CN101845434A - Method for rapidly screening thermophilic anaerobic ethanol microbe high-yield ethanol bacterial strain - Google Patents

Method for rapidly screening thermophilic anaerobic ethanol microbe high-yield ethanol bacterial strain Download PDF

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CN101845434A
CN101845434A CN201010149123A CN201010149123A CN101845434A CN 101845434 A CN101845434 A CN 101845434A CN 201010149123 A CN201010149123 A CN 201010149123A CN 201010149123 A CN201010149123 A CN 201010149123A CN 101845434 A CN101845434 A CN 101845434A
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ethanol
bacterial strain
thermophilic anaerobic
concentration
microbe
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CN101845434B (en
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籍月彤
宋厚辉
徐健
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Qingdao Institute of Bioenergy and Bioprocess Technology of CAS
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Qingdao Institute of Bioenergy and Bioprocess Technology of CAS
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Abstract

The invention relates to a method for rapidly screening thermophilic anaerobic ethanol microbe high-yield ethanol bacterial strain, comprising the following steps: 1) wild strains are cultured continuously in a fluid nutrient medium containing ethanol, the ethanol tolerance of the thermophilic anaerobic ethanol microbe is evolved directionally; 2) ethylmethylsulfone mutagenesis is carried out to ethanol-tolerance bacterial strains, and the mutagenesis bacterium liquid is coated on the solid culture medium containing the ethanol; 3) after individual bacterial colony is formed, the individual bacterial colony with large radius is dropwise added in 2,3,5-triphenyl chlorination tetrazole developing liquid, and single spots with deep color development are taken for re-screening on a CaCO3 rescreening plate containing high substrate concentration; 4) bacterial colonies with large radius and small radius of a transparent ring, pH indicators are dropwise added for eliminating false positive; 5) the ethanol concentration is tested by a gas chromatography or an ethanol quantitative kit after fermentation is carried out, and the bacterial strains with high ethanol output are cultured for at least 20 generations under a non-selection condition, and the bacterial strain which still has high yield is destination bacterial strain.

