CN102250801A - Microbial agent, and its preparation method and application - Google Patents
Microbial agent, and its preparation method and application Download PDFInfo
- Publication number
- CN102250801A CN102250801A CN2011101698214A CN201110169821A CN102250801A CN 102250801 A CN102250801 A CN 102250801A CN 2011101698214 A CN2011101698214 A CN 2011101698214A CN 201110169821 A CN201110169821 A CN 201110169821A CN 102250801 A CN102250801 A CN 102250801A
- Authority
- CN
- China
- Prior art keywords
- cgmcc
- preserving number
- pseudomonas
- microbiobacterial agent
- rhodococcus ruber
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Abstract
The invention provides a microbial agent which comprises a medium and thalli, wherein, the thalli comprise at least two selected from the group consisting of pseudoxanthomonas japonensis with an accession number CGMCC No.4797, pseudomonas with an accession number CGMCC No.4793, gordonia amicalis with an accession number CGMCC No.4794, rhodococcus rubber with an accession number CGMCC No. 4795 and gordonia alkanivorans with an accession number CGMCC No.4796. The invention also provides a preparation method for the microbial agent and application of the microbial agent in degradation of petroleum hydrocarbon, bioremediation of soil polluted by petroleum and bioremediation of water bodies polluted by petroleum.
Description
Technical field
The present invention relates to a kind of microbiobacterial agent and its production and application; Or rather, relate to a kind of microbiobacterial agent and preparation method thereof, also relate to application, application in biological restoration petroleum pollution, application in the soil of biological restoration petroleum pollution and the application in the water body of biological restoration petroleum pollution of described bacteria agent in decomposing petroleum hydrocarbon.
Background technology
The world today, oil has become one of human topmost energy, and a large amount of oil and processed goods thereof enter the petroleum pollution that soil causes soil, bring harm for human mater to whole biosphere, thereby become international environmental problem.In the research to petroleum pollution, biological restoration as a kind of potential efficiently, cleaning technique come into one's own just day by day cheaply.
Biological restoration (Bioremediation) is meant by biological (particularly microorganism) catalyze and degrade organic pollutants, thus the controlled or spontaneous process of carrying out of repairing the pollutent in contaminated environment or the elimination environment.
" petroleum hydrocarbon " is the main pollutant component in oil and the processed goods thereof, discovers that microorganism can play Degradation to petroleum hydrocarbon.The microbiological deterioration petroleum hydrocarbon is to find for 19 end of the centurys.Before the 1950's, Bel is representative with U.S. C.E. assistant, and the marine microorganism decomposing petroleum hydrocarbon has been carried out extensive studies.The beginning of the fifties, gas-chromatography was come out, and the widespread usage of radio-label method has play a part positive to the microbiological deterioration mechanism of studying petroleum hydrocarbon.Since the sixties,, impelled many maritime nations,, actively developed the research work of relevant marine microorganism decomposing petroleum hydrocarbon as states such as the U.S., Canada, Japan, Britain and the Soviet Union because offshore oil pollution is on the rise.The mid-1970s, American scholar have also been cultivated " super microorganism " with engineered technology, in the hope of decomposing petroleum hydrocarbon effectively.
China is from 1975, successively Jiaozhou Bay, Qingdao, the Bohai Sea, PORT OF XIAMEN, the Huanghai Sea and East Sea oil degradation microbial numbers, distribution, kind formed and influenced degradation factors etc. and investigate.
In influencing the principal element of microorganism to the degraded of petroleum hydrocarbon, the physiological property of microorganism is one of key factor of decision decomposing petroleum hydrocarbon ability.It is also lower to the petroleum hydrocarbon degradation ability at present to have separated the microorganism strains and the microbial inoculum that obtain, therefore, presses for and develops a kind of microbiobacterial agent that can the efficient degradation petroleum hydrocarbon.
Summary of the invention
The objective of the invention is to overcome the existing microbiobacterial agent shortcoming low, microbiobacterial agent of a kind of decomposing petroleum hydrocarbon efficiently and its production and application is provided the degradation capability of petroleum hydrocarbon.
In order to realize first goal of the invention, the invention provides a kind of microbiobacterial agent, this microbiobacterial agent comprises substratum and thalline, wherein, described thalline comprises at least two kinds that are selected from the following bacterial classification: preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797, preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796.
In order to realize second goal of the invention, the present invention also provides a kind of preparation method of microbiobacterial agent, wherein, this method comprises: will be selected from preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797, preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is that at least two kinds of bacterial classification inoculations in the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and the food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796 are cultivated in substratum.
The present invention also provides the application of described microbiobacterial agent in decomposing petroleum hydrocarbon.
The present invention also provides the application of described microbiobacterial agent in the biological restoration petroleum pollution.
The present invention also provides the application of described microbiobacterial agent in the soil of biological restoration petroleum pollution.
In addition, the present invention also provides the application of described microbiobacterial agent in the water body of biological restoration petroleum pollution.
