CN104560688B - A kind of for cultivating device and the cultural method thereof of mixed microorganism - Google Patents

A kind of for cultivating device and the cultural method thereof of mixed microorganism Download PDF

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CN104560688B
CN104560688B CN201310509183.5A CN201310509183A CN104560688B CN 104560688 B CN104560688 B CN 104560688B CN 201310509183 A CN201310509183 A CN 201310509183A CN 104560688 B CN104560688 B CN 104560688B
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inner tube
urceolus
fixing device
dialysis membrane
culture
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CN104560688A (en
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满都拉
杨伟超
张忠泽
徐慧
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Institute of Applied Ecology of CAS
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Institute of Applied Ecology of CAS
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/08—Flask, bottle or test tube
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/24—Gas permeable parts
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/38—Caps; Covers; Plugs; Pouring means
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
    • C12M29/04—Filters; Permeable or porous membranes or plates, e.g. dialysis
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/08—Chemical, biochemical or biological means, e.g. plasma jet, co-culture
    • 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
    • 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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Abstract

The present invention relates to the field of Mixed culture microorganism, be specially a kind of for cultivating device and the cultural method thereof of mixed microorganism. This device comprises porous plug, urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device; Urceolus one end is airtight, and the other end is open type, and inner tube one end connects dialysis membrane fixing device, inner tube is fixed in urceolus by the interior pipe fixing device of at least one, in urceolus and inner tube, add respectively culture medium, and access respectively different bacterial strains, urceolus port seals by porous plug. Device of the present invention, not only can simulate mixed bacterium condition of culture, and can obtain in Mixed culture the pure culture of two aimed strains under different cultivation conditions, for correlation between research Mixed Microbes is provided convenience. Apparatus of the present invention are made simple, easy to use, can be widely used in the interaction relationship research under various mixed bacterium conditions.

