CN104694445B - A kind of anaerobe culture medium is compound to go oxygen layer - Google Patents
A kind of anaerobe culture medium is compound to go oxygen layer Download PDFInfo
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- CN104694445B CN104694445B CN201510149249.3A CN201510149249A CN104694445B CN 104694445 B CN104694445 B CN 104694445B CN 201510149249 A CN201510149249 A CN 201510149249A CN 104694445 B CN104694445 B CN 104694445B
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- 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
- C12N1/20—Bacteria; Culture media therefor
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- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
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Abstract
A kind of anaerobe culture medium is compound to go oxygen layer.It is related to a kind of culture medium and goes oxygen layer.It, which solves existing anaerobic culture medium, can not go deoxygenation completely, and reducing agent has a risk for influenceing or endangering anaerobe growth, and anaerobic jar and anaerobic culture box it is complicated, purchase and maintain operating cost height, carry the problems such as inconvenient.Oxygen animalcule culture medium is compound to go oxygen layer to be from bottom to top followed successively by aseptic silica gel plug, the sterile letheen containing medium component, reducing agent and sterile letheen；And aseptic silica gel is provided with the pore penetrated vertically beyond the Great Wall.Anaerobe culture medium of the present invention is compound to go oxygen layer to make simply, and equipment requirement is low, and cost is cheap, is convenient for carrying and makes temporarily, and deaerating effect is good, is particularly suitable for applying in fields such as zymad monitoring, industrial microorganism primary dcreening operations.
Description
Technical field
The present invention relates to a kind of culture medium to go oxygen layer.
Background technology
Anaerobe is a kind of microorganism survived under the conditions of hypoxemia or oxygen-free environment, widely distributed in nature.
The research of anaerobe is long-standing, nowadays in petroleum resources utilization, new energy development utilization, the research and development of new material, stream
The various fields such as the sick prevention and control of row, the diagnosis of special infectious disease, antibiotic development, special fermenting organic material are required for using anaerobism
Microorganism.
Because growth of the oxygen for anaerobe is harmful to, culture anaerobe needs ad hoc approach or equipment, mesh
The method of preceding commonplace use is including the use of anaerobic culture medium, anaerobic jar, anaerobic culture box etc..Anaerobic culture medium passes through
The method that reducing agent is added into culture medium, part removes the oxygen in culture medium, to meet the living necessities of anaerobe, but
Due to that can not go deoxygenation completely, this culture medium is only applicable to the anaerobe existence under hypoxia；And what is added goes back
Immunogenic substance also in the presence of the risk for influenceing or endangering anaerobe growth, limits the application of this culture medium.Anaerobic culturel
Tank and anaerobic culture box are all by the oxygen inside means removal inventionculture tools physically or chemically, to realize to anaerobe
Culture.Although their deaerating effects are good, both equipment are purchased and maintained operating cost high all because complicated, take
The reasons such as band inconvenience can not be widely popularized use.
The content of the invention
The present invention is can not to go deoxygenation completely to solve existing anaerobic culture medium, and reducing agent has influence or endangers anaerobism
The risk of microorganism growth, and anaerobic jar and anaerobic culture box it is complicated, purchase and maintain operating cost high, carrying
The problems such as inconvenient, and a kind of anaerobe culture medium provided compound goes oxygen layer.
Anaerobe culture medium of the present invention is compound go oxygen layer be from bottom to top followed successively by aseptic silica gel plug, containing culture medium into
Sterile letheen, reducing agent and the sterile letheen divided；And aseptic silica gel is provided with the pore penetrated vertically beyond the Great Wall.
The compound aseptic silica gel plug diameter gone in oxygen layer of anaerobe culture medium of the present invention should be slightly bigger than in culture vessel
Footpath, it is easy to fix.Aseptic silica gel plug is located on anaerobe culture medium liquid level or surface, can be cultivated with anaerobe
The liquid level of base or surface contact.Aseptic silica gel plug has supporting role, and fixed anaerobe culture medium is compound to go to oxygen layer position；
Aseptic silica gel plug also has the function that heat-insulated simultaneously, and the sterile letheen containing medium component can be avoided to be killed in forming process
Go out anaerobe.Aseptic silica gel plug has the pore penetrated vertically, and the pore be able to can not be led to ventilating air but liquid agar
Crossing, while fixed aseptic silica gel plug position, the air above anaerobe culture medium is discharged along pore, without
It is extruded and is dissolved into anaerobe culture medium, will not also increases the pressure in culture vessel.