According to the present invention, the bacterial strain that uses in the described microbiobacterial agent screens by a large amount of screening operations, and these bacterial strains have very strong degradation capability to petroleum hydrocarbon.These bacterial strains have been stored in (address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City), China Committee for Culture Collection of Microorganisms common micro-organisms center on 04 28th, 2011, wherein, the preserving number of Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) is CGMCC No.4797, the preserving number of pseudomonas (Pseudomonas) is CGMCC No.4793, the preserving number of friendly Gordon Salmonella (Gordonia amicalis) is CGMCC No.4794, the preserving number of Rhodococcus ruber (Rhodococcus ruber) is CGMCC No.4795, the preserving number of food alkane Gordon Salmonella (Gordonia alkanivorans) is CGMCC No.4796.Microbiobacterial agent provided by the invention has very strong degradation capability to petroleum hydrocarbon, can be widely used in decomposing petroleum hydrocarbon and to the reparation of petroleum pollution, as to the reparation of the soil of petroleum pollution with to reparation of the water body of petroleum pollution etc.
Description of drawings
Fig. 1 has shown that the preserving number of the present invention's screening is the form that the Japanese false Xanthomonas campestris (Pseudoxant homonasjaponensis) of CGMCC No.4797 was cultivated 24 hours on solid TSA substratum;
Fig. 2 has shown that the preserving number of the present invention's screening is that the Japanese false Xanthomonas campestris (Pseudoxan thomonasjaponensis) of CGMCC No.4797 was being cultivated 24 hours on the solid TSA substratum after the form under 100 times of oily mirrors of lawn dyeing back at opticmicroscope;
Fig. 3 has shown that the preserving number of the present invention's screening is the form that the pseudomonas (Pseudomonas) of CGMCC No.4793 was cultivated 24 hours on solid TSA substratum;
Fig. 4 has shown that the preserving number of the present invention's screening is that the pseudomonas (Pseudomonas) of CGMCC No.4793 was being cultivated 24 hours on the solid TSA substratum after lawn dyeing back form of (10 * 100 times) under opticmicroscope oil mirror;
Fig. 5 has shown that the preserving number of the present invention's screening is the form that friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 was cultivated 24 hours on solid TSA substratum;
Fig. 6 has shown that the preserving number of the present invention's screening is that friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 was being cultivated 24 hours on the solid TSA substratum after the form under 100 times of oily mirrors of lawn dyeing back at opticmicroscope;
Fig. 7 for the preserving number of the present invention screening be stereoscan photograph after cultivating 24 hours on the solid TSA substratum of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 (* 10K);
Fig. 8 for the preserving number of the present invention screening be stereoscan photograph after cultivating 48 hours on the solid TSA substratum of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 (* 10K);
Fig. 9 has shown that the preserving number of the present invention's screening is the form that the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 was cultivated 24 hours on solid TSA substratum;
Figure 10 has shown that the preserving number of the present invention's screening is that the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 was being cultivated 24 hours on the solid TSA substratum after the form under 100 times of oily mirrors of lawn dyeing back at opticmicroscope;
Figure 11 for the preserving number of the present invention screening be stereoscan photograph after cultivating 24 hours on the solid TSA substratum of the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 (* 10K);
Figure 12 for the preserving number of the present invention screening be stereoscan photograph after cultivating 48 hours on the solid TSA substratum of the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 (* 10K);
Figure 13 has shown that the preserving number of the present invention's screening is the form that food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 was cultivated 24 hours on solid TSA substratum;
Figure 14 has shown that the preserving number of the present invention's screening is that food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 was being cultivated 24 hours on the solid TSA substratum after the form under 100 times of oily mirrors of lawn dyeing back at opticmicroscope;
Figure 15 for the preserving number of the present invention screening be stereoscan photograph after cultivating 24 hours on the solid TSA substratum of food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 (* 10K);
Figure 16 for the preserving number of the present invention screening be stereoscan photograph after cultivating 48 hours on the solid TSA substratum of food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 (* 10K).
Embodiment
The invention provides a kind of microbiobacterial agent, this microbiobacterial agent comprises substratum and thalline, wherein, described thalline comprises at least two kinds that are selected from the following bacterial classification: preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797, preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkan ivorans) that preserving number is CGMCC No.4796.
In a kind of preferred implementation, described thalline comprises that preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797, also comprises being selected from least a in the following bacterial classification: preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796.
Through 16s rDNA amplification and sequencing analysis, deposit number is that the bacterial strain of CGMCC No.4797 has the 16s rDNA sequence shown in the SEQ ID NO:1; Deposit number is that the bacterial strain of CGMCC No.4793 has the 16s rDNA sequence shown in the SEQ ID NO:2; Deposit number is that the bacterial strain of CGMCC No.4794 has the 16s rDNA sequence shown in the SEQ ID NO:3; Deposit number is that the bacterial strain of CGMCCNo.4795 has the 16s rDNA sequence shown in the SEQ ID NO:4; Deposit number is that the bacterial strain of CGMCC No.4796 has the 16s rDNA sequence shown in the SEQ ID NO:5.
The amount of the thalline that contains in the described microbiobacterial agent can in very large range change, and under the preferable case, the contained total viable count of the described microbiobacterial agent of every gram is 0.5-1 * 10
10Individual.
A kind of preferred embodiment in, in the described microbiobacterial agent, preserving number is that the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 is the 10-90% of total viable count; 30-70% more preferably.
According to the present invention, the kind of described substratum can in very large range change, can be the various substratum of cultivating Japanese false Xanthomonas campestris, pseudomonas, friendly Gordon Salmonella, Rhodococcus ruber and food alkane Gordon Salmonella that can be used in, for example, can be in beef-protein medium, oil screening culture medium and the PDA substratum one or more, be preferably beef-protein medium.Above-mentioned substratum can be commercially available or prepare according to the record of " microbiological culture media handbook " (Microbiology Culture Media Manual).For example, described beef-protein medium can contain extractum carnis, peptone, NaCl and water, wherein, extractum carnis with respect to 100 weight parts, the content of peptone can be the 150-250 weight part, the content of NaCl can be the 50-150 weight part, and the content of water can be the 15000-25000 weight part, and the pH value can be 7.2-7.4; The agar that adds 1.2 weight % in the substratum can be made into solid medium.
