CN108715813A - A method of collecting pure mycelia - Google Patents
A method of collecting pure mycelia Download PDFInfo
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- CN108715813A CN108715813A CN201810515281.2A CN201810515281A CN108715813A CN 108715813 A CN108715813 A CN 108715813A CN 201810515281 A CN201810515281 A CN 201810515281A CN 108715813 A CN108715813 A CN 108715813A
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- mycelia
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- 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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
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Abstract
The present invention provides a kind of method for collecting pure mycelia, includes the following steps：Prepare culture medium：Prepare a glass dish for cultivating mycelia, glassine paper is cut into the size less than glass dish；The culture medium prepared is fallen in sterile culture dish on superclean bench, after waiting for that culture medium cools down, sterile glassine paper is placed on culture medium upper surface, bacterial strain is accessed in the culture dish containing culture medium, culture dish sealing is cultivated under conditions of being placed on Suitable strains growth, and growth can be carried out across glassine paper hole for mycelia until mycelia covers with culture dish；On superclean bench bacterial strain is collected using the sterile glass slide of cooling；This method is collected using the glass slide to have gone out.In the medium using glassine paper patch, glassine paper separates mycelia and culture medium, but does not influence mycelia growth, and mycelia can be grown across glassine paper hole, finally be collected mycelia using glass slide, can quickly collect pure mycelia.The result shows that this method not only time-saving and efficiency, but also the quality for collecting sample mycelia is greatly improved.
Description
Technical field
The present invention relates to a kind of methods for collecting pure mycelia.
Background technology
It is essential link in microorganism field experimentation that mycelia, which is collected, uses the progress such as toothpick, pipette tips at present
The mycelia of collection, not only time and effort consuming, and easy to carry a small amount of culture medium, a degree of extraction for affecting later stage sample
And experimental analysis.The problems in collected for current mycelia, a kind of can collect of this experimental study quickly collects pure mycelia
Method.By taking two kinds of pathogens of phytophthora infestans and target bacterium as an example.
Invention content
The present invention improves the above problem, i.e., collects and imitate the technical problem to be solved by the present invention is to existing mycelia
Rate is low to be easy to mix culture medium and influence to detect.
Specific embodiments of the present invention are：1, a kind of method for collecting pure mycelia, which is characterized in that including following step
Suddenly：
（1）, prepare culture medium：In packing to sterile bottle, high pressure sterilization is carried out；
（2）, prepare one for cultivating the glass dish of mycelia, glassine paper is cut into the size less than glass dish, and place glass
High pressure sterilization is carried out together after being packed in ware, and glassine paper has gone out places baking oven drying after bacterium again；
（3）, on superclean bench by the culture medium prepared fall in sterile culture dish, wait for culture medium cooling after, will
Sterile glassine paper is placed on culture medium upper surface, and bacterial strain is accessed in the culture dish containing culture medium, and culture dish sealing is placed on
It is cultivated under conditions of Suitable strains growth, growth can be carried out across glassine paper hole for mycelia until mycelia covers with culture dish；
（4）, glass slide cleaned, place baking oven drying, glass slide be sub-packed in glass dish after drying, will be with glass slide
Glass dish packages laggard horizontal high voltage sterilizing, and baking oven drying is placed again after bacterium of having gone out；
（5）, on superclean bench using the sterile glass slide of cooling be collected bacterial strain, then will be collected by sterile toothpick
Good mycelia is put into the centrifuge tube of sterile 2mL and marks strain number；
（6）, the glass slide after use may be reused again after cleaning.
2, a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the step（4）In
The drying temperature of drying box is 65 DEG C.
3, a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the step（3）And
Step（5）Middle superclean bench uses ultraviolet irradiation 30min or more.
4, a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the mycelia is Ma Ling
When the pure mycelia of potato late disease bacteria, culture medium is rye culture medium：The specific manufacturing method of rye culture medium is as follows, uses electronics
Balance weighs 50g ryes, places and is cleaned in the beaker of 2L, impregnates 8h, is then ground three times using soy bean milk making machine, often
Secondary 1min is positioned over water-bath 2h in 65 DEG C of water-bath later, and water-bath is filtered after terminating using 4 layers of gauze, and 15g is weighed
Agar is added, and pure water is used in combination to be settled to 1L, dissolves by heating, and packing to bottle carries out high pressure sterilization.
5, a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the mycelia is Ma Ling
When the pure mycelia of potato early epidemic germ, culture medium is PDA culture medium, and the PDA culture medium preparation method includes：Use electronics day
It is flat to weigh 200g peeled potatoes, potato cutting is put into pot to and is added a small amount of pure water heating, then uses mashed potatoes
Filtered through gauze weighs 18g agar powders, and 20g glucose is used in combination pure water to be settled to 1L, dissolves by heating, and heating while stirs,
Packing carries out high pressure sterilization to bottle.
