CN108587987A - One plant of microbe oil production bacterium W-Y6 and its application - Google Patents
One plant of microbe oil production bacterium W-Y6 and its application Download PDFInfo
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- CN108587987A CN108587987A CN201810811820.7A CN201810811820A CN108587987A CN 108587987 A CN108587987 A CN 108587987A CN 201810811820 A CN201810811820 A CN 201810811820A CN 108587987 A CN108587987 A CN 108587987A
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- oil
- georgenia
- microbial inoculum
- microbe
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- 238000004519 manufacturing process Methods 0.000 title claims abstract description 18
- 241000894006 Bacteria Species 0.000 title abstract description 25
- 241000082871 Georgenia sp. Species 0.000 claims abstract description 28
- 244000005700 microbiome Species 0.000 claims abstract description 11
- 239000003921 oil Substances 0.000 claims description 57
- 239000002068 microbial inoculum Substances 0.000 claims description 26
- 239000010779 crude oil Substances 0.000 claims description 25
- 238000011084 recovery Methods 0.000 claims description 16
- 239000003208 petroleum Substances 0.000 claims description 15
- 238000000034 method Methods 0.000 claims description 8
- 238000004321 preservation Methods 0.000 claims description 8
- 239000007791 liquid phase Substances 0.000 claims description 2
- 239000004575 stone Substances 0.000 claims description 2
- 230000006872 improvement Effects 0.000 abstract description 3
- 235000019198 oils Nutrition 0.000 description 48
- 230000001580 bacterial effect Effects 0.000 description 27
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 24
- 239000007788 liquid Substances 0.000 description 10
- 239000002609 medium Substances 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 8
- 239000000047 product Substances 0.000 description 8
- 239000007787 solid Substances 0.000 description 8
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 7
- 239000008367 deionised water Substances 0.000 description 7
- 229910021641 deionized water Inorganic materials 0.000 description 7
- 230000000813 microbial effect Effects 0.000 description 7
- 239000000843 powder Substances 0.000 description 7
- 230000015556 catabolic process Effects 0.000 description 6
- 238000006731 degradation reaction Methods 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 108020004465 16S ribosomal RNA Proteins 0.000 description 5
- 238000004458 analytical method Methods 0.000 description 5
- 238000006073 displacement reaction Methods 0.000 description 5
- 239000012071 phase Substances 0.000 description 5
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Inorganic materials [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 description 5
- 239000001888 Peptone Substances 0.000 description 4
- 108010080698 Peptones Proteins 0.000 description 4
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 4
- 229910000396 dipotassium phosphate Inorganic materials 0.000 description 4
- 238000011081 inoculation Methods 0.000 description 4
- 230000007483 microbial process Effects 0.000 description 4
- 235000019319 peptone Nutrition 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 230000001954 sterilising effect Effects 0.000 description 4
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 230000035699 permeability Effects 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 239000002689 soil Substances 0.000 description 3
- 229910017677 NH4H2 Inorganic materials 0.000 description 2
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 239000000470 constituent Substances 0.000 description 2
- DIOQZVSQGTUSAI-UHFFFAOYSA-N decane Chemical compound CCCCCCCCCC DIOQZVSQGTUSAI-UHFFFAOYSA-N 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- AIUDWMLXCFRVDR-UHFFFAOYSA-N dimethyl 2-(3-ethyl-3-methylpentyl)propanedioate Chemical class CCC(C)(CC)CCC(C(=O)OC)C(=O)OC AIUDWMLXCFRVDR-UHFFFAOYSA-N 0.000 description 2
- SNRUBQQJIBEYMU-UHFFFAOYSA-N dodecane Chemical compound CCCCCCCCCCCC SNRUBQQJIBEYMU-UHFFFAOYSA-N 0.000 description 2
- 239000002552 dosage form Substances 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 239000000706 filtrate Substances 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 229910052500 inorganic mineral Inorganic materials 0.000 description 2
- 229910017053 inorganic salt Inorganic materials 0.000 description 2
- -1 liquor Substances 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 239000011707 mineral Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000000877 morphologic effect Effects 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- 229940066779 peptones Drugs 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 239000004576 sand Substances 0.000 description 2
- 239000005418 vegetable material Substances 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000726221 Gemma Species 0.000 description 1
