CN1203943A - Airlift cyclic immersion lighting plant cell tissue organ culture method and culture reactor - Google Patents
Airlift cyclic immersion lighting plant cell tissue organ culture method and culture reactor Download PDFInfo
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Abstract
The present invention relates to a method for culturing plant cell, tissue and organ and its culture reactor. Said culture reactor consists of main body, light source, separation layers, inoculating tube, gas distributor, liquid-storage tank, electromagnetic valve, gas-liquid circulating pipe and gas-liquid distributing pipe, etc.. Its culture method includes the following steps: steam-sterilization at 100-121 deg.C, for 40-200 min., introducing sterile air to make protection,and under the condition of room temp. and in sterile environment the culture material is added by layers, capping, introducing culture liquid and regulating time sequence apparatus according to the immersion time and airing period required for culture material to implement automatic culture.
Description
The present invention relates to the method for a kind of vegetable cell, tissue and organ culture, particularly relate to a kind of cultural method and reactor thereof that is suitable for air lift type cycle submergence illumination plant cell, tissue and organ that scale operation uses.
The culture plant cell bio-reactor has been studied more, mainly be divided into mechanical stirring bio-reactor, on-mechanical stirred bioreactor, bubble tower and airlift bioreactor, the packed bed bio-reactor, the fluidized bed bio reactor, (Chen Yingliang etc. " cell cultures engineering " such as membrane bioreactor, Huadong Chemical College press, 1992, Shanghai P285-294).Much be different from zooblast and microorganism because vegetable cell, tissue and organ have, less as oxygen requirement, cell is poor to shear stress tolerance, and cell volume is big etc., so the culture plant cell bio-reactor has been proposed different requirements.Wherein the easy pair cell of high-shear characteristic of mechanical stir-reactor causes damage, and must be improved agitator, reduces the shearing force in the whipping process, just can be used for vegetable cell and tissue etc. and cultivate (Xing Jianmin etc., biotechnology progress, 1997,17 (5), P49-53).Hydrodynamic shear has the effect of two aspects to vegetable cell.The first increases ventilation, keeps good mixing state and cell dispersiveness etc., its two be in most of the cases then pair cell physical abuse is arranged, change cellular form, influence compound release in the born of the same parents, change metabolism, influence productive rate, make cell loss of activity etc., therefore research and develop the novel bio-reactor that is applicable to all kinds of culture plant cell and become direction (Cao Mengde etc., biotechnology progress, 1996 that various countries biochemical engineering scholar makes great efforts, 16 (4), P51-53).Hydrodynamic shear has the effect of two aspects to vegetable cell.The first increases ventilation, keeps good mixing state and cell dispersiveness etc., its two be in most of the cases then pair cell physical abuse is arranged, change cellular form, influence compound release in the born of the same parents, change metabolism, influence productive rate, make cell loss of activity etc., therefore research and develop the novel bio-reactor that is applicable to all kinds of culture plant cell and become direction (Cao Mengde etc., biotechnology progress, 1996 that various countries biochemical engineering scholar makes great efforts, 16 (4), P51-53).As for plant organ, the tissue than vegetable cell big tens times even hundreds of times.These organs comprise with organizing mainly: the bud of growing thickly, hairly root (root of hair), somatic embryo, seedling etc.Because the specific form of these tissues, organ is mobile poor in culturing process, easily piles up, more responsive to shearing force, long-term easier generation vitrifying of submergence and variation, therefore, with respect to vegetable cell, realize that in reactor large scale culturing is just more complicated, difficult.Begin vegetable cell, tissue and organ culture from the seventies later stage and adopted airlift bioreactor morely.As (" Bioreactors for growth of plant roots " M.A.Rodrigueg-Mendiola et al.Enzyme Microb.Technol.1991 such as document M.A.Rodrigueg-Mendiola, Vol.13 sep.697-702) the 9 liter plant rootings of being introduced are cultivated reactor, and its structure is seen Fig. 1.Reactor body 1 is made by the thermal glass that has expanding reach.Fix the pipe core 28 that a nylon wire is rolled into by glass support pipe 15 in the main body.Be positioned in reactor below the pipe core, a sintered plate gas distributor 6 is set, required air enters pipe core through inlet pipe 30, because the inside and