Compared with prior art, the invention has the advantages that：In efficiency, the progress such as toothpick, pipette tips are used at present
The mycelia of collection, not only time and effort consuming, and easy to carry a small amount of culture medium, a degree of extraction for affecting later stage sample
And experimental analysis.In addition, the method for the pure mycelia of collection of the invention, can avoid carrying culture medium in the mycelia collected, carry
The high purity and quality of mycelia.
Specific implementation mode
The present invention will be further described in detail With reference to embodiment.
Embodiment one collects the specific method step of the pure mycelia of phytophthora infestans：
（1）, prepare 1L rye culture mediums：50g ryes are weighed using electronic balance, places and is cleaned in the beaker of 2L, are impregnated
Then 8h is ground three times using soy bean milk making machine, each 1min, places water-bath 2h in 65 DEG C of water-bath later, and water-bath terminates
It is filtered later using 4 layers of gauze, weighs the addition of 15g agar（Typically 12g/L, this experiment are 15g/L）, it is used in combination pure
Water is settled to 1L, dissolves by heating（It heats while stirring）, packing to bottle, progress high pressure sterilization.
（2）, glassine paper is cut into is slightly less than the circle of glass dish and places in glass dish, reuse newspaper and rubber band packaging
Carry out high pressure sterilization after good together, glassine paper has gone out places baking oven drying after bacterium again.
（3）, on superclean bench after ultraviolet irradiation 30min, the rye culture medium prepared is fallen in sterile training
It supports in ware, after waiting for the cooling of rye culture medium, sterile glassine paper is placed on rye culture medium, bacterial strain access is contained into rye
In the culture dish of culture medium, it is positioned under 19 DEG C of dark conditions and cultivates two weeks or so after culture dish sealing, i.e., mycelia covers with culture
When ware.
（4）, glass slide cleaned, place baking oven drying（65℃）, be sub-packed in glass dish after drying, reuse newspaper and
Rubber band packages laggard horizontal high voltage sterilizing, and baking oven drying is placed again after bacterium of having gone out.
（5）, before superclean bench after ultraviolet irradiation 30min, bacterial strain is collected using the sterile glass slide of cooling
（Glass slide forced area is far longer than toothpick, and mycelia collection efficiency greatly improves, and culture medium is not easy to be collected into）, then borrow
It helps sterile toothpick to be put into the mycelia gathered in the centrifuge tube of sterile 2mL and marks strain number.
（6）, the glass slide after use may be reused again after cleaning.
Embodiment two collects the specific method step of the pure mycelia of target bacterium：
（1）, prepare 1L PDA culture mediums：200g peeled potatoes are weighed using electronic balance, potato cutting is put into pot
And a small amount of pure water heating is added, then by mashed potatoes filtered through gauze, 18g agar powders are weighed, 20g glucose is used in combination pure
Water is settled to 1L, dissolves by heating（It heats while stirring）, packing to bottle, progress high pressure sterilization.
（2）, glassine paper is cut into is slightly less than the circle of glass dish and places in glass dish, reuse newspaper and rubber band packaging
Carry out high pressure sterilization after good together, glassine paper has gone out places baking oven drying after bacterium again.
（3）, on superclean bench after ultraviolet irradiation 30min, the PDA culture medium prepared is fallen in sterile culture
In ware, after waiting for PDA culture medium cooling, sterile glassine paper is placed in PDA culture medium, then by target bacteria strain
It accesses in the culture dish containing PDA culture medium, is positioned under 25 DEG C of dark conditions after culture dish sealing and cultivates one week or so, i.e. bacterium
When filament length expires culture dish.
（4）, glass slide cleaned, place baking oven drying（65℃）, be sub-packed in glass dish after drying, reuse newspaper and
Rubber band packages laggard horizontal high voltage sterilizing, and baking oven drying is placed again after bacterium of having gone out.
（5）, before superclean bench after ultraviolet irradiation 30min, bacterial strain is collected using the sterile glass slide of cooling,
Since glass slide forced area is far longer than toothpick, mycelia collection efficiency greatly improves, and culture medium is not easy to be collected into, then
The mycelia gathered is put into the centrifuge tube of sterile 2mL by sterile toothpick and marks upper strain number.
（6）, the glass slide after use may be reused again after cleaning.
Any technical solution disclosed in aforementioned present invention unless otherwise stated, if it discloses numberical range,
Disclosed numberical range is preferred numberical range, it is any it should be appreciated by those skilled in the art：Preferred numberical range
The only obvious or representative numerical value of technique effect in many enforceable numerical value.It, can not since numerical value is more
Exhaustion, so the present invention just discloses technical solution of the component values to illustrate the present invention, also, the above-mentioned numerical value enumerated is not
The limitation to the invention protection domain should be constituted.