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 235000010627 Phaseolus vulgaris Nutrition 0.000 description 1
- 244000046052 Phaseolus vulgaris Species 0.000 description 1
- 239000004372 Polyvinyl alcohol Substances 0.000 description 1
- 239000006004 Quartz sand Substances 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 229910021536 Zeolite Inorganic materials 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- LFVGISIMTYGQHF-UHFFFAOYSA-N ammonium dihydrogen phosphate Chemical compound [NH4+].OP(O)([O-])=O LFVGISIMTYGQHF-UHFFFAOYSA-N 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 235000013339 cereals Nutrition 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000004927 clay Substances 0.000 description 1
- 229910052570 clay Inorganic materials 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000002242 deionisation method Methods 0.000 description 1
- GUJOJGAPFQRJSV-UHFFFAOYSA-N dialuminum;dioxosilane;oxygen(2-);hydrate Chemical compound O.[O-2].[O-2].[O-2].[Al+3].[Al+3].O=[Si]=O.O=[Si]=O.O=[Si]=O.O=[Si]=O GUJOJGAPFQRJSV-UHFFFAOYSA-N 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- HNPSIPDUKPIQMN-UHFFFAOYSA-N dioxosilane;oxo(oxoalumanyloxy)alumane Chemical compound O=[Si]=O.O=[Al]O[Al]=O HNPSIPDUKPIQMN-UHFFFAOYSA-N 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 235000013312 flour Nutrition 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- NLYAJNPCOHFWQQ-UHFFFAOYSA-N kaolin Chemical compound O.O.O=[Al]O[Si](=O)O[Si](=O)O[Al]=O NLYAJNPCOHFWQQ-UHFFFAOYSA-N 0.000 description 1
- 229910052622 kaolinite Inorganic materials 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 229910052901 montmorillonite Inorganic materials 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 239000003129 oil well Substances 0.000 description 1
- 239000003002 pH adjusting agent Substances 0.000 description 1
- 239000003209 petroleum derivative Substances 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000012113 quantitative test Methods 0.000 description 1
- 238000007670 refining Methods 0.000 description 1
- 230000008439 repair process Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 239000000375 suspending agent Substances 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 235000015112 vegetable and seed oil Nutrition 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 239000004563 wettable powder Substances 0.000 description 1
- 239000010457 zeolite Substances 0.000 description 1
Classifications
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- 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
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- C—CHEMISTRY; METALLURGY
- C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
- C10G—CRACKING HYDROCARBON OILS; PRODUCTION OF LIQUID HYDROCARBON MIXTURES, e.g. BY DESTRUCTIVE HYDROGENATION, OLIGOMERISATION, POLYMERISATION; RECOVERY OF HYDROCARBON OILS FROM OIL-SHALE, OIL-SAND, OR GASES; REFINING MIXTURES MAINLY CONSISTING OF HYDROCARBONS; REFORMING OF NAPHTHA; MINERAL WAXES
- C10G31/00—Refining of hydrocarbon oils, in the absence of hydrogen, by methods not otherwise provided for
-
- 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
-
- E—FIXED CONSTRUCTIONS
- E21—EARTH OR ROCK DRILLING; MINING
- E21B—EARTH OR ROCK DRILLING; OBTAINING OIL, GAS, WATER, SOLUBLE OR MELTABLE MATERIALS OR A SLURRY OF MINERALS FROM WELLS
- E21B43/00—Methods or apparatus for obtaining oil, gas, water, soluble or meltable materials or a slurry of minerals from wells
- E21B43/16—Enhanced recovery methods for obtaining hydrocarbons
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- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Virology (AREA)
- Geology (AREA)
- Medicinal Chemistry (AREA)
- Mining & Mineral Resources (AREA)
- Biomedical Technology (AREA)
- Environmental & Geological Engineering (AREA)
- Tropical Medicine & Parasitology (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Biodiversity & Conservation Biology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Physics & Mathematics (AREA)
- Fluid Mechanics (AREA)
- Hydrology & Water Resources (AREA)
- Water Supply & Treatment (AREA)
- General Life Sciences & Earth Sciences (AREA)
- Geochemistry & Mineralogy (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses one plant of microbe oil production bacterium W Y6 and its applications.Microbe oil production bacterium W Y6 are specially Georgenia sp.W Y6, are CGMCC No.15903 in the deposit number of China Committee for Culture Collection of Microorganisms's common micro-organisms center.It it is demonstrated experimentally that Georgenia sp.W Y6 not only have the function of degraded oil, can be used for the improvement and reparation of oil pollution, and can be also used for microbe oil production, improve tar productivity.The present invention has important application value.