The present invention also provides the preparation method of described microbiobacterial agent, wherein, this method comprises and will be selected from the Japanese false Xanthomonas campestris that preserving number is CGMCC No.4797 (Pseudoxanthomonas japonensis), preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is that at least two kinds of bacterial classification inoculations in the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and the food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796 are cultivated in substratum.
In a kind of preferred implementation, described preparation method comprises: with preserving number is that the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 and at least a being inoculated in the substratum that be selected from the following bacterial classification are cultivated: preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796.
According to the present invention, described cultured method comprises: independently cultivating respectively in the culture system separately, and will cultivate the microorganism that obtains respectively and proportionally mix.Under the preferable case, by separately independently the cultivation in the culture system and cultivate after mixing, the total viable count that makes the every gram microbiobacterial agent that obtains is 0.5-1 * 10
10Individual.
In a kind of preferred implementation, by separately independently the cultivation in the culture system and cultivate after proportionally mixing, described blended condition has no particular limits, as long as the microbiobacterial agent that obtains is met the following conditions: make in the every gram microbiobacterial agent that obtains, preserving number be CGMCCNo.4797 Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) total viable count be the 10-90% of viable count; 30-70% more preferably.
According to the present invention, described preserving number is that the cultural method of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 has no particular limits, for example, can in the beef-protein medium of 100 weight parts, inoculate the bacterial strain of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of 2-5 weight part, 37 ℃ of cultivations are 0.5-1 * 10 until the viable count of the false Xanthomonas campestris of Japan (Pseudoxanthomonasjaponensis)
10The substratum of individual/gram.
Described preserving number is that the cultural method of the pseudomonas (Pseudomonas) of CGMCC No.4793 has no particular limits, for example, can in the beef-protein medium of 100 weight parts, inoculate the bacterial strain of the pseudomonas (Pseudomonas) of 2-5 weight part, 37 ℃ of cultivations are 0.5-1 * 10 until the viable count of pseudomonas (Pseudomonas)
10The substratum of individual/gram.
Described preserving number is that the cultural method of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 has no particular limits, for example, can in the beef-protein medium of 100 weight parts, inoculate the bacterial strain of friendly Gordon Salmonella (Gordonia amicalis) of 2-5 weight part, 37 ℃ of cultivations are 0.5-1 * 10 until the viable count of friendly Gordon Salmonella (Gordonia amicalis)
10The substratum of individual/gram.
Described preserving number is that the cultural method of the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 has no particular limits, for example, can in the beef-protein medium of 100 weight parts, inoculate the bacterial strain of the Rhodococcus ruber (Rhodococcus ruber) of 2-5 weight part, 37 ℃ of cultivations are 0.5-1 * 10 until the viable count of Rhodococcus ruber (Rhodococcus ruber)
10The substratum of individual/gram.
Described preserving number is that the cultural method of food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 has no particular limits, for example, can in the beef-protein medium of 100 weight parts, inoculate food alkane Gordon Salmonella (Gordonia alkanivorans) bacterial strain of 2-5 weight part, 37 ℃ of cultivations are 0.5-1 * 10 until the viable count of eating alkane Gordon Salmonella (Gordonia alkanivorans)
10The substratum of individual/gram.
The present invention also provides the application of described microbiobacterial agent in decomposing petroleum hydrocarbon.
The present invention also provides the application of described microbiobacterial agent in the biological restoration petroleum pollution.
The present invention also provides the application of described microbiobacterial agent in the soil of biological restoration petroleum pollution.
In addition, the present invention also provides the application of described microbiobacterial agent in the water body of biological restoration petroleum pollution.
Below by specific embodiment the present invention is further illustrated.
Experiment material
One, the oil sample of strain screening:
The crude oil and the commercially available 0# diesel oil that pick up from Shengli Oil Field are even by 1: 4 mixed, as the experiment oil.
Two, pedotheque is 1: 1: 1 the mixture that picks up from three kinds of different oil-polluted soils samples of Shandong Shengli Oil Field, and it is respectively:
1) the ground crude oil contaminated soil around the producing well;
2) oil sediment; With
3) stack the topsoil soil of oil sediment for a long time.
Three, substratum
1) enrichment medium: K
2HPO
43H
2O 1.0g, KH
2PO
41.0g, MgSO
47H
2O 0.5g, NH
4NO
31.0g, CaCl
20.02g distilled water 1000mL transfers to PH7.0,121 ℃ of sterilization 20min add the 5g oil after cooling.
2) isolation medium (PDA substratum): potato powder 6.0g, glucose 20g, agar 20g adds distilled water until 1000ml, and standby 115 ℃ of autoclavings 20 minutes.
3) strain store medium: beef-protein medium
4) oil screening culture medium (liquid): NaNO
31.5g, (NH4)
2SO
41.5g, K
2HPO
41g, MgSO
47H
2O 0.5g, KCl 0.5g, FeSO
47H
2O 0.01g, CaCl
20.002g distilled water 1000mL transfers to pH8.0,121 ℃ of sterilization 20min, and the cooling back adds the 2.5g oil.The agar that adds 1.2 weight % in the substratum is made the solid screening culture medium.