outside liquid apparent density official post nutritive medium of pipe core is realized circulating.Reactor head is provided with an inoculation mouth 25, and the root of hair seed inserts in the reactor thus.Gas in the reactor is discharged behind condenser 21.Reactor temperature is by well heater 19 controls.In culturing process, the root of hair seed hangs on the central network grows, and these reactor characteristics have solved inoculation and hung over stationary distribution and growth on the pipe core, and part has solved root of hair and piled up problem, and nutritive medium can circulate and have mixing to a certain degree.But because no jacket space is big, so not basic homogeneity that solves inoculation, the uniform distribution of root of hair process of growth, even particularly around pipe core is off the net mouthful, still can form the accumulation of local root of hair in the bottom, so that influence circulating of nutritive medium at the root of hair growing period, thereby influence mixed characteristic and mass transfer, heat transfer process, and this type of reactor belongs to full immersion reactor, the long-term submergence of plant tissue organ wherein can produce vitrification phenomenon and variation, owing in the reactor internal light source is not set, root of hair to happiness illumination is cultivated, and the amplification of reactor diameter must be restricted.Therefore, this type of reactor is difficult to amplify to adapt to the large scale culturing of plant rooting.
Chun-zhao Liu and for example, (" Production of arthmisininby shoot cultures of Arthmisia annua L in a modified inner-loop mist bioreactor " such as Yuchun Wang, Plant science, receive and publish, 1988) research and develop out the interior circulation ultrasonic atomization reactor that 2.5 liters are transformed.Its primary structure as shown in Figure 2.This reactor body 1 is made by the hot glass of anti-liquid, built-in one has the pipe core 18 in hole 20, around pipe core three layers of stainless steel filter 26 is arranged, and at reactor bottom one atomising head 17 is installed, and be connected with spraying gun 16, microbiological free air is entered in the reactor bottom nutritive medium by inlet pipe 30.Gas is by venting port 27 dischargings in the reactor, this reactor does not have classification inoculation apparatus, need reactor is placed on the inoculation of uncapping on the super clean bench, root of hair evenly is connected on each layer filter, starts spraying gun, atomising head produces ultrasonic wave makes liquid level be positioned at the nutrient liquid atomizing of pipe core below, nutrient mist is risen by pipe core, enter each interlayer through center pore and top exit, and descend and be full of whole reactor, offer root of hair, the long required nutrient of blastogenesis along the interannular between reactor wall and the pipe core.Atomizing is intermittently carried out, and feeds sterile air between twice atomizing, to satisfy root of hair and the bud needs to oxygen.This reactor has solved root of hair and has grown thickly bud uniform distribution and growth in reactor, utilizing the ultrasonic atomizatio principle to produce nutrient mist can spread rapidly and be evenly distributed, intermittently atomizing and air feed make and support abundance in the reactor, and have avoided in the submergence reactor because plant organ is organized vitrifying and the deformityization phenomenon of being brought by soaked with liquid for a long time.But the still unresolved classification inoculation apparatus of atomizing reactor makes that inoculation is not easy to operate, easily microbiological contamination.Because no internal light source, light distributes and light intensity is affected.Because droplet is tiny and intermittently produce mist, it is poor that the organ surface nutrient film upgrades, and makes new nutrition supply deficiency.The nutritive medium of gas entrainment simultaneously droplet is easily discharged from venting port, causes the loss of nutritive medium.Be restricted difficult amplification of feasible atomizing reactor owing to produce the mist height again to carry out the large scale culturing of plant tissue organ.
Above-mentioned present existing airlift reactor is compared with the reactor of other type with the atomizing reactor, though be more suitable in the cultivation of vegetable cell, histoorgan, but still have many problem and shortcomings that influence large scale culturing and improve useful secondary metabolite.And existing bio-reactor specificity when being used for vegetable cell, histoorgan cultivation is stronger, and range of application is restricted.Therefore, when concrete the application, must be improved it, could be adapted to the requirement of large scale culturing better at vegetable cell, histoorgan growth metabolism characteristic.
