C, relative air humidity 65%~70%, guarantee has scattering light (100Lx~200Lx) induction coremium to be formed.Culturing room will fit
When ventilation, keep bacterium germination room air it is fresh；
Harvesting and processing: harvesting coremium and mycelium；
Fluid nutrient medium: PSB culture medium (200 grams of potato, 20 grams of glucose, water 1000ml)；The 1% molten crystalline substance of height is added
Type calcium citrate；
Solid medium: wheat broth, high solvent type calcium citrate additional amount 6.25%, after suitable quantity of water mixes at ultrasound
Manage 15min.
Embodiment 2
Inclined-plane culture: by Cordyceps cicadae strain, (Paecilomyces cicadae bacterial strain [Paecilomyces cicadae (Miq.) Samson] is protected
Hiding number is CGMCC No.3453) it is inoculated in slant tube, then the slant tube for being inoculated with strain is put into 22 DEG C of incubators, it trains
Supporting the time is 7 days, covers with test tube to mycelia；
Liquid Culture: 200m1 fluid nutrient medium, the high pressure sterilization 30 under 0.11Mpa pressure are packed into 500m1 triangular flask
Minute, it is cooled to room temperature, the strain of 1 slant tube is inoculated into 500m1 triangular flask culture medium, constant temperature oscillation is placed in
Incubator is cultivated under the conditions of 22 ± 1 DEG C of temperature, 150 revs/min, and incubation time is 3 days；
Solid culture: the strain obtained by 25% inoculum concentration inoculation seed culture, the container after inoculation are put into culturing room
Culture, culture room temperature are 22-24 DEG C, relative air humidity 65%-70%；It first has to culturing room's shading making bacteria-producing room base
This dark.When mycelium covers with charge level (indoor culture 3~5 days), it is transferred to illumination cultivation.It inspects periodically after inoculation, picks in time
Except the culture vessel for growing potential difference, infecting miscellaneous bacteria.It hereafter is induced synthesis coremium period, keeping culture room temperature is 20-22
DEG C, relative air humidity 65%~70%, guarantee has scattering light (100Lx~200Lx) induction coremium to be formed.Culturing room will fit
When ventilation, keep bacterium germination room air it is fresh；
Harvesting and processing: harvesting coremium and mycelium；
Fluid nutrient medium: PSB culture medium (200 grams of potato, 20 grams of glucose, water 1000ml)；The 1% molten crystalline substance of height is added
Type calcium citrate；
Solid medium: wheat broth, high solvent type calcium citrate additional amount 6.25%, after suitable quantity of water mixes at ultrasound
Manage 15min.
Embodiment 3
Logical 1 method of embodiment, solid medium: corn culture medium, citric acid-calcium malate additional amount 0.25%, in right amount
Water mixes.Method is as a result as follows with embodiment 1:
Rich calcium effect of 6 citric acid-calcium malate of table in corn culture medium
Embodiment 4
Solid medium: barley culture medium, calcium citrate trihydrate additional amount 0.5%, suitable quantity of water mix.The same embodiment of method
1, as a result as follows:
Rich calcium effect of 7 calcium citrate trihydrate of table on barley culture medium
Embodiment 5
Solid medium: soybean powder medium, high soluble crystal calcium citrate additional amount 1%, suitable quantity of water mix.Method is the same as real
Example one is applied, as a result as follows:
Rich calcium effect of the 8 high soluble crystal calcium citrate of table on soybean powder medium
Embodiment 6
Solid medium: rice medium, calcium citrate additional amount 3%, suitable quantity of water mix.Method is as in the first embodiment, knot
Fruit is as follows:
Rich calcium effect of 9 calcium citrate of table on rice medium
Embodiment 7
Solid medium: buckwheat culture medium, high soluble crystal calcium citrate additional amount 2.5%, suitable quantity of water mix.Method is the same as real
Example one is applied, as a result as follows:
Rich calcium effect of the 10 high soluble crystal calcium citrate of table on buckwheat culture medium
Embodiment 8
Solid medium: polished rice culture medium, calcium citrate trihydrate additional amount 1.25%, suitable quantity of water mix.Method is the same as implementation
Example one is as a result as follows:
Rich calcium effect of 11 calcium citrate trihydrate of table on polished rice culture medium
Embodiment 9
Solid medium: wheat broth, high soluble crystal calcium citrate additional amount 6.25%, suitable quantity of water mix.Method is same
Embodiment one is as a result as follows:
Rich calcium effect of the 12 high soluble crystal calcium citrate of table on wheat broth
Embodiment 10
Subtropical zone and torrid areas low lying areas on the south China the Changjiang river, are adopted July according to the method for general Chinese medicine
Collection.It is aseptically, the fresh cordyceps sinensis for adopting next is clean with aseptic water washing on super mirror workbench, it is placed in sterilized
Diameter is interior sclerotium part (polypide) the absorbent cotton package drenched of cordyceps sinensis, with moisturizing in the culture dish of 15cm.The son of cordyceps sinensis
The glass slide of a sterilizing is placed below entity, aweto sclerotium part is paved with small slide so that can pregnant part not contact directly
Glass slide, distance about 0.5cm or so.Then culture dish is placed in 20 DEG C ± 0.5 DEG C of illumination box culture, to cicada fungus
Spore launches when on glass slide, and it is a little to be added dropwise the PDA culture medium just dissolved around spore, removes slide, is placed in another go out
Moisturizing culture in the culture dish of bacterium turns another plate (SDAY culture medium) training with a small amount of mycelia of transfer needle picking after growing mycelia
It supports (ibid), until the bacterium colony for growing single purifying.And verified through morphology or molecular biology method, which is
Periostracum cicadae (Isaria cicadae Miquel) phorozoon.
Claims (5)
1. a kind of cultural method of richness calcium cicada fungus, this method comprises the following steps:
Step 1, Cordyceps cicadae strain expands culture in liquid medium；
Step 2, the strain after step 1 amplification is inoculated into solid medium and carries out solid culture；
Step 3, coremium is harvested；
It is characterized in that, calcium salt is added in step 1 fluid nutrient medium and step 2 solid medium；The calcium salt is high molten crystalline substance
Type calcium citrate.
2. the method as described in claim 1, which is characterized in that be ultrasonically treated after the step 1 and 2 addition calcium salts.
3. method according to claim 1 or 2, which is characterized in that solid medium described in step 2 is grain culture medium, institute
It is any one in wheat, corn, rice, millet, soybean powder, buckwheat, barley, oat, brown rice, polished rice to state grain culture medium
Kind is several.
4. a kind of for cultivating the culture medium of cicada fungus, which is characterized in that the culture medium is that calcium salt is added in solid medium；
The calcium salt is high soluble crystal calcium citrate；The solid medium be grain culture medium, the grain culture medium be selected from wheat,
Corn, rice, millet, soybean powder, buckwheat, barley, oat, brown rice, any one or a few in polished rice.
5. culture medium as claimed in claim 4, which is characterized in that culture medium is ultrasonically treated after calcium salt is added.
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