Claims (5)
1. a kind of method for collecting pure mycelia, which is characterized in that include the following steps：
（1）, prepare culture medium：In packing to sterile bottle, high pressure sterilization is carried out；
（2）, prepare one for cultivating the glass dish of mycelia, glassine paper is cut into the size less than glass dish, and place glass
High pressure sterilization is carried out together after being packed in ware, and glassine paper has gone out places baking oven drying after bacterium again；
（3）, on superclean bench by the culture medium prepared fall in sterile culture dish, wait for culture medium cooling after, will
Sterile glassine paper is placed on culture medium upper surface, and bacterial strain is accessed in the culture dish containing culture medium, and culture dish sealing is placed on
It is cultivated under conditions of Suitable strains growth, growth can be carried out across glassine paper hole for mycelia until mycelia covers with culture dish；
（4）, glass slide cleaned, place baking oven drying, glass slide be sub-packed in glass dish after drying, will be with glass slide
Glass dish packages laggard horizontal high voltage sterilizing, and baking oven drying is placed again after bacterium of having gone out；
（5）, on superclean bench using the sterile glass slide of cooling be collected bacterial strain, then will be collected by sterile toothpick
Good mycelia is put into the centrifuge tube of sterile 2mL and marks strain number；
（6）, the glass slide after use may be reused again after cleaning.
2. a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the step（4）Middle drying
The drying temperature of case is 65 DEG C.
3. a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the step（3）And step
（5）Middle superclean bench uses ultraviolet irradiation 30min or more.
4. a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the mycelia is potato evening
When the pure mycelia of epidemic disease bacterium, culture medium is rye culture medium：The specific manufacturing method of rye culture medium is as follows, uses electronic balance
50g ryes are weighed, places and is cleaned in the beaker of 2L, 8h is impregnated, is then ground three times using soy bean milk making machine, every time
1min is positioned over water-bath 2h in 65 DEG C of water-bath later, and water-bath is filtered after terminating using 4 layers of gauze, and 15g fine jades are weighed
Fat is added, and pure water is used in combination to be settled to 1L, dissolves by heating, and packing to bottle carries out high pressure sterilization.
5. a kind of method for collecting pure mycelia according to claim 1, which is characterized in that the mycelia is that potato is early
When the pure mycelia of epidemic disease bacterium, culture medium is PDA culture medium, and the PDA culture medium preparation method includes：Claimed using electronic balance
200g peeled potatoes are taken, potato cutting is put into pot and a small amount of pure water heating is added, then by mashed potatoes gauze
Filtering, weighs 18g agar powders, and 20g glucose is used in combination pure water to be settled to 1L, dissolves by heating, and heating while stirs, and dispenses
To bottle, high pressure sterilization is carried out.
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| CN201810515281.2A CN108715813A (en) | 2018-05-25 | 2018-05-25 | A method of collecting pure mycelia |
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| CN201810515281.2A CN108715813A (en) | 2018-05-25 | 2018-05-25 | A method of collecting pure mycelia |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110618056A (en) * | 2019-09-29 | 2019-12-27 | 遵义市林业科学研究所 | Method for measuring biomass of fungal hyphae |
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| CN203613177U (en) * | 2013-11-13 | 2014-05-28 | 华南农业大学 | Culture device applicable to microorganism separation and purification |
| CN104611239A (en) * | 2015-02-11 | 2015-05-13 | 广东省微生物研究所 | Culture collection method for fungi mycelium |
| CN105483032A (en) * | 2016-01-27 | 2016-04-13 | 福建农林大学 | Method for separating monosporangiums of potato late blight pathogens |
| CN106591286A (en) * | 2016-11-15 | 2017-04-26 | 南京农业大学 | Method for improving extraction quality of sclerotinia homoeocarpa genome DNA |
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2018
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| CN203613177U (en) * | 2013-11-13 | 2014-05-28 | 华南农业大学 | Culture device applicable to microorganism separation and purification |
| CN104611239A (en) * | 2015-02-11 | 2015-05-13 | 广东省微生物研究所 | Culture collection method for fungi mycelium |
| CN105483032A (en) * | 2016-01-27 | 2016-04-13 | 福建农林大学 | Method for separating monosporangiums of potato late blight pathogens |
| CN106591286A (en) * | 2016-11-15 | 2017-04-26 | 南京农业大学 | Method for improving extraction quality of sclerotinia homoeocarpa genome DNA |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110618056A (en) * | 2019-09-29 | 2019-12-27 | 遵义市林业科学研究所 | Method for measuring biomass of fungal hyphae |
| CN110618056B (en) * | 2019-09-29 | 2022-04-19 | 遵义市林业科学研究所 | Method for measuring biomass of fungal hyphae |
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Application publication date: 20181030 |
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