Description

The method of rapidly screening thermophilic anaerobic ethanol microbe high-yield ethanol bacterial strain
Technical field
The invention belongs to the sudden change and the chemomorphosis screening field of specified microorganisms physiological character, specifically be the means that adopt orthogenesis and chemomorphosis to combine, thermophilic anaerobic ethanol microbe is suddenlyd change, by color reaction, the rapid screening high-yield ethanol bacterial strain.
Background technology
Fast development along with global economy, oil, coal, Nonrenewable energy resources such as Sweet natural gas are exhausted day by day, and the destruction that a large amount of greenhouse gases of burning fossil energy discharging are serious ecotope ([Cifuentes, L., Borja-Aburto, V.H., Gouveia, N., Thurston, G., Davis, D.L.2001.Climate change.Hidden health benefits of greenhouse gas mitigation.Science 293,1257-1259.], [Lashof, D.A., Ahuja, D.R.1990.RelativeContributions of Greenhouse Gas Emissions to Global Warming.Nature 344,529-531.]), the renewable energy source of green low-carbon is had to make great efforts to develop and utilize in countries in the world.Bio-ethanol is as a kind of novel clean energy ([Hahn-Hagerdal, B., Galbe, M., Gorwa-Grauslund, M.F., Liden, G., Zacchi, G.2006.Bio-ethanol--the fuel oftomorrow from the residues of today.Trends Biotechnol 24,549-556.]), just progressively add in the gasoline, reduced the dependence of people to a great extent fossil energy as power fuel.The U.S. and Brazil are the majoies country of production of bio-ethanol, owing to use corn, food crop such as sugarcane are as raw material, not only there is the problem of striving grain with the people, also need large-area deforestation to open up wasteland and plant starting material ([Timothy Searchinger, R.H., R.A.Houghton, Fengxia Dong.AmaniElobeid, Jacinto Fabiosa, Simla Tokgoz, Dermot Hayes, Tun-Hsiang Yu..2008.Use of U.S.Croplands for biofuels increases greenhouse gases throughemissions from land-use change..Science.319,1238-1240.]), after research was in a few years explored, people recognized that gradually must greatly develop with non-food crop is the bio-ethanol of raw material.
Lignocellulose is the abundantest in the world renewable raw materials, cellulosic ethanol has utilized these agriculture and forestry organic waste materials that contain Mierocrystalline cellulose, hemicellulose to come fermentative production of ethanol ([Lynd just, L.R.1996.Overview and evaluation of fuel ethanol from cellulosic biomass:Technology, economics, the environment, and policy.Annual Review of Energy and theEnvironment 21,403-465.]).But the subject matter of running in industrial application at present is that cost is too high, tracing it to its cause, to be that the course of processing is loaded down with trivial details cause, need be monose with cellulose degradation at first promptly, could utilize production of ethanol from microbial fermentation such as yeast then by physics, chemistry or biological means.Thereby, must shorten the production cycle if cellulosic hydrolytic process and sugar-fermenting process conformity can be in the same place, reduce production costs.Associating biological processing (Consolidated Bioprocessing, CBP) technology has solved this difficult problem ([Lynd, L.R., van Zyl, W.H., McBride, J.E., Laser, M.2005.Consolidated bioprocessing of cellulosic biomass:an update.CurrOpin Biotechnol 16,577-583.]), promptly under the condition of high temperature anaerobic, adopt one or more microorganism mixed fermentation, fermentative production of ethanol when degraded cellulose produces reducing sugar can be an ethanol with cellulose conversion once going on foot.But it is too low that the present bottleneck of this method is an ethanol production, can not put into production.The mutagenesis screening high-yield ethanol bacterial strain is to address this problem the most direct effective way.
The comparatively general screening method of high-yield ethanol bacterial strain is directly to screen ethanol tolerance bacterial strain at present, screening as saccharomycetic high-yield ethanol bacterial strain, usually be applied on the high concentration ethanol flat board after adopting chemistry or ultraviolet mutagenesis, ([(Hou Lihua, horse all one's life to select the clone that can tolerate high concentration ethanol then.2007, a kind of method that makes up the yeast saccharomyces cerevisiae ethanol high-yield bacterial strain.CN 101323837A.]), therefrom screen high-yield ethanol bacterial strain again.This method workload is big, length consuming time, screening efficiency is low, and be not suitable for thermophilic anaerobic ethanol microbe, reason is that alcohol resistance is the principal element that the restriction yeast produces alcohol, but for thermophilic anaerobic ethanol microbe, as thermophilic anaerobic ethanol bacillus ([Wiegel J., L.L.G.1981.Thermoanaerobacter ethanolicus gen.nov., spec.nov., a new, extreme thennophilic, anaerobic bacterium.Arch Microbiol.128,343-348.]), the product kind of glycolytic pathway is a lot of in its born of the same parents, in low concentration of substrate, main tunning is an ethanol, but in a single day concentration of substrate surpasses 1% (w/v), and tunning is then turned to by products such as acetate and lactic acid by ethanol.This shows that the accumulation of by product is the key factor of restriction ethanol production during high concentration of substrate.For this reason, it is dull and stereotyped that we have designed following two screenings: ethanol-TTC (2,3,5-triphenyl tetrazolium chloride) colour developing flat board and CaCO 3 -High sugar is dull and stereotyped, to screen ethanol high-yield bacterial strain efficiently.(Alcohol dehydrogenase ADH) is key enzyme in the ethanol generation approach to ethanol dehydrogenase, and its active height directly influences the output of product, and TTC and ADH reaction generate red material, and its shade is directly proportional with the ADH activity.Dull and stereotyped can the filtering out of ethanol-TTC colour developing has certain alcohol resistance and the active high bacterial strain of ADH.CaCO 3Can be with being secreted into born of the same parents outer by-product acetic acid, lime acetate and the calcium lactate that lactic acid generates solubility produce the CaCO of the many more consumption of acid 3Many more, CaCO 3The transparent circle that forms on the flat board is just big more, and promptly under the situation of identical colony radius, what of product acid are the size of transparent circle diameter directly reflect.CaCO 3-Gao sugar flat board can filter out and produce the few bacterial strain of acid under the high concentration of substrate, for avoiding the false positive bacterium colony, by drip the reliability that the pH indicator improves The selection result on bacterium colony.