5) solid TSA substratum: Tryptones 15g, soya peptone 5g, sodium-chlor 30g, agar 15g, adding distilled water is 1000ml until final volume, the pH value is 7.1-7.5.
Embodiment 1
1, the concentration and separation of petroleum hydrocarbon degradation bacterial strain
Taking by weighing the adding of 5g oil-polluted soils mixture is equipped with in the 250mL triangular flask of 100mL enrichment medium, under 28 ℃, the constant temperature shaking table condition of 180r/min, cultivate, draw the 5mL nutrient solution after 5 days and be forwarded to fresh enrichment medium, the same terms was cultivated 5 days, enrichment culture is 25 days so continuously, nutrient solution is through being coated on behind the gradient dilution on the solid PDA substratum, in 28 ℃ of cultivations; After treating that flat board grows bacterium colony, single bacterium colony of picking different shape adopts the line partition method to carry out separation and purification repeatedly, and up to the pure growth that obtains single strain, the bacterial strain behind the purifying is stored in the beef-protein medium inclined-plane.
2, the screening of high degrading activity bacterial strain
The bacterial strain of above-mentioned purifying is inoculated into respectively in the 250mL triangular flask that the 100mL screening culture medium is housed, cultivated 5 days under 28 ℃, the condition of 180r/min, be coated on the solid screening culture medium after 100 times of the nutrient solution dilutions, observe the growing state of each bacterial strain in solid medium, filter out grow very fast, the more bacterial strain of bacterium colony.
Single bacterium colony of each bacterial strain of picking is inoculated into respectively in the 250ml triangular flask that 50ml liquid beef-protein medium is housed, in 28 ℃, the constant temperature shaking table of 180r/min, cultivate 24h, measure the absorbancy of bacterium liquid at the 600nm place, regulate its mass concentration with the beef-protein medium of sterilization and make OD600nm be about 0.4, make seed inoculation screening culture medium with this bacterium liquid that dilutes.
Drawing the ready seed liquor of 5ml is inoculated in the 250mL triangular flask that the 100mL screening culture medium is housed, measure oil content after in 28 ℃, the constant temperature shaking table of 180r/min, cultivating 4 days, the processing triplicate of each seed liquor, calculate average degradation rate, filter out 5 the highest bacterial strains of oil degradation rate, and this bacterial strain carried out biological preservation, preserving number is respectively CGMCC No.4793, CGMCC No.4794, CGMCC No.4795, CGMCC No.4796 and CGMCC No.4797.
Embodiment 2
According to experiment content and experimental technique of record in " common bacteria system identification handbook " [4] and " actinomycetes systematics principle, method and put into practice " [5], respectively to sieve preserving number be that the bacterial strain of CGMCC No.4793, CGMCC No.4794, CGMCC No.4795, CGMCC No.4796 and CGMCCNo.4797 carries out strain identification.
1, morphological analysis
Be that preserving number is that the bacterial strain of CGMCC No.4793, CGMCC No.4794, CGMCC No.4795, CGMCC No.4796 and CGMCC No.4797 carries out morphological analysis and observes under opticmicroscope to preserving number respectively, the result is respectively shown in Fig. 1-16.
Colonial morphology is observed and is carried out on solid TSA substratum, after the bacterial strain streak inoculation, observes colonial morphology in 24 hours in 28 ℃ of constant temperature culture.
Preserving number is that the bacterium colony of the bacterial strain of CGMCC No.4793 is colourless translucent, circle, and the edge is irregular, and quality is smooth, and the surface is flat.Picking is cultivated 24 hours lawn dyeing backs and is observed under 100 times of oily mirrors of opticmicroscope.Can see clearly that at microscopically preserving number is that the somatic cells of the bacterial strain of CGMCC No.4793 is stock shape, and single appearance.
Preserving number is that the bacterium colony of the bacterial strain of CGMCC No.4794 is a light orange, circle, and quality is smooth, rat.Picking is cultivated 24 hours lawn dyeing backs and is observed under 100 times of oily mirrors of opticmicroscope.Can see clearly that at microscopically preserving number is that the somatic cells of bacterial strain of CGMCC No.4794 is less relatively, and in pairs or gather into Xiao Cong and occur.
Preserving number is that the bacterium colony of the bacterial strain of CGMCC No.4795 is a pink colour, and circle, quality are slightly coarse, surface elevation.Picking is cultivated 24 hours lawn dyeing backs and is observed under 100 times of oily mirrors of opticmicroscope.Can see clearly that at microscopically preserving number is that the somatic cells of bacterial strain of CGMCC No.4795 is less relatively, and in pairs or gather into Xiao Cong and occur.
Preserving number is that the bacterium colony of the bacterial strain of CGMCC No.4796 is orange, circle, and quality is smooth, rat.Picking is cultivated 24 hours lawn dyeing backs and is observed under 100 times of oily mirrors of opticmicroscope.Can see clearly that at microscopically preserving number is that the somatic cells of bacterial strain of CGMCC No.4796 is less relatively, and in pairs or gather into Xiao Cong and occur.
Preserving number is that the bacterium colony of the bacterial strain of CGMCC No.4797 is light yellow, circle, and quality is smooth, surface elevation.Picking is cultivated 24 hours lawn dyeing backs and is observed under 100 times of oily mirrors of opticmicroscope.Can see clearly that at microscopically preserving number is that the somatic cells of the bacterial strain of CGMCC No.4797 is stock shape, and single appearance.