Medium component in sterile letheen of the invention containing medium component should be with anaerobe medium component
It is identical；The sterile letheen containing medium component can be avoided to be contacted with anaerobe culture medium (particularly fluid nutrient medium)
And change or influence the concentration and composition of anaerobe culture medium.Sterile letheen or reducing agent containing medium component
Supporting layer.
Anaerobe culture medium of the present invention is compound to go addition in oxygen layer to have reducing agent, can absorb in culture vessel and remain
Oxygen and incubation in the extraneous oxygen penetrated into；And reducing agent is not entered in anaerobe culture medium, is not interfered with
Or endanger the growth of anaerobe.
Reducing agent is hedged off from the outer world by sterile letheen of the present invention, avoids the Quick Oxidation of reducing agent, and more effectively barrier is outer
The infiltration of boundary's oxygen.
Anaerobe culture medium of the present invention is compound to go oxygen layer to make simply, and equipment requirement is low, and cost is cheap, is convenient for carrying
Made with interim, and deaerating effect is good, is particularly suitable for applying in fields such as zymad monitoring, industrial microorganism primary dcreening operations.
Embodiment
Technical solution of the present invention is not limited to act embodiment set forth below, in addition between each embodiment
Any combination.
Embodiment one：Present embodiment anaerobe culture medium is compound, and to go oxygen layer to be from bottom to top followed successively by sterile
Silica gel plug, the sterile letheen containing medium component, reducing agent and sterile letheen；And aseptic silica gel is provided with vertical insertion beyond the Great Wall
Pore.
Present embodiment aseptic silica gel plug has the pore penetrated vertically, and the pore aperture can be with ventilating air but liquid fine jade
Fat can not pass through.
Embodiment two：The difference of present embodiment and embodiment one is：Anaerobe culture medium
Molecular sieve bed course or bacteria filters are set between the compound sterile letheen and reducing agent containing medium component for going oxygen layer.It is other
Step and parameter are identical with embodiment one.
Molecular sieve bed course and bacteria filters are used to prevent reducing agent from entering the nothing containing medium component in present embodiment
Bacterio-agar, avoid passing through diffusion reducing agent and enter anaerobe culture medium.
Embodiment three：The difference of present embodiment and embodiment one or two is：Anaerobe is trained
Support that base is compound goes oxygen layer also to include N layers reducing agent and sterile letheen on sterile letheen, reducing agent is set with sterile letheen interval
Put, wherein N is natural number.Other steps and parameter are identical with embodiment one or two.
It is repeated several times and sets, starvation effect is more preferable.
Embodiment four：The difference of present embodiment and embodiment one, two or three is：Contain culture medium
Agar concentration is 25g/L in the sterile letheen of composition.Other steps and parameter are identical with embodiment one, two or three.
Agar concentration is the sterile letheens containing medium component prepared of 25g/L to solidify hardness in present embodiment
It is moderate, and will not dry, ftracture again in incubation.
Embodiment five：The difference of present embodiment and one of embodiment one to four is：Sterile letheen
Middle agar concentration is 25g/L.Other steps and parameter are identical with one of embodiment one to four.
Agar concentration is that the sterile letheen that 25g/L is prepared is moderate to solidify hardness in present embodiment, and is being cultivated
During again will not dry, ftracture.
Embodiment six：The difference of present embodiment and one of embodiment one to five is：Reducing agent is
Vulcanized sodium.Other steps and parameter are identical with one of embodiment one to five.
Embodiment 1
Prepare, go oxygen layer using anaerobe culture medium is compound：
1st, will needed for all equipment, culture medium sterilized respectively with high temperature and high pressure steam according to respective feature, filtration sterilization etc.
Method sterilizes；
2nd, anaerobe culture medium is first distributed into culture vessel, then sterilized with high temperature and high pressure steam, now cultivated
Container can be sealed first with interim tampon or silica gel plug, in order to sterilize；
3rd, aseptically, by experimental design, to the anaerobism of the anaerobe inoculation of medium culture after sterilizing
Microbial strains, specific method are identical with traditional vaccination method；
4th, aseptically, aseptic silica gel plug is carefully filled in culture vessel with aseptic inoculation rod, as culture
Base fluid face is in contact, whole air discharge between liquid level；
5th, aseptically, into culture vessel irrigate the sterile letheen containing medium component, containing culture medium into
The nutritional ingredient of the sterile letheen divided is identical with anaerobe medium component, treats its cooled and solidified；
6th, aseptically, molecular sieve bed course or bacteriological filter are placed on the sterile letheen containing medium component
Film, empty its air between the sterile letheen containing medium component；
7th, aseptically, sterile vulcanized sodium or other reducing agents are added；
8th, aseptically, continue to irrigate sterile letheen, treat its cooled and solidified；Realize that anaerobe culture medium is answered
Close the preparation for going oxygen layer.