The objective of the invention is to: the shortcoming and defect that overcomes above-mentioned prior art, for realizing the even inoculation of plant tissue, organ, dirt length is evenly distributed, have good mixing, mass transfer, heat transfer characteristic, overcome the vitrifying of plant tissue, the long-term submerged culture generation of organ and the phenomenon of variation simultaneously, and realize that gas-liquid replaces the adjusting and the control in half submergence cycle, periodicity of illumination and intensity are adjustable, thereby the air lift type half submergence illumination plant cell, the histoorgan that provide a kind of suitable scale operation to use are cultivated reactor.
The object of the present invention is achieved like this: it is by reactor body that air lift type cycle submergence illumination plant cell provided by the invention, histoorgan are cultivated reactor, light source, the stainless steel interlayer, inoculated tube, the gas distributor that have stainless steel mesh are formed, and it is characterized in that also comprising reservoir, magnetic valve, gas-liquid circulation tube, gas-liquid distribution piping.Multilayer stainless steel mesh interlayer wherein is installed in reactor body, a centre hole is arranged on the interlayer, built in light is looked after from the hole and is inserted, and dividing plate is fixed thereon, and the interlayer spacing determines according to culture.The side of pipe core has the inoculated tube patchhole, flexibly connects the turnover panel that has stainless steel mesh in a lid hole on the hole, and a segmentation inoculated tube passes this hole, and every section inoculated tube screws togather fixing; Reservoir be installed in reactor body below, by a pipe gas distributor of the gas distributor in the reservoir with magnetic valve and reactor body inner bottom part is connected, another stainless steel tube is communicated with the reactor body bottom by a magnetic valve with the reservoir top.Another sidewall of reservoir picks out a gas-liquid circulation tube and is communicated on the interior gas-liquid distribution piping of reactor body.Cover on the reactor body and insert a vapor pipe; Light source is placed on during built in light looks after.The gas-liquid distribution piping is in reactor body between each interlayer.Have the through hole of different diameter and number according to different depths, be beneficial to the uniformly distributing of gas with various and nutritive medium.
The cultural method of employing air lift type cycle submergence illumination plant cell tissue provided by the invention:
1. feed vapor sterilization after gas, water-cooled examination, sterilising temp is 100-121 ℃, and sterilization time is 40-200 minute, feeds the sterile air protection again.
2. at room temperature open inoculated tube top sealing cap, in gnotobasis, culture seed (vegetable cell, tissue, organ etc.) is successively added, finish one deck inoculation back and improve one section inoculated tube and take off, new one deck flap falls automatically and inoculates, and finishes inoculation successively from low to high.
3. build sealing cap, the input nutrient solution
4. by the temperature required temperature controller that mixes up
5. Immersion time and the ventilation cycle by required culture requirement mixes up the sequential instrument, starts native system, realizes automatically cultivating according to following conditional request.By the required Immersion time 3-60 of the requirement of culture minute, non-Immersion time 30-100 minute, container for storing liquid aeration-agitation time 1-20 minute, 10-20 minute loading and unloading time, light application time 8-24 hour, intensity of illumination 500lux-5000lux.
Culture condition is adjustable in the culturing process, and different cultures need different nutrient solutions and culture condition.
The invention has the advantages that: Airlift cyclic immersion lighting plant cell tissue organ reactor of the present invention is a kind of new-type bioreactor of large scale culturing, can the submergence of property performance period, ventilation, periodically illumination and stop illumination and unglazed cultivation operation, can realize the even inoculation of plant tissue organ, growth distribution is even, have good mixing mass transfer, heat transfer characteristic, do not have mechanical moving parts, simple in structure, easily sealing is difficult for microbiological contamination and is easy to be amplified to industrially scalable.