Summary of the invention
The object of the present invention is to provide a kind of method of rapid screening ethanol high-yield bacterial strain.
Novelty of the present invention is that an ethanol adaptive evolution and chemomorphosis combine, screening ethanol tolerance bacterial strain, and on screening method, start with from producing sour approach, united the dull and stereotyped and high sugar flat board of CaCO3-of ethanol-TTC colour developing, can filter out at short notice have certain alcohol resistance, alcohol dehydrogenase enzyme activity height and under high concentration of substrate, produce the few bacterial strain of acid, and by on bacterium colony, dripping the reliability that the pH indicator improves The selection result.Thereby in the change known technology to the screening of zymic high-yield ethanol bacterial strain, be with low alcohol resistance as main restricting factor, this method workload is big, length consuming time, screening efficiency is low, and is not suitable for the defective of thermophilc anaerobe.
For achieving the above object, the method for rapidly screening thermophilic anaerobic ethanol microbe ethanol high-yield bacterial strain of the present invention provided by the invention comprises the steps:
1) with wild type strain cultured continuously in containing the alcoholic acid liquid nutrient medium, the alcohol resistance of orthogenesis thermophilic anaerobic ethanol microbe;
2) tolerance alcoholic acid bacterial strain is carried out ethylmethane sulfonate (EMS) mutagenesis, the bacterium liquid after the mutagenesis is applied to contains on the alcoholic acid solid medium;
3) behind single bacterium colony to be formed, select the big single bacterium colony of colony radius to drip 2,3,5-triphenyl tetrazolium chloride (TTC) colour developing liquid dips in aseptic toothpick and to get the saturate single spot that develops the color to the CaCO that contains high concentration of substrate 3Carry out multiple sieve on the flat board;
4) select the big but little bacterium colony of transparent circle radius of colony radius to drip the pH indicator and get rid of false positive;
5) alcohol concn is measured with gas-chromatography or ethanol quantification kit in the fermentation back, selects the high bacterial strain of ethanol production to cultivate at least 20 generations under non-selection condition, still can keep the bacterial strain of high ethanol production to be the purpose bacterial strain.
In the described method, thermophilic anaerobic ethanol microbe in the step 1 is meant that optimum growth temperature is higher than 50 ℃, strictness or amphimicrobian, there is the gram-positive microorganism that produces acid and producing and ethanol pathways metabolism, mainly comprises: thermophilic anaerobic ethanol bacillus (Thermoanaerobacter ethanolicus), Bu Shi thermophilic anaerobic bacterium (Thermoanaerobium brockii), thermophilic hydrogen sulfide clostridium (Clostridiumthermohydrosulfuricum), Clostridium thermocellum (Clostridium thermocellum) and thermophilic clostridium saccharolyticum (Clostridium thermosaccharolyticum).
In the described method, in the step 1 the tolerance final concentration of ethanol orthogenesis by volume concentration count 2%-15%.
In the described method, in the step 2 the ethylmethane sulfonate working concentration by volume concentration count 0.1%-5%.
In the described method, in the step 32,3, in the 5-triphenyl tetrazolium chloride ethanol final concentration by volume concentration count 3%-16%.
In the described method, in the step 32,3,5-triphenyl tetrazolium chloride concentration is 1-100mg/ml.
In the described method, sieve flat board in the step 3 again and contain 0.1-2g/L CaCO 3, 1-10g/L glucose.
In the described method, in the step 5 non-selection condition cultivate down at least 20 generations still can keep ethanol production by volume densitometer more than 2%.
The means that the present invention adopts orthogenesis and chemomorphosis to combine are suddenlyd change to thermophilic anaerobic ethanol microbe, by color reaction, and the rapid screening ethanol high-yield bacterial strain.Method of the present invention is applicable to that all are thermophilic, anaerobism and have the gram positive bacterium that produces acid and producing and ethanol pathways metabolism.The present invention can screen high-yield ethanol bacterial strain fast and effectively, for biological ethanol industry provides strain excellent.
Embodiment
Following embodiment can make the technician of this professional skill field more fully understand the present invention, but does not limit the present invention in any way.
Embodiment (is example with the thermophilic anaerobic ethanol bacillus)
1) orthogenesis alcohol resistance