Repeatable above-mentioned 5th~8 step operation several times, forms sterile letheen-reducing agent composite bed of multilayer.
Culture terminates, and aseptically, carefully successively takes out each layer above culture, particularly careful to take out reduction
Agent, avoid it from remaining in culture vessel, then take out bacterium silica gel plug, culture is drawn with sterile pipette.
Agar concentration is 25g/L in sterile letheen and sterile letheen containing medium component in the present embodiment.
Line blank inoculation test is entered using the present embodiment method, while oxygen indicator methylene blue is added in nutrient solution,
After 28 DEG C of culture 48h, oxygen indicator is negative, illustrates successfully to establish oxygen-free environment.
Carried out using the present embodiment method inoculation anaerobic bacteria C.perfringens (Clostridium perfringens)
Anaerobic culturel, while be inoculated with C.perfringens (Clostridium perfringens) with anaerobic culture box conventional method and enter
Row culture, each inoculation 15,28 DEG C of insulating box culture 48h, initial cell density is 1.0 × 103/mL after inoculation.This reality
It is (5.65 ± 0.12) × 107/mL to apply the C. perfringens cell concentration that a method is turned out, and anaerobic culture box is turned out
C. perfringens cell concentration be (5.70 ± 0.13) × 107/mL.In two kinds of cultural method cultures cell concentration without
Significant difference, illustrate that the culture effect using this method and the culture effect using anaerobic culture box are essentially identical, can be used for
The culture of anaerobe.
Claims (6)
1. a kind of anaerobe culture medium is compound to go oxygen layer, it is characterised in that anaerobe culture medium is compound to go oxygen layer under
Aseptic silica gel plug, the sterile letheen containing medium component, reducing agent and sterile letheen are followed successively by and；And aseptic silica gel is beyond the Great Wall
The pore penetrated vertically is provided with, pore be able to can not be passed through with ventilating air but liquid agar.
2. a kind of anaerobe culture medium according to claim 1 is compound to go oxygen layer, it is characterised in that anaerobe
Culture medium is compound to be gone between the sterile letheen and reducing agent containing medium component of oxygen layer to set molecular sieve bed course or bacterium filter
Film.
3. a kind of anaerobe culture medium according to claim 1 is compound to go oxygen layer, it is characterised in that anaerobe
Culture medium is compound to go oxygen layer also to include N layers reducing agent and sterile letheen, reducing agent and sterile letheen interval on sterile letheen
Set, wherein N is natural number.
4. a kind of anaerobe culture medium according to claim 1 is compound to go oxygen layer, it is characterised in that contains culture medium
Agar concentration is 25g/L in the sterile letheen of composition.
5. a kind of anaerobe culture medium according to claim 1 is compound to go oxygen layer, it is characterised in that in sterile letheen
Agar concentration is 25g/L.
6. a kind of anaerobe culture medium according to claim 1 is compound to go oxygen layer, it is characterised in that reducing agent is sulphur
Change sodium.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201510149249.3A CN104694445B (en) | 2015-04-01 | 2015-04-01 | A kind of anaerobe culture medium is compound to go oxygen layer |
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| CN201510149249.3A CN104694445B (en) | 2015-04-01 | 2015-04-01 | A kind of anaerobe culture medium is compound to go oxygen layer |
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| CN104694445A CN104694445A (en) | 2015-06-10 |
| CN104694445B true CN104694445B (en) | 2018-01-05 |
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| CN108410944A (en) * | 2018-06-05 | 2018-08-17 | 郑州安图生物工程股份有限公司 | A kind of detection clostridium difficile culture medium and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4014748A (en) * | 1975-12-22 | 1977-03-29 | Marion Laboratories, Inc. | Anaerobic culture collecting and transporting apparatus |
| JPS5816673A (en) * | 1981-07-21 | 1983-01-31 | Showa Yakuhin Kako Kk | Device for ready cultivation of anaerobic bacteria |
| CN102358881B (en) * | 2011-10-27 | 2013-01-16 | 成都瑞琦科技实业有限责任公司 | Disposable anaerobe culturing flat plate |
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Effective date of registration: 20211122 Address after: 152400 Xinyue Mingyuan community, Qingyuan community, Qing'an County, Suihua City, Heilongjiang Province Patentee after: Heilongjiang Ruixing Municipal Engineering Co.,Ltd. Address before: 150080 No. 74, Xuefu Road, Nangang District, Heilongjiang, Harbin Patentee before: Heilongjiang University |