Cultural method of the present invention has solved the inoculation inequality that occurs in present vegetable cell, tissue and the organ culture, long-term submerged culture generation vitrifying and variation phenomenon, atomizing is cultivated and is difficult for amplifying, gas-liquid-solid three-phase mixing and mass transfer are poor, illumination is difficult for regulating, growth the gas-liquid short circuit occurs at the later stage, temperature distributing disproportionation and this type of reactor specificity was stronger in the past, problems such as function singleness.Invention makes inoculation simple to operate by segmentation inoculated tube and the design of interlayer turnover panel, and inoculation evenly.Gas-liquid alternately half submergence periodic adjustment is cultivated, and meets the biological growth metabolic rhythm, has strengthened the homogeneity of mass transfer, and intensity of illumination and cycle are adjustable realizes that illumination-unglazed cycle of shining replaces cultivation and dark cultivation.Nutrition and gas-liquid uniform mixing have further been strengthened in the design of container for storing liquid, have increased the culture dissolved oxygen, have realized the periodical operation that gas-liquid is alternately switched in the reactor.Simultaneously, gas temperature can be regulated by cultivating optimum temps.Therefore culture systems of the present invention has multifunctionality, vegetable cell, histoorgan growth and metabolism Modulatory character, and environmental factor is adjustable and best selective, is easy to characteristics such as amplification, is the innovation and the breakthrough of present plant tissue organ culture system.The present invention will be described in detail below in conjunction with drawings and Examples:
Fig. 1 is that a kind of existing 9 liter plant rootings cultivation reactor synoptic diagram Fig. 2 is that circulation ultrasonic atomizatio reactor synoptic diagram Fig. 3 is that structural representation Fig. 4 of the present invention is an air lift type cycle submergence illumination plant cell of the present invention in a kind of existing 2.5 liters, the structural representation drawing that histoorgan is cultivated a kind of embodiment of reactor is described as follows: 1-reactor body 2-segmentation inoculated tube 3-interlayer 4-turnover panel 5-built in light is looked after 6- gas distributor 7,8,10, the external light source 12-of 11-magnetic valve 9-reservoir 13-gas, liquid distribution piping 14-gas, liquid circulation tube 15-glass support pipe 16-spraying gun 17-atomising head 18-perforated base pipe 19-well heater 20-center pore 21-condenser 25-inoculation 26-stainless steel filter 27-venting port 28-nylon wire pipe core 29-tensimeter 30-inlet pipe 31-temperature controller 32-pH probe and controller 33-dissolved oxygen probe and controller 34-tail gas gas chromatographic detection 35-sequential control instrument 36-gas temperature regulating device 37-fluid infusion jar 38-gas meter 39-sterilizing filter 40-safety valve 41-peristaltic pump 42-acid solution storage tank 43-alkali lye storage tank
Embodiment 1:
Air lift type half submergence illumination plant cell provided by the invention, histoorgan is cultivated reactor, reactor body 1 by the manufacturing of φ 230 * 1400 thermal glasses, the stainless steel interlayer and the screen cloth 3 that will have 10 layers of φ 204 of turnover panel 4 pass through stainless steel gas-liquid distribution piping 13, insert in the reactor body 1 after fixing, be on each layer stainless steel interlayer of 10cm for inserting stainless steel segmentation inoculated tube 2 in the hole of φ 42 by spacing, screen cloth center drilling on each layer stainless steel separator, insert φ 40 thermal glass built in light through this hole and look after 5, fluorescent lamp inserts built in light and looks after in 5 as built-in light source.Reactor body 1 bottom is provided with gas distributor 6, and after passing reactor body 1 bottom with stainless steel tube gas distributor is connected with commercially available magnetic valve 7.Be positioned at reactor lower part, the stainless steel reservoir 12 of setting-φ 408 * 408 is installed a gas distributor 6, one stainless steel tubes and is connected with magnetic valve 11 by reservoir 12 welded top one stainless steel tube, and then is communicated with reactor bottom in the reservoir 12.Another stainless steel tube connects with magnetic valve 10, and is communicated on the gas-liquid circulation tube 14.Gas, liquid circulation tube 14 are communicated with two the gas-liquid distribution pipings 13 that stretch in the reactor body 1 above reactor body 1.Gas-liquid distribution piping 13 passes stainless steel interlayer in the reactor body 1 and fixes, and the space segment between each layer has the through hole of different diameter and number according to different depths, is beneficial to the uniformly distributing of different interlayers and nutritive medium and gas.Embodiment 2:
