CN104396572A - Cordyceps sinensis strain spore separation, purification and rejuvenation method - Google Patents
Cordyceps sinensis strain spore separation, purification and rejuvenation method Download PDFInfo
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- CN104396572A CN104396572A CN201410777023.3A CN201410777023A CN104396572A CN 104396572 A CN104396572 A CN 104396572A CN 201410777023 A CN201410777023 A CN 201410777023A CN 104396572 A CN104396572 A CN 104396572A
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- test tube
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- stroma
- aweto
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N3/00—Spore forming or isolating processes
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Abstract
The invention provides a cordyceps sinensis strain spore separation, purification and rejuvenation method. The cordyceps sinensis strain spore separation, purification and rejuvenation method comprises the following steps of 1 preparing a solid culture medium, namely thoroughly cooling and filtering potatoes 100 g-140 g, adding glucose10 g-14 g, beef extract 4 g-6 g, KH2PO4 0.4 g-0.6 g, MgSO4 0.2g-0.3 g and agar 8 g-12g, performing heating and boiling till the agar is completely molten, adding distilled water, performing high-pressure steam disinfection and installing an oblique plane; 2 selecting cordyceps sinensis strains with ripening spores, using 0.1% of corrosive sublimate solution to perform disinfection and using sterile water for washing; 3 hamming a small hook at the lower end of a test tube plug, cutting a strain seed seat under the aseptic condition, inversely hanging the strain seed seat on the small hook, and plugging the test tube plug into an oblique plane test tube; 4 keeping the oblique plane test tube in a dark place at the temperature of 22-24 DEG C to perform cultivation for 3-4 days, then performing cultivation at the temperature of 20-22 DEG C for 4-5 days, performing cultivation under the natural light condition at the temperature of 18-22 DEG C for 24 hours and selecting bright-yellow microbe-free strains with developed hyphae.
Description
Technical field
The invention belongs to biological field, relate to a kind of aweto strain spore separation purification and rejuvenation method.
Background technology
Cordyceps sinensis is the compound that Clavicipitaceae Cordyceps Militaris parasitizes stroma on bat moth larvae body and larva body.There is wild and cultivation two kinds.The active ingredient played a role to human body is cordycepin, cordycepic acid, Cordyceps sinensis polysaccharide, superoxide dismutase.There is kidney tonifying benefit lung, hemostasis and phlegm, anticancer, improve immunity, antifatigue, the several functions such as protection lung, kidney, liver.Deeply welcome by many people, be called the soft gold in health products.
After the separation of Cordyceps sinensis wild mushroom and cultivation Chinese caterpillar fungus several generations, degenerating easily appears in kind, need purificate and rejuvenate, but be separated by fruit body, sterilize very difficult, often also mycelia killed or miscellaneous bacteria can not be removed completely, by spore separation again because spore is difficult to obtain, being difficult to realize.
Summary of the invention
The object of the invention is for the deficiencies in the prior art, a kind of low cost, high production are provided, the separating-purifying rejuvenation method of highly active aweto strain spore can be turned out.
The object of the present invention is achieved like this: a kind of aweto strain spore separation purification and rejuvenation method, comprises the steps:
2) solid culture medium is prepared: filling a prescription is: potato 100-140g, glucose 10-14g, beef extract 4-6g, KH
2pO
40.4-0.6g, MgSO
40.2-0.3g, agar 8-12g; First potato is boiled, filter, add glucose, beef extract, KH
2pO
4, MgSO
4, agar, heating is boiled and is allowed agar melt completely, adding distil water, and high pressure steam sterilization, fills inclined-plane processed, stand-by;
2) select the fast ripe aweto strain of spore, with 0.1% mercuric chloride liquid, surface sterilization is carried out, then together with stroma sterile water wash to bacterial classification polypide portion;
3) a little hook is hammered in test tube plug lower end, aseptically cut aweto seed holder, aseptically stroma lower end is hung upside down on the little hook of test tube plug, make stroma point downward, the test tube plug hanging up properly stroma is filled in slant tube stand-by in step 1;
4) cultivate 3-4 days by the slant tube lucifuge 22-24 in step 3 DEG C, then cultivate 4-5 days at 20-22 DEG C, remove the test tube having varied bacteria growing, then 18 ~ 22 DEG C of natral light cahures 24 hours, select without miscellaneous bacteria, mycelia prosperity, the bacterial strain of color cadmium yellow, to obtain final product.
Described aweto strain is wild-type strain or cultivation Chinese caterpillar fungus.
Described worm summer grass wild-type strain is in Nagqu or Yushu district, Qinghai collection.
The invention has the beneficial effects as follows: provide a kind of low cost, high production, the separating-purifying rejuvenation method of highly active aweto strain spore can be turned out.The spore obtained is large, vigor is high, anti-miscellaneous bacteria power strong, can be used as expanding production bacterial classification, successfully achieves the purification and rejuvenation of aweto strain spore separation.