Description
Technical field
The invention belongs to biotechnologies, and in particular to one plant of microbe oil production bacterium W-Y6 and its application.
Background technology
Oil pollution is widely distributed, and it is various to be related to the various industry such as oil recovery, oil refining, chemical industry, machinery and Oil spills etc.
Pollute environment.Petroleum hydrocarbon complicated components are various, and hydrophobicity is strong and is difficult to be contacted by common micro-organisms, degrades, utilizes.
Microbial Enhanced Oil Recovery is exactly by injecting nutrition or microorganism into stratum, utilizing the growth of microorganism in oil reservoir
Metabolic activity improves crude output and recovery ratio.Traditional oil function stem is located away from water body, soil more.It is existing to grind
Study carefully and shows that microorganism except water, soil China and foreign countries are distributed in, also has extensive distribution, therefore divide from oil oil phase in oil oil phase
It is of great significance in Microbial Enhanced Oil Recovery from relevant function stem.
Invention content
The purpose of the present invention is degraded oils.
The present invention protects Georgenia sp.W-Y6 first, and it is micro- which has been preserved in China on 06 05th, 2018
(abbreviation CGMCC, address are biological inoculum preservation administration committee common micro-organisms center：BeiChen West Road, Chaoyang District, BeiJing City 1
Institute 3), deposit number is CGMCC No.15903.Georgenia sp.W-Y6CGMCC No.15903 abbreviations Georgenia
sp.W-Y6。
The present invention also protects a kind of microbial inoculum, which contains the Georgenia sp.W-Y6.The purposes of the microbial inoculum is
A1) or A2) or A3) or A4)：A1) microbe oil production；A2) emulsified crude oil；A3) degraded oil；A4) petroleum pollution and/or
It repairs.
The preparation method of the microbial inoculum may include following steps：Georgenia sp.W-Y6 are seeded to bacteria culture media
And cultivated, obtain Georgenia sp.W-Y6 a concentration of 107-109CFU/mL (such as 107-108CFU/mL、108-109CFU/
mL、107CFU/mL、108CFU/mL or 109CFU/mL bacterium solution), the as described microbial inoculum.
The bacteria culture media can be LB liquid medium.
In the preparation method of the microbial inoculum, the condition of the culture can be：35~39 DEG C (such as 35~37 DEG C, 37~39 DEG C,
35 DEG C, 37 DEG C or 39 DEG C), 100-200r/min (such as 100-150r/min, 150-200r/min, 100r/min, 150r/min or
200r/min) cultivate 20~30h (such as 20~for 24 hours, 24~30h, 20h, for 24 hours or 30h).
In addition to active constituent, the microbial inoculum can also include carrier.The carrier can be solid carrier or liquid-carrier.Institute
It can be mineral material, vegetable material or high-molecular compound to state solid carrier.The mineral material can be clay, talcum, kaolinite
At least one of soil, montmorillonite, white carbon, zeolite, silica and diatomite.The vegetable material can be corn flour, bean powder and shallow lake
At least one of powder.The high-molecular compound can be polyvinyl alcohol.The liquid-carrier can be organic solvent, vegetable oil,
Mineral oil or water.The organic solvent can be decane and/or dodecane.In the microbial inoculum, the active constituent can be to be trained
The foster zymotic fluid of living cells, living cells, the filtrate of cell culture or the form of cell and the mixture of filtrate exists.It is described
The dosage form of composition can be a variety of dosage forms, such as liquor, emulsion, suspending agent, pulvis, granule, wettable powder or water-dispersible grain
Agent.
As needed, surfactant (such as polysorbas20, Tween 80), adhesive, stabilization can be also added in the microbial inoculum
Agent (such as antioxidant), pH adjusting agent.
The application for the microbial inoculum that the present invention also protects the Georgenia sp.W-Y6 or any of the above-described described, can be a1) or
a2)：A1) microbe oil production；A2 the product of microbe oil production) is prepared.
The application for the microbial inoculum that the present invention also protects the Georgenia sp.W-Y6 or any of the above-described described, can be b1) or
b2)：B1) emulsified crude oil；B2 the product of emulsified crude oil) is prepared.
The application for the microbial inoculum that the present invention also protects the Georgenia sp.W-Y6 or any of the above-described described, can be c1) or
c2)：C1) degraded oil；C2 the product of degraded oil) is prepared.