Make the air lift type cycle submergence illumination plant cell tissue reactor of one 40 liter by Fig. 4, look after 5 by the thermal glass center light, thermal glass reactor body 1, the stainless steel interlayer 3 and the turnover panel 4 of band screen cloth, stainless steel gas, liquid distribution piping 13 has fluorescent tube to make external light source 9 outside reactor body, well heater, temp probe and temperature controller 31, stainless steel gas sparger 6, stainless steel magnetic valve 7,8,10,11, stainless steel reservoir 12, pH probe and controller 32, dissolved oxygen probe and controller 33, tail gas gas chromatographic detection 34, sequential control instrument 35, stainless steel segmentation inoculated tube 2, gas temperature regulating device 36, tail gas condenser 21, fluid infusion jar 37, gas meter 38, sterilizing filter 39, tensimeter 29, safety valve 40, peristaltic pump 41, acid solution storage tank 42, alkali lye storage tank 43 is formed; Multilayer stainless steel mesh interlayer 3 wherein is installed in reactor body, and porose on the interlayer 3, built in light is looked after from the hole and is inserted, and dividing plate is fixed thereon, and the interlayer spacing determines according to culture.The side of center pore 20 has inoculated tube 2 patchholes, and the turnover panel that has stainless steel mesh 4, the one segmentation inoculated tubes that flexibly connect a lid hole on the hole pass this hole, and every section inoculated tube screws togather fixing; Reservoir 12 be installed in reactor body below, by a pipe gas distributor 6 in the reservoir 12 is connected with the gas distributor 6 of magnetic valve and reactor body inner bottom part, another stainless steel tube is communicated with the reactor body bottom by a magnetic valve with reservoir 12 tops.Reservoir 12 another sidewalls pick out a gas-liquid circulation tube 14 and are communicated on the gas-liquid distribution piping 13 in the reactor body 1.Cover on the reactor body 1 and insert a vapor pipe; Light source is placed on built in light and looks after in 5.Gas-liquid distribution piping 13 is each interlayer 3 in reactor body 1, and wherein built in light is looked after 5 and inserted from the hole, and dividing plate is fixed thereon, and interlayer 3 spacings determine according to culture.The side of center pore 20 has inoculated tube 2 patchholes, and the turnover panel that has stainless steel mesh 4, the one segmentation inoculated tubes 2 that flexibly connect a lid hole on the hole pass this hole, and every section inoculated tube 2 screws togather fixing; Reservoir be installed in reactor body below, by a pipe gas distributor 6 in the reservoir 12 is connected with the gas distributor 6 of magnetic valve and reactor body inner bottom part, another stainless steel tube is communicated with the reactor body bottom by a magnetic valve with the reservoir top.Reservoir 12 another sidewalls pick out a gas, and wherein internal light source 5 can be regulated light intensity and periodicity of illumination with external light source 9 in reactor body 1.Segmentation inoculated tube 2, interlayer 3, screen cloth 3 and turnover panel 4 are easy to inoculation and seed uniform distribution in reactor, avoid the culture local accumulation. Magnetic valve 7,8,10,11 combine with sequential control instrument 35 and can realize that gas-liquid cycle alternately immerse cultivates.Heating and control when well heater, temp probe and temperature controller 31 can be realized liquid-immersed the cultivation.Gas temperature adjustment temperature regulating device 36 can be controlled at gas temperature required, and gas flows to 38 adjustments of gas flows after sterilizing filter 39 degerming.Gas distributor 6 can be realized gas uniform distribution in solution.Gas, liquid distribution piping 13 are transported to each layer of reactor with gas and nutrient solution.Entrapped moisture in the tail gas condenser 21 condensable tail gas.Fluid infusion jar 37 can be added loss moisture.Reactor body and container for storing liquid setting pressure table 29 and safety valve 40 guarantee operational safety.Embodiment 3:
In 40 liters reactor assembly of the present invention, use method of the present invention, the Agrobacterium rhizogenes R1601 that Chinese Academy of Sciences plant is provided induces sweet wormwood blade formation plant organ hairly root to carry out large scale culturing, inoculum size be 0.1-1 gram dry weight/liter, half submergence cycle was: submergence in 3-10 minute, non-submergence in 30-60 minute and ventilation.Intensity of illumination 500lux-5000lux, periodicity of illumination: illumination every day 8-20 hour, temperature 20-35 ℃, cultivated 20-30 days, whole reactor covers with root of hair, obtain the root of hair dry weight and be 5-30 gram dry weight/liter, biomass increases 30-60 doubly.Operation steps: embodiment 4:
In 40 liter scales reactor assembly of the present invention, the sweet wormwood that Chinese Academy of Sciences plant the is provided bud of growing thickly carries out large scale culturing, inoculum size be 0.1-0.6 gram dry weight/liter, submergence cycle, intensity of illumination, periodicity of illumination and temperature are with example 1.Cultivate after 20-30 days, whole reactor covers with the bud of growing thickly, obtain sweet wormwood grow thickly shoot dry matter be 5-30 gram dry weight/liter, biomass increases 15-20 doubly, the gained biomass is respectively solid culture and shake-flask culture 1.7 times and 2.5 times.