Description

A kind of for cultivating device and the cultural method thereof of mixed microorganism
Technical field
The present invention relates to the field of Mixed culture microorganism, be specially a kind of for cultivating device and the cultural method thereof of mixed microorganism.
Background technology
Long-term production practices and increasing scientific research show, numerous microbial processes needs the synergy of multiple bacterial strain (or efficient) to complete, not only in fermentation industry field, as fermentation process for vitamin production, food fermentation and wine, vinegar are brewageed etc., in the environmental protection and the bioenergy field that relate to microorganism, the also reports that have bacterial strain Mixed culture to improve conversion and bioaccumulation efficiency more. Along with mixed bacterium culture technique is in the extensive use of every field, the mixed bacterium interphase interaction relation of research has become one of study hotspot of domestic and international scientist to improve mixed bacterium conversion (degraded) efficiency.
In the research of hybrid bacterial strain interphase interaction relation, obtain the purebred of aimed strain after interacting through Mixed culture and with other bacterial strain, be first to need the problem that solves. Design one can ensure highly to imitate Mixed culture condition, can obtain fast again the purebred experimental provision of aimed strain, could meet us and study the needs that mix bacterium relation. And such device rarely has report. Similarly patent as disclosed in Chinese Patent Application No. 201220446864.2 " use for laboratory anaerobic bacteria dialysis culturing device " is to utilize pellicle as dialysis membrane, and is that this device is unsuitable for studying aerobic bacteria for the Dialysis culture of independent anaerobe. The dialysis culturing device complex structure that " microbial dialysis incubator " of Chinese Patent Application No. 200920104149.9 mentioned, cost is higher. Also there is similar patent report abroad, but be all to utilize pellicle as dialysis membrane, and complex structure, high expensive. Traditional method that is used for studying the relation between bacterium of mixing is that the nutrient solution of a kind of microorganism is joined in objective microbe nutrient solution after filtration, carries out shaking flask cultivation. This method is not only difficult to simulate Mixed culture condition, and wastes time and energy, and microbiological contamination probability is high.
Summary of the invention
The technical issues that need to address of the present invention are to provide a kind of not only simple but also efficiently a kind of for cultivating device and the cultural method thereof of mixed microorganism.
For solving above technical problem, the technical scheme that this method adopts is:
For cultivating a device for mixed microorganism, comprise porous plug, urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device; Urceolus one end is airtight, and the other end is open type, and inner tube one end connects dialysis membrane fixing device, and inner tube is fixed in urceolus by the interior pipe fixing device of at least one, and urceolus open end seals by porous plug.
Outer tube one end is round bottom or flat, described in be connected with inner tube one end of dialysis membrane fixing device and urceolus bottom butt or hung in outer tube by interior pipe fixing device.
Described inner tube one end is provided with external screw thread, is provided with externally threaded inner tube one end and is connected by engage thread with dialysis membrane fixing device.
Described inner tube is provided with and between externally threaded one end and dialysis membrane fixing device, is provided with successively silicone gasket, dialysis membrane, dialysis membrane fixed network from top to bottom.
Described silicone gasket overall diameter equates with the interior diameter of dialysis membrane fixing device.
Described dialysis membrane is the cellulose filter membrane of aperture at 0.22-0.45 μ m.
Described porous plug, urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device are thermostable transparent material and make.
Cultural method: add respectively culture medium in urceolus and inner tube, and access respectively different bacterial strains, utilize the pressure reduction between two pipes, bacterial metabolism material in shaken cultivation inner tube culture medium with medium flow to outer tube, after cultivating a period of time, take out inner tube, the bacterium in urceolus culture medium is the objective microbe of the pure culture obtaining. Described urceolus and inner tube pressure reduction are 1-3cm water column.
The present invention has advantages of:
The present invention adopts thermostable transparent material to make, and by after apparatus of the present invention sterilization treatment, adds respectively culture medium in urceolus and inner tube, makes the culture medium liquid level of inner tube higher than outer tube. Inner and outer pipe accesses respectively after different bacterial strains, and owing to having pressure reduction between two pipes, in the time of shaken cultivation, the bacterial metabolism material in inner tube culture medium can be with medium flow to outer tube, and thalline can not pass through. Cultivate and carefully after a period of time take out inner tube, and bacterium in urceolus culture medium is the objective microbe of the pure culture obtaining, and this microorganism is subject to the effect of inner tube microorganism or has affected. The objective microbe of the pure culture obtaining, can be applicable to carry out the comparative analysis of various biological methods and technology, finally reaches the object of understanding two bacterium correlations and effect.
Device of the present invention, not only can mix bacterium condition of culture by altitude simulation, and can obtain at short notice the pure culture of each aimed strain under different cultivation conditions in Mixed culture, for correlation between research Mixed Microbes is provided convenience. Apparatus of the present invention are made simple, easy to use, can be widely used in the interaction relationship research under various mixed bacterium conditions.
Brief description of the drawings
Fig. 1 is the structural representation of this experimental provision;
Fig. 2 is the dialysis membrane fixing device schematic diagram of this experimental provision;
Wherein: 1, porous plug, 2, outer tube, 3, inner tube, 4, interior pipe fixing device, 5, dialysis membrane fixing device, 6, silicone gasket, 7, dialysis membrane, 8, dialysis membrane fixed network.
Detailed description of the invention
Embodiment 1
For cultivating the device of mixed microorganism, comprise porous plug, urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device; Urceolus one end airtight (blind end bottom can be round bottom or flat), the other end is open type, and inner tube one end connects dialysis membrane fixing device, and inner tube is fixed in urceolus by the interior pipe fixing device of at least one, and urceolus port seals by porous plug. Described inner tube one end is provided with external screw thread, is provided with externally threaded inner tube one end and is connected by engage thread with dialysis membrane fixing device. Described inner tube is provided with and between externally threaded one end and dialysis membrane fixing device, is provided with successively silicone gasket, dialysis membrane, dialysis membrane fixed network from top to bottom. Described silicone gasket overall diameter equates with dialysis membrane fixing device interior diameter. Said apparatus is wrapped and put into autoclave sterilizing with brown paper, stand-by.
Wherein, ventilative plug can be plug or gauze sealing; Urceolus material is glass, external diameter 35.00mm, internal diameter 30.00mm, long 200.00mm; Inner tube material is glass, external diameter 15.00mm, internal diameter 10mm, long 120.00mm; Interior pipe fixing device material is high-temperature resistance plastice, and there is air-vent external diameter 35.00mm, internal diameter 30.00mm, centre; Dialysis membrane fixing device material is high-temperature resistance plastice nut, external diameter 20.00mm, internal diameter 18.00mm, mesopore internal diameter 10.00mm, long 20.00mm; Mesoporous silica gel packing ring material is silica gel, external diameter 18.00mm, mesopore internal diameter 10.00mm; Dialysis membrane is the high temperature resistant filter membranes such as material cellulose filter membrane, aperture 0.22 μ m-0.45 μ m diameter 18.00mm; Dialysis membrane fixed network material is stainless steel, diameter 18.00mm.
The assembling of device:
First dialysis membrane is cut by the internal diameter size of inner tube filter membrane fixture 5, and get wet wetting after, dialysis membrane fixed network 8, filter membrane 7, silicone gasket 6 are put into filter membrane fixture 5 successively. Put careful inner tube rotation into film fixture 5, after screwing, can add suitable water to detect whether completely sealing. Interior pipe fixing device 4 is placed in inner tube 3, then the inner tube that installs interior pipe fixing device is carefully put into urceolus 2, inner tube is pushed up outer tube bottom or is hung in urceolus. Utilize ventilative plug 1 by outer duct occlusion, wrap and put into autoclave sterilizing with brown paper.
Application examples
Existing a kind of aerobic microbiological A bacterium, it can transformation substance a be b, but the poor growth of A bacterium own and conversion capability are low. The effect that A bacterium must just can obtain with another kind of aerobic microbiological B bacterium co-incubation. B bacterium itself can be good at growth. So in order to study B bacterium for which type of material A bacterium provides, when the metabolism and growth that has caused A bacterium changes and A bacterium which itself has occurred after the impact of the B bacterium being subject to changes, the A bacterium that will obtain A bacterium and be subject to the pure culture after B bacterium affects. In gnotobasis, to the culture medium that adds respectively suitable A bacterium and B bacterium in inner tube 3 and urceolus 2, wherein in inner tube, add B bacterium respectively, urceolus adds a small amount of A bacterium, fill in ventilative plug, tiltedly put after a period of time is cultivated in concussion and measure corresponding index according to the training method of existing A bacterium and B bacterium. After cultivation completes in gnotobasis to the nutrient solution coating in urceolus or microscopy to guarantee to obtain pure A bacterium.
The measurement result of material b:
After cultivating, obtained the purebred A bacterium through the impact of B bacterium in apparatus of the present invention, and not by B fungi pollution.