The application for the microbial inoculum that the present invention also protects the Georgenia sp.W-Y6 or any of the above-described described, can be d1) or
d2)：D1) petroleum pollution and/or reparation；D2 petroleum pollution and/or the product of reparation) are prepared.
The present invention also protects a kind of product, contains the microbial inoculum described in the Georgenia sp.W-Y6 or any of the above-described；
At least one of the product can have the function of following A1)-A4)：A1) microbe oil production；A2) emulsified crude oil；A3) degradation stone
Oil；A4) petroleum pollution and/or reparation.
The present invention also protects a kind of method of microbe oil production, it may include following steps：In oil recovery process, described in addition
The microbial inoculum described Georgenia sp.W-Y6 or any of the above-described.
A kind of method that the present invention also protects emulsified crude oil and/or degraded oil, it may include following steps：To crude oil or contain
The Georgenia sp.W-Y6 or any of the above-described microbial inoculums are added in the liquid-phase system for having crude oil.
A kind of method that the present invention also protects petroleum pollution and/or reparation, it may include following steps：Using described
The microbial inoculum described Georgenia sp.W-Y6 or any of the above-described handles petroleum pollution.
It is demonstrated experimentally that Georgenia sp.W-Y6CGMCC No.15903 provided by the invention not only have degraded oil
Function, can be used for the improvement and reparation of oil pollution, and can be also used for microbe oil production, improve tar productivity.The present invention
With important application value.
Description of the drawings
Fig. 1 is the effect of W-Y6 bacterial strain emulsified crude oils.
Fig. 2 is object mould displacement of reservoir oil result curve.
Preservation explanation
Latin name：Georgenia sp.
Strain number：W-Y6
Preservation mechanism：China Committee for Culture Collection of Microorganisms's common micro-organisms center
Preservation mechanism is referred to as：CGMCC
Address：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3
Preservation date：On 06 05th, 2018
Collection is registered on the books number：CGMCC No.15903
Specific implementation mode
Embodiment below facilitates a better understanding of the present invention, but does not limit the present invention.Experiment in following embodiments
Method is unless otherwise specified conventional method.Test material as used in the following examples is unless otherwise specified certainly
What routine biochemistry reagent shop was commercially available.Quantitative test in following embodiment is respectively provided with three repeated experiments, as a result makes even
Mean value.
Liquid inorganic salt culture medium：By 5g NaCl, 1g NH4H2PO4、1g(NH4)2SO4、1g K2HPO4With 3g KNO3It is molten
In deionized water, it then is settled to 1L with deionized water, adjusts pH value to 7-8；121 DEG C of sterilizing 30min.
LB liquid medium：10g NaCl, 10g peptones and 5g yeast powders are dissolved in deionized water, then use deionization
Water is settled to 1L, adjusts pH value to 7-8；121 DEG C of sterilizing 30min.
LB solid mediums：10g NaCl, 10g peptones, 5g yeast powders and 15g agar powders are dissolved in deionized water, so
It is settled to 1L with deionized water afterwards, adjusts pH value to 7-8；121 DEG C of sterilizing 30min.
LB solid plates：About 55 DEG C of LB solid mediums are poured into sterile petri dish, LB solid plates are obtained after cooling.
Embodiment 1, the separation of W-Y6 bacterial strains, identification and preservation
One, the separation of W-Y6 bacterial strains
It takes 1g crude oil (in August, 2017 picks up from Chinese Liaohe Oil Field oil well), is added 100mL liquid inorganic salt culture mediums, 37
DEG C, after 150rpm shaken cultivations 30d, dilution spread on LB solid plates, cross after purification by 37 DEG C of culture 7d, picking single bacterium colony
It preserves, the isolate for isolating and purifying acquisition is named as W-Y6 bacterial strains.
Two, the identification of W-Y6 bacterial strains
1, Morphological Identification
By in W-Y6 inoculations to LB solid plates, 37 DEG C of light cultures the form of observation bacterium colony and pass through high score after 5d
Resolution transmission electron microscope analysis observes the morphological feature of thalline.
The experimental results showed that the bacterium colony of W-Y6 bacterial strains is glassy yellow；W-Y6 bacterial strains round bar shape does not generate gemma.