2,16s rDNA identifies
The pcr amplification of 16s rDNA and order-checking: adopt test kit (the full Shi Jin in Beijing biotech company) to extract and respectively the purifying preserving number be total DNA of the bacterial classification of CGMCC No.4793, CGMCC No.4794, CGMCC No.4795, CGMCC No.4796 and CGMCC No.4797, be template with total DNA then, 16s rDNA conserved sequence carried out pcr amplification obtain target gene fragment.
The pcr amplification primer is respectively:
27f (5 '-AGAGTTTGATCCTGGCTCAG-3 ') and;
1492r(5’-TACGGCTACCTTGTTACGACTT-3’)。
PCR total reaction system is 50uL, specifically comprises:
| 10×Buffer | 5μL |
| 10mmol/L dNTP | 4μL |
| Reverse primer | 1μL |
| Forward primer | 1μL |
| Taq enzyme (5U/ μ L) | 1μL |
| Template DNA | 2μL |
| ddH 2O | Complement to 50 μ L |
The PCR reaction conditions is: 94 ℃ of pre-sex change 5min, and 94 ℃ of sex change 1min, 50 ℃ of annealing 1min, 72 ℃ are extended 2min, and this step is carried out 35 circulations altogether, and last 72 ℃ are extended 10min.
Amplified production carries out the electrophoresis check with 1% sepharose.The examining order of PCR product is finished by Beijing three rich polygala root biotech companies.
Sequencing result shows that deposit number is that the bacterial strain of CGMCC No.4797 has the 16s rDNA sequence shown in the SEQ ID NO:1; Deposit number is that the bacterial strain of CGMCC No.4793 has the 16s rDNA sequence shown in the SEQ IDNO:2; Deposit number is that the bacterial strain of CGMCC No.4794 has the 16s rDNA sequence shown in the SEQID NO:3; Deposit number is that the bacterial strain of CGMCC No.4795 has the 16s rDNA sequence shown in the SEQ ID NO:4; Deposit number is that the bacterial strain of CGMCC No.4796 has the 16s rDNA sequence shown in the SEQ ID NO:5.
Through Phylogenetic Analysis: land http://www.ncbi.nlm.nih.gov, check order row are analyzed by the sequence in blast program and the nucleic acid database search same or analogous nucleotide sequence, utilize sequence alignment software ClustalX2.0 and Phylogenetic Analysis software MEGA4.0.2 constructing system evolutionary tree then.In MEGA software, the method of constructing system evolutionary tree is used the Neighbor-Joining method, the base alternative model is selected the Kimura two-parameter model, the evolutionary tree certificate authenticity is selected 1000 duplicate detection of check (bootstrap) of bootstrapping for use, and the conversion in the mutant dna sequence is given identical weighted value with transversion.Show by analysis, preserving number be CGMCC No.4797 bacterial strain belong to Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis), preserving number is that the bacterial strain of CGMCC No.4793 belongs to pseudomonas (Pseudomonas), preserving number is that the bacterial strain of CGMCCNo.4794 belongs to friendly Gordon Salmonella (Gordonia amicalis), preserving number is that the bacterial strain of CGMCCNo.4795 belongs to Rhodococcus ruber (Rhodococcus ruber), and preserving number is that the bacterial strain of CGMCC No.4796 belongs to food alkane Gordon Salmonella (Gordonia alkanivorans).
Embodiment 3-7
Use preserving number as food alkane Gordon Salmonella (Gordonia alkanivorans) of CGMCC No.4796 the oil-polluted soils mixture to be handled as Rhodococcus ruber (Rhodococcus ruber) and the preserving number of CGMCC No.4795 as friendly Gordon Salmonella (Gordonia amicalis), the preserving number of CGMCC No.4794 as pseudomonas (Pseudomonas), the preserving number of CGMCC No.4793 as Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis), the preserving number of CGMCC No.4797 respectively, concrete steps are as follows:
Taking by weighing 10g oil-polluted soils mixture (oil content 1.25 gram) adds and the 100mL beef-protein medium is housed (inoculum size of bacterial classification is that the contained total viable count of every gram substratum is 0.3 * 10
10Individual) the 250mL triangular flask in, under 28 ℃, the constant temperature shaking table condition of 180r/min, cultivate, detect the oil content in the oil-polluted soils mixture after 3 days, detect the residual petroleum content in the oil-polluted soils mixture after handling, and the calculating microbiobacterial agent is to the degradation rate of oil, oil content before oil content/processing after oil content-processing before degradation rate=processing, the result is as shown in table 1 below.
Table 1
From the data of last table 1 as can be seen, the preserving number that the present invention filters out is the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797, preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796, petroleum hydrocarbon had good degradation capability, can be widely used in decomposing petroleum hydrocarbon and to the reparation of petroleum pollution, as to the reparation of the soil of petroleum pollution with to reparation of the water body of petroleum pollution etc.
Embodiment 8
(1) preserving number of inoculation 3 weight parts is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.5 * 10 until the viable count of the false Xanthomonas campestris of Japan
10The substratum of individual/gram.
(2) preserving number of inoculation 4 weight parts is the pseudomonas (Pseudomonas) of CGMCC No.4793 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.5 * 10 until the viable count of pseudomonas
10The substratum of individual/gram.
(3) Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) and the pseudomonas (Pseudomonas) that obtains respectively in the step (1) to (2) proportionally mixed, make in the microbiobacterial agent that obtains, preserving number be the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797 be total viable count 70%, preserving number is that the viable count of the pseudomonas (Pseudomonas) of CGMCC No.4793 is 30% of total viable count, obtains microbiobacterial agent A1.