The thermophilic anaerobic ethanol bacillus was inoculated into containing in 1% (v/v) alcoholic acid RCM substratum (prescription sees Appendix 1) of deoxygenation in 1: 10 by volume, after treating that bacterium liquid length is dense, be inoculated at 1: 10 again and contain in 1% (v/v) alcoholic acid RCM substratum, pass 5-10 repeatedly after generation; Bacterium liquid is inoculated at 1: 10 contains in 1.5% (v/v) alcoholic acid RCM substratum, pass 5-10 repeatedly after generation; Bacterium liquid is inoculated at 1: 10 contains in 2% (v/v) alcoholic acid RCM substratum, pass 5-10 repeatedly after generation; Bacterium liquid is inoculated at 1: 10 contains in 2.5% (v/v) alcoholic acid RCM substratum, pass 5-10 repeatedly after generation, bacterium liquid is coated on contains on 2.5% (v/v) alcoholic acid RCM solid medium, a picking 10-100 mono-clonal is chosen the fastest clone of the speed of growth in containing 2.5% (v/v) alcoholic acid RCM substratum.
Annex 1: the anaerobion culture medium prescription difference of different genera, RCM substratum only are fit to thermophilic anaerobic ethanol bacillus (Themroanaerobacter ethanolicus).For specific microorganism, need select corresponding substratum for use.Only need in substratum, add ethanol and get final product, down together according to above-mentioned steps.RCM substratum preparation: yeast extract paste 3g/L, extractum carnis 10g/L, peptone 10g/L, Zulkovsky starch 1g/L, glucose 5g/L, cysteine hydrochloride 0.5g/L, NaCl 3g/L, NaAc3g/L, agar 0.5g/L, resazurin 2mg/L, adding distil water mend to cumulative volume be 1L, adjusting pH value is 6.8, leads to N 2Deoxygenation, 115 ℃ of moist heat sterilization 15min.The preparation of RCM solid medium: agar 1.25%, other is with the RCM substratum.
2) EMS mutagenesis ethanol tolerance bacterial strain
(1) activatory thermophilic anaerobic ethanol bacillus ethanol tolerance bacterial strain is inoculated at 1: 100 contains 10ml among the Hungate-tube of ATCC 1190 substratum (prescription sees Appendix 2) of deoxygenation, leave standstill in 60 ℃ of incubators and be cultured to OD 600Be worth about 0.4.
The centrifugal 5min of (2) 6,000rpm collects thalline, and being resuspended in 500 μ l does not have in the ATCC1190 substratum of carbon source, gets 150 μ l bacterium liquid and is resuspended in 3ml and does not have in the ATCC1190 substratum of carbon source, leaves standstill in 60 ℃ of incubators and cultivates 2h.
(3) in stink cupboard, in bacterium liquid, add 40 μ l EMS and 100 μ l Tris-HCl (0.1M, pH7.4,1% sucrose), behind the whirlpool concussion mixing, 60 ℃ of shaking table concussions (120rpm) were cultivated 30 minutes.
(4) Na of adding 150 μ l 2M in bacterium liquid 2S 2O 3Stopped reaction, the centrifugal 20min of 6000rpm abandons supernatant, washes twice with the ATCC1190 substratum, is resuspended in the 500 μ l ATCC1190 substratum.Remarks ATCC1190 substratum preparation: KH 2PO 41.5g/L, Na 2HPO 412H 2O 4.2g/L, NH 4Cl0.5g/L, MgCl 26H 2O 0.18g/L, yeast extract 2.0g/L, glucose 5g/L, cysteine hydrochloride 0.1g/L, Na 2S 0.1g/L, vitamin solution 0.5ml/L, Wolfe ' s mineral salt solution 5.0ml/L, resazurin (0.1%) 1.0ml/L, adding distil water mend to cumulative volume be 1L, adjusting pH value is 7.3, leads to N 2Deoxygenation, 115 ℃ of moist heat sterilization 15min.
Annex 2:
Vitamin solution (500ml): Biotin 20.0mg, p-Aminobenzoic acid 50.0mg, Folicacid 20.0mg, Pantothenic acid calcium salt 50.0mg, Nicotinic acid 50.0mg, Vitamin B12 1.0mg, Thiamine HCl 5.0mg, Pyridoxine hydrochloride 100.0mg, Thioctic acid 50.0mg, Riboflavin 5.0mg.
Wolfe ' s mineral salt solution (1L): Nitrilotriacetic acid 1.5g, MgSO 47H 2O 3.0g, MnSO 4H 2O 500.0mg, NaCl 1.0g, FeSO 47H 2O 100.0mg, Co (NO 3) 26H 2O100.0mg, CaCl 2100.0mg, ZnSO 47H 2O 100.0mg, CuSO 45H 2O 10.0mg, ALK (SO 4) 210.0mg, Boric acid 10.0mg, Na 2MoO 42H 2O 10.0mg, Na 2SeO 31.0mg.
3) primary dcreening operation ethanol high-yield bacterial strain
With 1000 times of the bacterium liquid after EMS mutagenesis dilutions, get 50 μ l and be applied to and contain on 3% (v/v) alcoholic acid RCM solid plate, flat board is upside down in the anaerobic jar of anaerobic, in 60 ℃ of incubators, leave standstill cultivation.After treating the single bacterium colony of dull and stereotyped upward formation, select the big single spot of colony radius to drip the TTC solution (pH7.4,0.1M phosphate buffered saline buffer, 0.22 μ m membrane filtration degerming) of 10mg/ml, develop the color after 10 minutes, be wine-colored bacterium colony after the picking colour developing and carry out next step multiple sieve.
4) sieve ethanol high-yield bacterial strain again
Dip in aseptic toothpick and to get the scarlet bacterium colony to sieving again on the flat board, multiple sieve is dull and stereotyped for containing 0.5g/LCaCO 3With the RCM solid plate of 10g/L glucose, flat board is upside down in the anaerobic jar of anaerobic, leave standstill cultivation in 60 ℃ of incubators, after treating the single bacterium colony of dull and stereotyped upward formation, the picking colony radius is big but the little bacterium colony of transparent circle radius drips Congo red pH indicator, selects red partially single bacterium colony; Perhaps drip methyl red pH indicator, select inclined to one side xanchromatic list bacterium colony.
Congo red pH indicator prescription: take by weighing Congo red 0.5g, be dissolved in the ethanol of 100ml 10% (0.22 μ m membrane filtration degerming), color change interval is pH 3.0～5.0, and color is red by blue stain.Methyl red pH indicator prescription: take by weighing methyl red 0.1g, be dissolved in the sodium hydroxide solution of 7.4ml 0.05mol/L, thin up is to 200ml (0.22 μ m membrane filtration degerming) again, and color change interval is pH4.2～6.3, and color is by the red stain Huang.
5) product stability detects
The bacterial strain and the wild type strain that screen are carried out fermenting experiment in containing the ATCC1190 substratum of 1g/L glucose, leave standstill in 60 ℃ of incubators and cultivated 60 hours, sampling gas Chromatographic Determination alcohol concn.Ethanol production is significantly higher than 20 generations of bacterial strain cultured continuously of wild type strain, and the bacterial strain that output height and proterties are stable is ethanol high-yield bacterial strain.
Utilize aforesaid method obtaining within 1 month more than the alcohol resistance 6% (volume ratio), the bacterial strain of the high-yield ethanol that output 3% (volume ratio) is above.