2, analysis of physio biochemical characteristics
With reference to《Common bacteria system identification handbook》(east show pearl, the wonderful Beijing English common bacterias system identification handbook of Cai:Section
Publishing house, 2011.) and《Microbiology Experiment》(Beijing Shen Ping, Fan Xiurong, Li Guang force Microbiology Experiment (third edition):
Higher Education Publishing House, 1999.) physiological and biochemical property of W-Y6 bacterial strains is measured.
The result shows that the physiological and biochemical property of W-Y6 bacterial strains is as follows：Gram-positive, oxydase reaction is negative, mistake
Hydrogen oxide enzyme reaction is positive.
3,16s rDNA sequence homology analysis
(1) by W-Y6 inoculations in LB liquid medium, 37 DEG C, 150rpm shaken cultivations for 24 hours, obtain culture bacterium solution.
(2) after completing step (1), fresh culture bacterium solution, 4 DEG C, 8000rpm centrifugation 5min is taken to collect thalline in centrifugation
It manages in (specification 2mL), then DNA extraction kit is used to extract DNA.
(3) after completing step (2), the DNA of thalline is taken, electrophoresis detection is first carried out, then use universal primer 8F/1492R
PCR amplification is carried out, the PCR product containing the conserved regions W-Y6 bacterial strain 16S rDNA is obtained.Primer sequence is as follows：8F：5’-
AGAGTTTGATCCTGGCTCAG-3’；1492R：5’-GGTTACCTTGTTACGACTT-3’.
Electrophoresis detection PCR product is simultaneously sequenced.Sequencing result shows the nucleotide sequence such as sequence of W-Y6 bacterial strain 16S rDNA
In table shown in sequence 1.
By sequence 1 and NCBI (http://www.ncbi.nlm.nih.gov) in published 16S rDNA sequences carry out
Online sequence analysis.Comparison result shows W-Y6 bacterial strains and the homology highest of Georgeniasp..
In view of above-mentioned form, analysis of physio biochemical characteristics and 16S rDNA sequence homology analysis as a result, by step 1 point
The W-Y6 bacterial strains obtained from purifying are accredited as Georgeniasp..
Three, the preservation of W-Y6 bacterial strains
W-Y6 bacterial strains were preserved in China Committee for Culture Collection of Microorganisms's commonly micro- life on 06 05th, 2018
(abbreviation CGMCC, address are at object center：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3), deposit number CGMCC
No.15903.The full name of W-Y6 bacterial strains is Georgeniasp.W-Y6CGMCC No.15903.
The application of embodiment 2, W-Y6 bacterial strains in emulsified crude oil and degraded oil
One, application of the W-Y6 bacterial strains in emulsified crude oil
1, by W-Y6 inoculations in LB liquid medium, 37 DEG C, 150rpm shaken cultivations for 24 hours, obtain W-Y6 bacterium solutions (W-
In Y6 bacterium solutions, a concentration of the 10 of W-Y6 bacterial strains8CFU/mL)。
2, the W-Y6 bacterium solutions that step 1 obtains are inoculated according to the ratio of 5% (v/v) in crude oil minimal medium, 37
DEG C, 150rpm shaken cultivations 7d.
When temperature is 37 DEG C, the viscosity of crude oil is 50500mPa.s.
Crude oil minimal medium：By 1g crude oil, 5g NaCl, 1g NH4H2PO4、1g(NH4)2SO4、1g K2HPO4And 3g
KNO3It is dissolved in deionized water, is then settled to 1L with deionized water, adjusts pH value to 7-8；121 DEG C of sterilizing 30min.
3, LB liquid medium is inoculated according to the ratio of 5% (v/v) in crude oil minimal medium, 37 DEG C,
150rpm shaken cultivations 7d.As a contrast.
Experimental result is shown in Fig. 1 (left figure is that LB liquid medium handles crude oil, and right figure is that W-Y6 bacterium solutions handle crude oil).As a result
Show that W-Y6 bacterial strains have apparent emulsification at 37 DEG C to crude oil.
Two, application of the W-Y6 bacterial strains in degraded oil
1, in step 11.
2, in step 12.
3, the system for taking into step 2 is surveyed according to the method in document " separation of oil degradation bacterium and its degradation property "
Determine petroleum residual amount, calculates petroleum degradation rate.Petroleum degradation rate=(oil addition-petroleum residual amount)/oil addition.
Result of calculation shows that petroleum degradation rate is 57.6%.The result shows that W-Y6 bacterial strains have the function of degraded oil,
It can be used for the improvement and reparation of oil pollution.