Embodiment 9
(1) preserving number of inoculation 5 weight parts is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 1 * 10 until the viable count of the false Xanthomonas campestris of Japan
10The substratum of individual/gram.
(2) preserving number of inoculation 3 weight parts is the pseudomonas (Pseudomonas) of CGMCC No.4793 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 1 * 10 until the viable count of pseudomonas
10The substratum of individual/gram.
(3) preserving number of inoculation 4 weight parts is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 1 * 10 until the viable count of friendly Gordon Salmonella
10The substratum of individual/gram.
(4) with the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) that obtains respectively in the step (1) to (3), pseudomonas (Pseudomonas) and friendly Gordon Salmonella (Gordonia amicalis) proportionally mix, make in the microbiobacterial agent that obtains, preserving number is that the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 is 40% of total viable count, preserving number is that the viable count of the pseudomonas (Pseudomonas) of CGMCC No.4793 is 30% of total viable count, preserving number is that the viable count of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 is 30% of total viable count, obtains microbiobacterial agent A2.
Embodiment 10
(1) preserving number of inoculation 4 weight parts is the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of the false Xanthomonas campestris of Japan
10The substratum of individual/gram.
(2) preserving number of inoculation 2 weight parts is the pseudomonas (Pseudomonas) of CGMCC No.4793 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of pseudomonas
10The substratum of individual/gram.
(3) preserving number of inoculation 3 weight parts is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of friendly Gordon Salmonella
10The substratum of individual/gram.
(4) preserving number of inoculation 4 weight parts is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of Rhodococcus ruber
10The substratum of individual/gram.
(5) with the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) that obtains respectively in the step (1) to (4), pseudomonas (Pseudomonas) and friendly Gordon Salmonella (Gordonia amicalis), Rhodococcus ruber (Rhodococcus ruber) proportionally mixes, make in the microbiobacterial agent that obtains, preserving number is that the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonas japonensis) of CGMCC No.4797 is 30% of total viable count, preserving number is that the viable count of the pseudomonas (Pseudomonas) of CGMCC No.4793 is 20% of total viable count, preserving number is that the viable count of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 is 25% of total viable count, preserving number is that the viable count of the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 is 25% of total viable count, obtains microbiobacterial agent A3.
Embodiment 11
(1) preserving number of inoculation 2 weight parts is the pseudomonas (Pseudomonas) of CGMCC No.4793 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of pseudomonas
10The substratum of individual/gram.
(2) preserving number of inoculation 3 weight parts is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of friendly Gordon Salmonella
10The substratum of individual/gram.
(3) preserving number of inoculation 4 weight parts is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 in the beef-protein medium of 100 weight parts, 37 ℃ of cultivations, taking a sample in culturing process and observe by the microscope direct-counting method, is 0.8 * 10 until the viable count of Rhodococcus ruber
10The substratum of individual/gram.
(4) with the pseudomonas (Pseudomonas) and the friendly Gordon Salmonella (Gordonia amicalis) that obtain respectively in the step (1) to (3), Rhodococcus ruber (Rhodococcus ruber) proportionally mixes, make in the microbiobacterial agent that obtains, preserving number is that the viable count of the pseudomonas (Pseudomonas) of CGMCC No.4793 is 25% of total viable count, preserving number is that the viable count of friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794 is 25% of total viable count, preserving number is that the viable count of the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 is 50% of total viable count, obtains microbiobacterial agent A4.
Embodiment 12-15
Use the microbiobacterial agent A1-A4 that makes among the embodiment 8-11 that the oil-polluted soils mixture is handled respectively, concrete steps are as follows:
Taking by weighing 10g oil-polluted soils mixture (oil content 1.25 grams) adds and is equipped with in the 250mL triangular flask of 100mL microbiobacterial agent, under 28 ℃, the constant temperature shaking table condition of 180r/min, cultivate, detect the oil content in the oil-polluted soils mixture after 3 days, detect the residual petroleum content in the oil-polluted soils mixture after handling, and the calculating microbiobacterial agent is to the degradation rate of oil, oil content before oil content/processing after oil content-processing before degradation rate=processing, the result is as shown in table 2 below.
Table 2
By the result in the last table 2 as can be seen, microbiobacterial agent provided by the invention has good degradation capability to petroleum hydrocarbon, can be widely used in decomposing petroleum hydrocarbon and to the reparation of petroleum pollution, as to the reparation of the soil of petroleum pollution with to reparation of the water body of petroleum pollution etc.
Claims (11)
1. microbiobacterial agent, this microbiobacterial agent comprises substratum and thalline, it is characterized in that described thalline comprises at least two kinds that are selected from the following bacterial classification: preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797, preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkan ivorans) that preserving number is CGMCC No.4796.
2. microbiobacterial agent according to claim 1, wherein, described thalline comprises that preserving number is the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797, also comprises being selected from least a in the following bacterial classification: preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796.
3. microbiobacterial agent according to claim 1 and 2, wherein, the contained total viable count of the described microbiobacterial agent of every gram is 0.5-1 * 10
10Individual.
4. microbiobacterial agent according to claim 3, wherein, the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 is the 10-90% of total viable count, is preferably 30-70%.