The inventive method and cultivate reactor and realized that sweet wormwood organ hairly root, the sweet wormwood bud of growing thickly is periodically submerged culture.Well-grown in the test, the reactor effective volume makes full use of, and culture does not have vitrification phenomenon and variation takes place, the whole reactor good seal, no microbiological contamination phenomenon, culture, uniformity of temperature profile in the reactor, mass transfer is good between gas-liquid, periodically carries liquids and gases.PH, temperature, dissolved oxygen can monitor and regulate and control to optimum range, and the total system operation is reliable.Domestic 40 liters of plant tissues, the organ large scale culturing of design are first succeedd first.
Claims (10)
1. Airlift cyclic immersion lighting plant cell tissue organ is cultivated reactor, comprises reactor body, light source, and interlayer, inoculated tube, gas distributor is characterized in that also comprising reservoir, magnetic valve, gas-liquid circulation tube, gas-liquid distribution piping.The interlayers of many band screen clothes wherein are installed in reactor body, and interlayer is provided with a hole, built in light look after from the hole, insert fixed thereon.The interlayer side has the inoculated tube patchhole, flexibly connects the turnover panel that has stainless steel mesh in a lid hole on the limit, hole, and a segmentation inoculated tube passes the hole of an interlayer, and every section inoculated tube passes interlayer and the hypomere inoculated tube screws togather fixing; Reservoir be installed in reactor body below, by a pipe gas distributor of the gas distributor in the reservoir with magnetic valve and reactor body inner bottom part is connected, another stainless steel tube is communicated with the reactor body bottom by a magnetic valve with the reservoir top.Another sidewall of reservoir picks out a gas-liquid circulation tube and is communicated on the interior gas-liquid distribution piping of reactor body.Cover on the reactor body and insert a vapor pipe; Light source is placed on during built in light looks after.The gas-liquid distribution piping has the through hole of different diameter and number according to different depths in reactor body between each interlayer, is beneficial to the uniformly distributing of nutritive medium and gas in the different interlayers.
2. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: the outside that also is included in reactor body is provided with external illumination pipe.
3. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: comprise that also magnetic valve is connected with the sequential control instrument, the sequential control instrument is electrically connected with built-in light source.
4. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: the gas/liquid distribution piping has improved the late stage of culture ventilation and nutrition is carried the problem of discharging, prevented gas-liquid short circuit as growth later stage appearance such as hairly root, the mass transfer inequality, problems such as temperature distributing disproportionation.
5. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: in half submergence circulation culturing process, guaranteed the thorough mixing of nutritive medium and enough dissolved oxygens etc., for the even growth of culture in the reactor provides condition with reservoir.
6. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: also be included in reactor body peripheral hardware one gas constant temperature device.
7. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: having one to live and connect on the interlayer, in the middle of the turnover panel is screen cloth, and turnover panel, screen cloth are that stainless material is made.
8. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: do not have mechanical moving parts, operation has automatically reduced microbiological contamination chance in the cultivation.