The application of embodiment 3, W-Y6 bacterial strains in Microbial Enhanced Oil Recovery
The oil displacement efficiency of microorganism fungus kind is evaluated by indoor object mould oil displacement experiment, experimental procedure is specific as follows：
1, according to (0.250 μm of oil reservoir block permeability2), using High Pressure Model pipe (specificationIt purchases to Hai'an
Oil scientific research Instrument Ltd.) in load different meshes quartz sand be made as sandpack column pipe.
2, using permeability measuring apparatus measurement model pipe permeability, the model pipe met is selected；Model pipe back-up sand parameter
As shown in table 1.
1. model pipe back-up sand parameter of table
3, saturation water flooding is vacuumized, porosity is calculated.
4, oily expelling water establishes irreducible water, measures initial oil saturation；And aging 3 days under reservoir temperature (37 DEG C).
5, the displacement of reservoir oil is carried out with injection water, moisture content (98%) stops water drive until outlet production fluid reaches capacity, and calculates water drive
Recovery ratio.
6, by W-Y6 inoculations in LB liquid medium, 37 DEG C, 150rpm shaken cultivations for 24 hours, obtain W-Y6 bacterium solutions (W-
In Y6 bacterium solutions, a concentration of the 10 of W-Y6 bacterial strains8CFU/mL)。
7, NH is added in injecting water4H2PO4、(NH4)2SO4、K2HPO4、KNO3, peptone and yeast powder and W-Y6 bacterium
Liquid obtains the mixed liquor containing W-Y6.In mixed liquor containing W-Y6, NH4H2PO4、(NH4)2SO4And K2HPO4Concentration be 1g/L,
KNO3A concentration of 3g/L, a concentration of 2g/L of peptone, a concentration of 1g/L of yeast powder, a concentration of 5% (v/ of W-Y6 bacterium solutions
v).In mixed liquor containing W-Y6, a concentration of the 10 of W-Y6 bacterial strains8CFU/mL。
It 8, will be in the above-mentioned mixed liquor injection model pipe containing W-Y6 by 1.0PV injection rates；It is trained under reservoir temperature (37 DEG C)
It supports 15 days.
9, secondary water drive, water drive to aqueous 98% or more, during calculating separately note microorganism (mixed liquor containing W-Y6),
Microorganism (mixed liquor containing W-Y6) acts on the raising recovery ratio of the recovery ratio and W-Y6 bacterial strains of subsequent waterflooding.
In water drive, note microbial process, result such as 2 institute of table of the recovery ratio of different phases such as microbial action subsequent waterflooding
Show.To in water drive, note microbial process, the recovery ratio of the different phases such as microbial action subsequent waterflooding carry out comparison discovery：Make
It is recovered the oil with W-Y1 bacterial strains, recovery ratio improves 5.06%.Object mould displacement of reservoir oil result curve is as shown in Figure 2.
In 2. water drive of table, note microbial process, the recovery ratio of the different phases such as microbial action subsequent waterflooding
| Object mode step section | Recovery ratio (%) | Improve recovery ratio (%) |
| Water drive | 31.81 | / |
| Note microbial process | 32.05 | 0.24 |
| Microbial action subsequent waterflooding | 36.87 | 5.06 |
Note："/" expression is not present.
<110>Beijing Run Shi Enertech Co., Ltd.