5. the preparation method of the described microbiobacterial agent of claim 1, it is characterized in that this method comprises and will be selected from the Japanese false Xanthomonas campestris that preserving number is CGMCC No.4797 (Pseudoxanthomonas japonensis), preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is that at least two kinds of bacterial classification inoculations in the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and the food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796 are cultivated in substratum.
6. preparation method according to claim 5, wherein, this method comprises with preserving number being that the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCC No.4797 and at least a being inoculated in the substratum that be selected from the following bacterial classification are cultivated: preserving number is the pseudomonas (Pseudomonas) of CGMCC No.4793, preserving number is friendly Gordon Salmonella (Gordonia amicalis) of CGMCC No.4794, preserving number is the Rhodococcus ruber (Rhodococcus ruber) of CGMCC No.4795 and food alkane Gordon Salmonella (Gordonia alkanivorans) that preserving number is CGMCC No.4796; Described cultured method comprises: independently cultivating respectively in the culture system separately, and will cultivate the microorganism that obtains respectively and proportionally mix, the total viable count that makes the every gram microbiobacterial agent that obtains is 0.5-1 * 10
10Individual.
7. preparation method according to claim 6, wherein, by separately independently the cultivation in the culture system and cultivate after mixing, make in the every gram microbiobacterial agent that obtains, preserving number is that the viable count of the Japanese false Xanthomonas campestris (Pseudoxanthomonasjaponensis) of CGMCCNo.4797 is the 10-90% of total viable count, is preferably 30-70%.
8. any application of described microbiobacterial agent in decomposing petroleum hydrocarbon among the claim 1-4.
9. any application of described microbiobacterial agent in the biological restoration petroleum pollution among the claim 1-4.
10. any application of described microbiobacterial agent in the soil of biological restoration petroleum pollution among the claim 1-4.
11. any application of described microbiobacterial agent in the water body of biological restoration petroleum pollution among the claim 1-4.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110169821.4A CN102250801B (en) | 2011-06-22 | 2011-06-22 | Microbial agent, and its preparation method and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110169821.4A CN102250801B (en) | 2011-06-22 | 2011-06-22 | Microbial agent, and its preparation method and application |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102250801A true CN102250801A (en) | 2011-11-23 |
| CN102250801B CN102250801B (en) | 2013-11-06 |
Family
ID=44978373
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201110169821.4A Active CN102250801B (en) | 2011-06-22 | 2011-06-22 | Microbial agent, and its preparation method and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102250801B (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102250770A (en) * | 2011-06-22 | 2011-11-23 | 中国农业科学院农业环境与可持续发展研究所 | Microbial agent and preparation method and application thereof |
| CN103555637A (en) * | 2013-11-15 | 2014-02-05 | 江南大学 | Denitrifying bacterium and culture conditions thereof |
| WO2014135735A3 (en) * | 2013-03-08 | 2014-11-06 | Bio-Iliberis Research & Development S.L. | Biological method for the degradation of complex mixtures of hydrocarbons in an aqueous phase |
| CN104830723A (en) * | 2015-04-30 | 2015-08-12 | 大连民族学院 | Bacterium AW25 with petroleum degradation function, application thereof and bacterial vector |
| CN106957809A (en) * | 2017-05-15 | 2017-07-18 | 中国农业科学院农业环境与可持续发展研究所 | The biological synergetic restorative procedure of soil organic contamination |
| CN108546659A (en) * | 2018-04-12 | 2018-09-18 | 陕西省微生物研究所 | A kind of alkane and degrading polycyclic aromatic hydrocarbons composite bacteria agent and preparation method thereof |
| JP2020103219A (en) * | 2018-12-28 | 2020-07-09 | 学校法人日本大学 | Hydrocarbon treatment agent, hydrocarbon treatment method, and method for producing hydrocarbon treatment agent |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105309371B (en) * | 2015-04-20 | 2017-12-12 | 中国农业科学院农业环境与可持续发展研究所 | The method of circulation in the three-dimensional joint breeding wastewater of one boar fish |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH1133572A (en) * | 1997-07-16 | 1999-02-09 | Unitika Ltd | Method for purifying water or soil contaminated with polycyclic aromatic hydrocarbon |
| CN101603018A (en) * | 2009-07-07 | 2009-12-16 | 南开大学 | Degraded oil, restoring petroleum polluted soil ecology bacteria preparation and preparation method thereof |
-
2011
- 2011-06-22 CN CN201110169821.4A patent/CN102250801B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH1133572A (en) * | 1997-07-16 | 1999-02-09 | Unitika Ltd | Method for purifying water or soil contaminated with polycyclic aromatic hydrocarbon |
| CN101603018A (en) * | 2009-07-07 | 2009-12-16 | 南开大学 | Degraded oil, restoring petroleum polluted soil ecology bacteria preparation and preparation method thereof |
Non-Patent Citations (6)
| Title |
|---|