9. cultivate reactor by the described Airlift cyclic immersion lighting plant cell tissue organ of claim 1, it is characterized in that: also be included in the reactor body bottom well heater is set, and be electrically connected with temp probe and temperature controller, temperature controller is electrically connected with the sequential control instrument again.
10. the described Airlift cyclic immersion lighting plant cell tissue organ of a claim 1 using method of cultivating reactor, it is characterized in that: (1) feeds vapor sterilization after gas, water-cooled examination, sterilising temp is 100-121 ℃, sterilization time is 40-2.00 minute, feeds the sterile air protection again;
(2). at room temperature open inoculated tube top sealing cap, in gnotobasis, culture seed (vegetable cell, tissue, organ etc.) is successively added, finish one deck inoculation back and improve one section inoculated tube and take off, new one deck flap falls automatically and inoculates, and finishes inoculation successively from low to high;
(3). build sealing cap, the input nutrient solution;
(4) by the temperature required temperature controller that mixes up;
(5) Immersion time and the ventilation cycle by required culture requirement mixes up the sequential instrument, starts native system, realizes automatically cultivating according to following conditional request.By the required Immersion time 3-60 of the requirement of culture minute, non-Immersion time 30-100 minute, container for storing liquid aeration-agitation time 1-20 minute, 10-20 minute loading and unloading time, light application time 8-24 hour, intensity of illumination 500lux-5000lux.
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| CN98102396A CN1063224C (en) | 1998-06-16 | 1998-06-16 | Airlift cyclic immersion lighting plant cell tissue organ culture method and culture reactor |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006122498A1 (en) * | 2005-05-20 | 2006-11-23 | Tsinghua University | Airlift circumfluent reactor needing no foreign air source |
| CN101953296B (en) * | 2009-07-17 | 2012-07-25 | 中国科学院过程工程研究所 | Pressure pulsation ultrasonic atomization culture device and cultural method using same |
| CN105210172A (en) * | 2012-12-31 | 2015-12-30 | 太阳能爱迪生半导体有限公司 | Processes and apparatus for preparing heterostructures with reduced strain by radial compression |
| CN111757927A (en) * | 2017-12-22 | 2020-10-09 | 爱沃逻辑克技术有限责任公司 | Bioreactor for root organ cultures |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| US11299700B1 (en) | 2021-02-19 | 2022-04-12 | Acequia Biotechnology, Llc | Bioreactor containers and methods of growing hairy roots using the same |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| FR2621323B1 (en) * | 1987-10-02 | 1990-06-15 | Commissariat Energie Atomique | INTENSIVE AND CONTROLLED PRODUCTION DEVICE FOR MICROORGANISMS BY PHOTOSYNTHESIS |
| CN2252818Y (en) * | 1996-04-16 | 1997-04-23 | 中国科学院化工冶金研究所 | Plant tissue culture reactor comprising of elastic valve sieve plate |
| CN1051575C (en) * | 1996-05-15 | 2000-04-19 | 中国科学院化工冶金研究所 | Reactor for nutrient liquid atomizing culture of plant |
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1998
- 1998-06-16 CN CN98102396A patent/CN1063224C/en not_active Expired - Fee Related
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006122498A1 (en) * | 2005-05-20 | 2006-11-23 | Tsinghua University | Airlift circumfluent reactor needing no foreign air source |
| AU2006246862B2 (en) * | 2005-05-20 | 2011-04-21 | Tsinghua University | Airlift circumfluent reactor needing no foreign air source |
| CN101953296B (en) * | 2009-07-17 | 2012-07-25 | 中国科学院过程工程研究所 | Pressure pulsation ultrasonic atomization culture device and cultural method using same |
| CN105210172A (en) * | 2012-12-31 | 2015-12-30 | 太阳能爱迪生半导体有限公司 | Processes and apparatus for preparing heterostructures with reduced strain by radial compression |
| CN111757927A (en) * | 2017-12-22 | 2020-10-09 | 爱沃逻辑克技术有限责任公司 | Bioreactor for root organ cultures |
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| CN1063224C (en) | 2001-03-14 |
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