Claims (8)

1. the method for a rapidly screening thermophilic anaerobic ethanol microbe high-yield ethanol bacterial strain, key step is as follows:
1) with wild type strain cultured continuously in containing the alcoholic acid liquid nutrient medium, the alcohol resistance of orthogenesis thermophilic anaerobic ethanol microbe;
2) tolerance alcoholic acid bacterial strain is carried out ethylmethane sulfonate mutation, the bacterium liquid after the mutagenesis is applied to contains on the alcoholic acid solid medium;
3) behind single bacterium colony to be formed, select the big single bacterium colony of colony radius to drip 2,3,5-triphenyl tetrazolium chloride colour developing liquid is got the saturate single spot of colour developing to the CaCO that contains high concentration of substrate 3Sieve again and carry out multiple sieve on the flat board;
4) select the big but little bacterium colony of transparent circle radius of colony radius to drip the pH indicator and get rid of false positive;
5) alcohol concn is measured with gas-chromatography or ethanol quantification kit in the fermentation back, selects the high bacterial strain of ethanol production to cultivate at least 20 generations under non-selection condition, still can keep the bacterial strain of high ethanol production to be the purpose bacterial strain.
2. method according to claim 1, wherein, thermophilic anaerobic ethanol microbe in the step 1 is meant that optimum growth temperature is higher than 50 ℃, strictness or amphimicrobian, there is the gram-positive microorganism that produces acid and producing and ethanol pathways metabolism, mainly comprises: thermophilic anaerobic ethanol bacillus (Thermoanaerobacter ethanolicus), Bu Shi thermophilic anaerobic bacterium (Thermoanaerobiumbrockii), thermophilic hydrogen sulfide clostridium (Clostridium thermohydrosulfuricum), Clostridium thermocellum (Clostridium thermocellum) and thermophilic clostridium saccharolyticum (Clostridiumthermosaccharolyticum).
3. method according to claim 1, wherein, in the step 1 the tolerance final concentration of ethanol orthogenesis by volume concentration count 2%-15%.
4. method according to claim 1, wherein, in the step 2 the ethylmethane sulfonate working concentration by volume concentration count 0.1%-5%.
5. method according to claim 1, wherein, in the step 32,3, in the 5-triphenyl tetrazolium chloride ethanol final concentration by volume concentration count 3%-16%.
6. method according to claim 5, wherein, in the step 32,3,5-triphenyl tetrazolium chloride concentration is 1-100mg/ml.
7. method according to claim 1 wherein, is sieved flat board again and is contained 0.1-2g/LCaCO in the step 3 3, 1-10g/L glucose.
8. method according to claim 1, wherein, in the step 5 non-selection condition cultivate down at least 20 generations still can keep ethanol production by volume densitometer more than 2%.
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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102399734A (en) * 2011-12-16 2012-04-04 昆明理工大学 Thermophilic anaerobic strain and application thereof
CN102808000A (en) * 2012-08-23 2012-12-05 辽宁科技大学 Recycling grading utilization and carbon sequestration treatment method for municipal solid waste
CN106957761A (en) * 2017-04-28 2017-07-18 德保县广鑫贸易有限公司 A kind of preparation method of spirulina wine
CN112210583A (en) * 2020-10-15 2021-01-12 广东中微环保生物科技有限公司 Application of TTC (time to temperature) dehydrogenase testing method in bacterium screening