Embodiment
Embodiment 1 pair of Cordyceps sinensis wild-type strain spore separation purification and rejuvenation
To the purification and rejuvenation of Cordyceps sinensis wild-type strain spore separation, operate in accordance with the following steps:
1) solid culture medium is prepared: filling a prescription is: potato 120g, glucose 12g, beef extract 5g, KH
2pO
40.5g, MgSO
40.25g, agar 10g; First potato is boiled, filter, add glucose, beef extract, KH
2pO
4, MgSO
4, agar, heating is boiled and is allowed agar melt completely, adding distil water to 500ml, bacterium 120 DEG C under high steam, 30min, stir dress inclined-plane processed, stand-by;
2) select the fast ripe Cordyceps sinensis wild-type strain of spore, the collection of Cordyceps sinensis wild-type strain should about May, the ground collections such as preferred Nagqu, Yushu district, Qinghai, selects the sturdy prosperity of stroma, the fast fresh wild kind forming spore; With 0.1% mercuric chloride liquid, surface sterilization is carried out, then together with stroma sterile water wash to bacterial classification polypide portion;
3) hammer a little fish hook of stainless steel in test tube stopper lower end central authorities, expose little hook.Aseptically cut aweto seed holder, aseptically stroma lower end is hung upside down on the little hook of test tube plug, make stroma point downward, the test tube plug hanging up properly stroma is filled in slant tube stand-by in step 1; Make stroma point downward, the stopper hanging up properly stroma filled in test tube or triangular flask, stroma not with glass contact, stroma tip to be aimed in test tube in the middle part of medium slant as well.
4) the slant tube lucifuge in step 3 24 DEG C cultivated 4 days, then cultivate 4 days at 20 DEG C, remove and have the test tube of varied bacteria growing, then 18 ~ 22 DEG C of natral light cahures 24 hours, select without miscellaneous bacteria, mycelia prosperity, the bacterial strain of color cadmium yellow, to obtain final product.
Embodiment 2 is to the aweto strain spore separation purification and rejuvenation of cultivation
To the aweto strain spore separation purification and rejuvenation of cultivation, operate in accordance with the following steps:
1) solid culture medium is prepared: filling a prescription is: potato 130g, glucose 13g, beef extract 6g, KH
2pO
40.6g, MgSO
40.3g, agar 11g; First potato is boiled, filter, add glucose, beef extract, KH
2pO
4, MgSO
4, agar, heating is boiled and is allowed agar melt completely, adding distil water to 500ml, bacterium 120 DEG C under high steam, 30min, stir dress inclined-plane processed, stand-by;
2) intensive, the sturdy prosperity of selection stroma, tip are expanded soon, and the aweto strain of the fast ripe cultivation of bright-colored spore, carries out surface sterilization, then together with stroma sterile water wash with 0.1% mercuric chloride liquid to bacterial classification polypide portion;
3) hammer a little fish hook of stainless steel in test tube stopper lower end central authorities, expose little hook.Aseptically cut aweto seed holder, aseptically stroma lower end is hung upside down on the little hook of test tube plug, make stroma point downward, the test tube plug hanging up properly stroma is filled in slant tube stand-by in step 1; Make stroma point downward, the stopper hanging up properly stroma filled in test tube or triangular flask, stroma not with glass contact, stroma tip to be aimed in test tube in the middle part of medium slant as well.
4) the slant tube lucifuge in step 3 24 DEG C cultivated 3 days, then cultivate 5 days at 22 DEG C, remove and have the test tube of varied bacteria growing, then 18 ~ 22 DEG C of natral light cahures 24 hours, select without miscellaneous bacteria, mycelia prosperity, the bacterial strain of color cadmium yellow, to obtain final product.
Claims (3)
1. an aweto strain spore separation purification and rejuvenation method, is characterized in that, comprise the steps:
1) solid culture medium is prepared: filling a prescription is: potato 100-140g, glucose 10-14g, beef extract 4-6g, KH
2pO
40.4-0.6g, MgSO
40.2-0.3g, agar 8-12g; First potato is boiled, filter, add glucose, beef extract, KH
2pO
4, MgSO
4, agar, heating is boiled and is allowed agar melt completely, adding distil water, and high pressure steam sterilization, fills inclined-plane processed, stand-by;
2) select the fast ripe aweto strain of spore, with 0.1% mercuric chloride liquid, surface sterilization is carried out, then together with stroma sterile water wash to bacterial classification polypide portion;
3) hammer a little hook in test tube plug lower end, aseptically cut aweto seed holder, stroma lower end hung upside down on the little hook of test tube plug, make stroma point downward, the test tube plug hanging up properly stroma is filled in slant tube stand-by in step 1;
4) cultivate 3-4 days by the slant tube lucifuge 22-24 in step 3 DEG C, then cultivate 4-5 days at 20-22 DEG C, remove the test tube having varied bacteria growing, then 18 ~ 22 DEG C of natral light cahures 24 hours, select without miscellaneous bacteria, mycelia prosperity, the bacterial strain of color cadmium yellow, to obtain final product.