| 《APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY》 20110131 She, YH "Investigation of Biosurfactant-Producing Indigenous Microorganisms that Enhance Residue Oil Recovery in an Oil Reservoir After Polymer Flooding" 第163卷, 第2期 * |
| 《中国优秀硕士学位论文全文数据库（工程科技Ⅰ辑）》 20091015 崔丽虹 "石油烃降解菌的筛选、鉴定及复合菌群降解效果的研究" , 第10期 * |
| 《生物技术通报》 20090930 崔丽虹 "石油烃降解菌的筛选与鉴定" , 第9期 * |
| SHE, YH: ""Investigation of Biosurfactant-Producing Indigenous Microorganisms that Enhance Residue Oil Recovery in an Oil Reservoir After Polymer Flooding"", 《APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY》 * |
| 崔丽虹: ""石油烃降解菌的筛选、鉴定及复合菌群降解效果的研究"", 《中国优秀硕士学位论文全文数据库（工程科技Ⅰ辑）》 * |
| 崔丽虹: ""石油烃降解菌的筛选与鉴定"", 《生物技术通报》 * |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102250770A (en) * | 2011-06-22 | 2011-11-23 | 中国农业科学院农业环境与可持续发展研究所 | Microbial agent and preparation method and application thereof |
| WO2014135735A3 (en) * | 2013-03-08 | 2014-11-06 | Bio-Iliberis Research & Development S.L. | Biological method for the degradation of complex mixtures of hydrocarbons in an aqueous phase |
| CN103555637A (en) * | 2013-11-15 | 2014-02-05 | 江南大学 | Denitrifying bacterium and culture conditions thereof |
| CN104830723A (en) * | 2015-04-30 | 2015-08-12 | 大连民族学院 | Bacterium AW25 with petroleum degradation function, application thereof and bacterial vector |
| CN106957809A (en) * | 2017-05-15 | 2017-07-18 | 中国农业科学院农业环境与可持续发展研究所 | The biological synergetic restorative procedure of soil organic contamination |
| CN106957809B (en) * | 2017-05-15 | 2020-05-22 | 中国农业科学院农业环境与可持续发展研究所 | Biological synergistic remediation method for soil organic pollution |
| CN108546659A (en) * | 2018-04-12 | 2018-09-18 | 陕西省微生物研究所 | A kind of alkane and degrading polycyclic aromatic hydrocarbons composite bacteria agent and preparation method thereof |
| CN108546659B (en) * | 2018-04-12 | 2021-08-13 | 陕西省微生物研究所 | Alkane and polycyclic aromatic hydrocarbon degradation complex microbial inoculum and preparation method thereof |
| JP2020103219A (en) * | 2018-12-28 | 2020-07-09 | 学校法人日本大学 | Hydrocarbon treatment agent, hydrocarbon treatment method, and method for producing hydrocarbon treatment agent |
| JP7088510B2 (en) | 2018-12-28 | 2022-06-21 | 学校法人日本大学 | Hydrocarbon treatment agent, hydrocarbon treatment method, and hydrocarbon treatment agent manufacturing method |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102250801B (en) | 2013-11-06 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102250798B (en) | Pseudomonas and microbial agent as well as applications thereof | |
| CN102250801B (en) | Microbial agent, and its preparation method and application | |
| CN102250794B (en) | Pseudoxanthomonasjaponensis and microorganism microbial inoculum as well as applications thereof | |
| CN101580808B (en) | Rhodococcus ruber and application thereof in degradation of hydrocarbon compounds | |
| CN102533601A (en) | Bacillus simplex, and culture method and application thereof | |
| EP3800243B1 (en) | Bacterium degrading las and n and use thereof | |
| CN109182178B (en) | Strain with chromium tolerance and Cr (VI) removal capacity and application thereof in-situ remediation of moderately and slightly chromium-polluted soil | |
| CN102994428B (en) | One strain ocean surfactant producing bacteria strain LHOD-1 and application thereof | |
| CN108102979A (en) | A kind of degradation bacteria strains JN5 of oily sludge petrochina hydro carbons and its application | |
| CN110217895A (en) | A kind of complex micro organism fungicide and its application for water environment treatment | |
| CN102250770B (en) | Microbial agent and preparation method and application thereof | |
| CN104087525A (en) | Novel seawater fermentation strain generating biosurfactant | |
| CN102399719A (en) | Bacterium DW3 capable of degrading marine diesel oil pollutants | |
| CN102250797B (en) | Gordoniaamicalis and microbial agent and applications thereof | |
| CN108300674A (en) | A kind of oil degradation bacteria and its preparation method and the application in degrading crude oil | |
| CN102250796B (en) | Rhodococcusruber and microbial agent and application of Rhodococcusruber and microbial agent | |
| CN102250795B (en) | Gordoniaalkanivorans, microbial agent and use thereof | |
| CN111733100B (en) | Acinetobacter scherzei for degrading polyethylene mulching film and application thereof | |
| CN102492637A (en) | Atrazine degrading bacterium | |
| CN101525585B (en) | Bacillus guangzhouensis GIMN1.001 and application thereof | |
| CN104726370A (en) | Enterobacter with crude oil degradation effect and application of enteric bacilli | |
| CN104818234A (en) | Streptomycete with cadmium tolerant characteristic and application thereof | |
| CN107217017A (en) | One plant of acinetobacter calcoaceticus and its application in oil degradation | |
| CN104694439A (en) | Crude oil degradation bacterium and application thereof | |
| CN103074283A (en) | Bacillussp., microbial agent and applications of Bacillussp. and microbial agent |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20220831 Address after: 15th Floor, Enterprise Headquarters Building, Ningdong Energy and Chemical Base, Yinchuan City, Ningxia Hui Autonomous Region, 750409 Patentee after: Ningxia Zhongwei teke Biotechnology Co.,Ltd. Address before: 100081 No. 12 South Main Street, Haidian District, Beijing, Zhongguancun Patentee before: INSTITUTE OF ENVIRONMENT AND SUSTAINABLE DEVELOPMENT IN AGRICULTURE, CHINESE ACADEMY OF AGRICULTURAL SCIENCES |
|
| TR01 | Transfer of patent right |