Claims (4)

1. for cultivating a device for mixed microorganism, it is characterized in that: device comprise porous plug,Urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device; Urceolus one end is airtight, and the other end isOpen type, inner tube one end connects dialysis membrane fixing device, and inner tube is by the fixing dress of inner tube of at least onePut and be fixed in urceolus, urceolus open end seals by porous plug;
Described inner tube one end is provided with external screw thread, is provided with externally threaded inner tube one end and dialysis membrane fixing deviceConnect by engage thread;
Described inner tube is provided with between externally threaded one end and dialysis membrane fixing device and is from top to bottom provided with successivelySilicone gasket, dialysis membrane, dialysis membrane fixed network;
Described silicone gasket overall diameter equates with the interior diameter of dialysis membrane fixing device;
Described dialysis membrane is the cellulose filter membrane of aperture at 0.22-0.45 μ m.
By claimed in claim 1 for cultivating the device of mixed microorganism, it is characterized in that: instituteState porous plug, urceolus, inner tube, interior pipe fixing device and dialysis membrane fixing device and be thermostable transparentMaterial is made.
3. utilize a cultural method for the cultivation mixed microorganism device described in claim 1, its spyLevy and be: in urceolus and inner tube, add respectively culture medium, make the culture medium liquid level of inner tube higher than outer tube;Inner and outer pipe accesses respectively different bacterial strains, utilizes the pressure reduction between two pipes, shaken cultivation inner tube culture mediumIn bacterial metabolism material with medium flow to outer tube, cultivate after a period of time and take out inner tube, urceolus trainingBacterium in foster base is the objective microbe of the pure culture obtaining.
4. by cultural method claimed in claim 3, it is characterized in that: described urceolus and inner tube pressure reductionFor 1-3cm water column.
CN201310509183.5A 2013-10-23 2013-10-23 A kind of for cultivating device and the cultural method thereof of mixed microorganism Active CN104560688B (en)

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CN105950458B (en) * 2016-05-27 2018-09-04 哈尔滨师范大学 Mixed microorganism culture apparatus
CN106148164B (en) * 2016-06-30 2018-09-07 南京师范大学 A kind of dialysis apparatus and the microcosm experiment device comprising the dialysis apparatus
CN108238360A (en) * 2018-01-12 2018-07-03 长春理工大学 A kind of steam sterilizing Packaging Bottle
CN111139174B (en) * 2020-01-15 2023-08-29 辽东学院 Mixed microorganism culture apparatus

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CN2570288Y (en) * 2002-09-30 2003-09-03 周晋 Disposable semipermeabile connection type cell separating combined culture bottle
CN201325964Y (en) * 2008-12-18 2009-10-14 崔生发 Open circulation type tubular microorganism dialysis cultivation system
CN201459137U (en) * 2009-08-19 2010-05-12 林涛 Microbial dialysis incubator
CN202786236U (en) * 2012-09-04 2013-03-13 西北民族大学 Dialysis culture device of anaerobic bacteria for laboratory
CN203112832U (en) * 2012-11-02 2013-08-07 中华人民共和国上海出入境检验检疫局 Multifunctional test tube for culturing bacteria
CN203559051U (en) * 2013-10-23 2014-04-23 中国科学院沈阳应用生态研究所 Device for cultivating mixed microbes

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