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101323837A (en) * 2007-12-28 2008-12-17 天津大学 Method for creating distiller's yeast ethanol high yield bacterial strain
CN101333553A (en) * 2008-07-29 2008-12-31 东南大学 Method for producing fuel ethanol by mixed fermentation cellulosic

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101323837A (en) * 2007-12-28 2008-12-17 天津大学 Method for creating distiller's yeast ethanol high yield bacterial strain
CN101333553A (en) * 2008-07-29 2008-12-31 东南大学 Method for producing fuel ethanol by mixed fermentation cellulosic

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
YU JIANG ET AL.: "A highly efficient method for liquid and solid cultivation of the anaerobic hyperthermophilic eubacterium Thermotoga maritima", 《FEMS MICROBIOL》 *
蒋宇,邵蔚蓝: "嗜热厌氧产乙醇杆菌乙醇代谢途径的初步研究", 《南京师大学报(自然科学版)》 *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102399734A (en) * 2011-12-16 2012-04-04 昆明理工大学 Thermophilic anaerobic strain and application thereof
CN102808000A (en) * 2012-08-23 2012-12-05 辽宁科技大学 Recycling grading utilization and carbon sequestration treatment method for municipal solid waste
CN102808000B (en) * 2012-08-23 2014-06-25 辽宁科技大学 Recycling grading utilization and carbon sequestration treatment method for municipal solid waste
CN106957761A (en) * 2017-04-28 2017-07-18 德保县广鑫贸易有限公司 A kind of preparation method of spirulina wine
CN112210583A (en) * 2020-10-15 2021-01-12 广东中微环保生物科技有限公司 Application of TTC (time to temperature) dehydrogenase testing method in bacterium screening

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