2. aweto strain spore separation purification and rejuvenation method as claimed in claim 1, is characterized in that, described aweto strain is wild-type strain or cultivation Chinese caterpillar fungus.
3. aweto strain spore separation purification and rejuvenation method as claimed in claim 1, is characterized in that, described worm summer grass wild-type strain is in Nagqu or Yushu district, Qinghai collection.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410777023.3A CN104396572A (en) | 2014-12-15 | 2014-12-15 | Cordyceps sinensis strain spore separation, purification and rejuvenation method |
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|---|---|---|---|
| CN201410777023.3A CN104396572A (en) | 2014-12-15 | 2014-12-15 | Cordyceps sinensis strain spore separation, purification and rejuvenation method |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106612982A (en) * | 2016-12-21 | 2017-05-10 | 广东省微生物研究所 | Efficient pleurotus tuber-regium multi-spore isolation method and multi-spore isolation device used in same |
| CN112342145A (en) * | 2020-11-04 | 2021-02-09 | 青海久实虫草生物科技有限公司 | Rejuvenation method of hirsutella hepiali Chen et Shen fungi |
| JP2021132637A (en) * | 2020-02-24 | 2021-09-13 | 広東省科学院動物研究所 | Application of methylfarnesyl ester in promoting generation of germinated spores of cordy ceps sinensis sacc |
Citations (7)
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|---|---|---|---|---|
| JPH06233627A (en) * | 1993-01-14 | 1994-08-23 | Nobuo Yahagi | Method for artificial mass cultivation of cordyceps sinensis sacc. of imperfect state and perfect stage |
| JPH0947149A (en) * | 1995-08-03 | 1997-02-18 | Fukushima Pref Gov | Artificial cultivation of fruit body of cordyceps sinensis (berk) sacc. |
| CN1766083A (en) * | 2005-09-23 | 2006-05-03 | 田向荣 | Chinese caterpillar fungus bionic culturing method |
| CN101050426A (en) * | 2007-03-28 | 2007-10-10 | 邹玉华 | Method for breeding north caterpillar fungus rich in selenium |
| CN101690463A (en) * | 2009-08-21 | 2010-04-07 | 吉林大学 | Mutagenic strain of cordyceps militaris and breeding method |
| CN102428834A (en) * | 2011-09-20 | 2012-05-02 | 骆新春 | Cultivation method of selenium-enriched Cordyceps militaris mother culture |
| CN102668880A (en) * | 2012-05-14 | 2012-09-19 | 刘继军 | Method for culturing cordyceps militaris bacterium |
-
2014
- 2014-12-15 CN CN201410777023.3A patent/CN104396572A/en active Pending
Patent Citations (7)
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|---|---|---|---|---|
| JPH06233627A (en) * | 1993-01-14 | 1994-08-23 | Nobuo Yahagi | Method for artificial mass cultivation of cordyceps sinensis sacc. of imperfect state and perfect stage |
| JPH0947149A (en) * | 1995-08-03 | 1997-02-18 | Fukushima Pref Gov | Artificial cultivation of fruit body of cordyceps sinensis (berk) sacc. |
| CN1766083A (en) * | 2005-09-23 | 2006-05-03 | 田向荣 | Chinese caterpillar fungus bionic culturing method |
| CN101050426A (en) * | 2007-03-28 | 2007-10-10 | 邹玉华 | Method for breeding north caterpillar fungus rich in selenium |
| CN101690463A (en) * | 2009-08-21 | 2010-04-07 | 吉林大学 | Mutagenic strain of cordyceps militaris and breeding method |
| CN102428834A (en) * | 2011-09-20 | 2012-05-02 | 骆新春 | Cultivation method of selenium-enriched Cordyceps militaris mother culture |
| CN102668880A (en) * | 2012-05-14 | 2012-09-19 | 刘继军 | Method for culturing cordyceps militaris bacterium |
Non-Patent Citations (1)
| Title |
|---|
| 刘锡琎 等: ""冬虫夏草菌无性阶段的分离和鉴定"", 《真菌学报》, vol. 8, no. 1, 31 January 1989 (1989-01-31) * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106612982A (en) * | 2016-12-21 | 2017-05-10 | 广东省微生物研究所 | Efficient pleurotus tuber-regium multi-spore isolation method and multi-spore isolation device used in same |
| JP2021132637A (en) * | 2020-02-24 | 2021-09-13 | 広東省科学院動物研究所 | Application of methylfarnesyl ester in promoting generation of germinated spores of cordy ceps sinensis sacc |
| JP7006988B2 (en) | 2020-02-24 | 2022-01-24 | 広東省科学院動物研究所 | Use of methyl farnesyl ester in promoting germination spore production of Cordyceps sinensis |
| CN112342145A (en) * | 2020-11-04 | 2021-02-09 | 青海久实虫草生物科技有限公司 | Rejuvenation method of hirsutella hepiali Chen et Shen fungi |
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Application publication date: 20150311 |
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