<120>One plant of microbe oil production bacterium W-Y6 and its application
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1372
<212> DNA
<213>Artificial sequence
<220>
<223>
<400> 1
cgatgaagcc cagcttgctg ggtggattag tggcgaacgg gtgagtaaca cgtgagtaac 60
ctgcccctga cttcgggata actgcgagaa atcgtggcta ataccggata tgcacatgtg 120
cctgcatggg tgtgtgtgga aagatttatc ggttggggat gggctcgcgg cctatcagct 180
tgttggtggg gtgatggcct accaaggcga cgacgggtag ccggcctgag agggtgaccg 240
gccacactgg gactgagaca cggcccagac tcctacggga ggcagcagtg gggaatattg 300
cacaatgggc gcaagcctga tgcagcgacg ccgcgtgagg gatgacggcc ttcgggttgt 360
aaacctcttt cagtagggaa gaagctcttc ggagtgacgg tacctgcaga agaagcgccg 420
gctaactacg tgccagcagc cgcggtaata cgtagggcgc aagcgttgtc cggaattatt 480
gggcgtaaag agctcgtagg cggtttgtcg cgtctgctgt gaaaacgcaa ggcttaacct 540
tgcgctgcag tgggtacggg cagactagag tgcggtaggg gagactggaa ttcctggtgt 600
agcggtggaa tgcgcagata tcaggaggaa caccgatggc gaaggcaggt ctctgggccg 660
ttactgacgc tgaggagcga aagcatgggg agcgaacagg attagatacc ctggtagtcc 720
atgccgtaaa cgttgggcac taggtgtggg atccattcca cgggttccgt gccgcagcta 780
acgcattaag tgccccgcct ggggagtacg gccgcaaggc taaaactcaa aggaattgac 840
gggggcccgc acaagcggcg gagcatgcgg attaattcga tgcaacgcga agaaccttac 900
caaggcttga catacaccgg aaaagtgcag agatgtgctc cccgtaaggt cggtgtacag 960
gtggtgcatg gttgtcgtca gctcgtgtcg tgagatgttg ggttaagtcc cgcaacgagc 1020
gcaacccttg tcctgtgttg ccagcacgta atggtgggga ctcataggag actgccgggg 1080
tcaactcgga ggaaggtggg gatgacgtca aatcatcatg ccccttatgt cttgggcttc 1140
acgcatgcta caatggccgg tacaaagggc tgcgataccg cgaggtggag cgaatcccaa 1200
aaagccggtc tcagttcgga ttggggtctg caactcgacc ccatgaagtc ggagtcgcta 1260
gtaatcgcag atcagcaacg ctgcggtgaa tacgttctcg ggccttgtac acaccgcccg 1320
tcacgtcatg aaagtcggta acacccgaag ccggtggccc aacccttgtg gg 1372
Claims (10)
1.Georgenia sp.W-Y6, in the preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms center
Number is CGMCC No.15903.
2. a kind of microbial inoculum, it is characterised in that：The microbial inoculum contains Georgenia sp.W-Y6 described in claim 1.
3. the application of the microbial inoculum described in Georgenia sp.W-Y6 or claim 2 described in claim 1 is a1) or a2)：
A1) microbe oil production；A2 the product of microbe oil production) is prepared.
4. the application of the microbial inoculum described in Georgenia sp.W-Y6 or claim 2 described in claim 1 is b1) or b2)：
B1) emulsified crude oil；B2 the product of emulsified crude oil) is prepared.
5. the application of the microbial inoculum described in Georgenia sp.W-Y6 or claim 2 described in claim 1 is c1) or c2)：
C1) degraded oil；C2 the product of degraded oil) is prepared.
6. the application of the microbial inoculum described in Georgenia sp.W-Y6 or claim 2 described in claim 1 is d1) or d2)：
D1) petroleum pollution and/or reparation；D2 petroleum pollution and/or the product of reparation) are prepared.
7. a kind of product contains Georgenia sp.W-Y6 described in claim 1 or the microbial inoculum described in claim 2；It is described
At least one of product has the function of following A1)-A4)：A1) microbe oil production；A2) emulsified crude oil；A3) degraded oil；A4) stone
Oily pollution is administered and/or is repaired.
8. a kind of method of microbe oil production, includes the following steps：In oil recovery process, it is added described in claim 1
Microbial inoculum described in Georgenia sp.W-Y6 or claim 2.
9. a kind of method of emulsified crude oil and/or degraded oil, includes the following steps：Liquid-phase system to crude oil or containing crude oil
The middle microbial inoculum being added described in Georgenia sp.W-Y6 or claim 2 described in claim 1.
10. a kind of petroleum pollution and/or the method for reparation, include the following steps：Using described in claim 1
Microbial inoculum described in Georgenia sp.W-Y6 or claim 2 handles petroleum pollution.
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| CN109234200A (en) * | 2018-10-25 | 2019-01-18 | 北京润世能源技术有限公司 | One plant of microbe oil production bacterium W-Y9 and its application |
| CN109306333A (en) * | 2018-10-25 | 2019-02-05 | 北京润世能源技术有限公司 | One plant of microbe oil production bacterium W-Y8 and its application |
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| CN109234201A (en) * | 2018-10-25 | 2019-01-18 | 北京润世能源技术有限公司 | One plant of microbe oil production bacterium W-Y10 and its application |
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