CN108060094B - Soil composting bacillus and microbial agent and application thereof - Google Patents
Soil composting bacillus and microbial agent and application thereof Download PDFInfo
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Abstract
The invention discloses a soil composting bacillus (Comptobacillus humi) B L G74 CGMCC No. 11219. the soil composting bacillus provided by the invention can grow by taking crude oil as a unique carbon source under the condition of an oil reservoir at 37-60 ℃, degrade crude oil and heavy components and increase the fluidity, thereby improving the oil recovery rate, and moreover, a biosurfactant, a biogas, an organic acid and the like generated by the metabolism of the strain interact with the crude oil, so that the fluidity of the crude oil in the oil reservoir is further increased, and the oil recovery rate is improved.
Description
Technical Field
The invention relates to the technical field of microbial oil production, in particular to soil composting bacillus and a microbial agent and application thereof.
Background
The microbial enhanced oil recovery technology (MEOR) is a biological technology for enhancing the yield of crude oil and the recovery rate of crude oil by using the life activities and metabolites of microbes in oil reservoirs, has the characteristics of low implementation cost, simple operation, remarkable economic benefit and the like, and is widely regarded by people. The method for improving the crude oil recovery rate by utilizing the microorganisms is to take an oil reservoir as a large fermentation field, promote the growth and the propagation of the microorganisms in the oil reservoir by adding bacterial liquid, nutritional agents and microbial metabolites into the oil reservoir, reduce the viscosity of the crude oil by utilizing the direct action of the microorganisms and the crude oil to increase the fluidity of the crude oil, and improve the physical properties of the crude oil by utilizing biogas, surfactants and organic acids generated by the metabolism of the microorganisms, thereby improving the crude oil recovery rate.
Pseudomonas and Bacillus are currently the most studied strains with the ability to emulsify crude oil. The method has good application prospect in the field of oilfield exploitation. Pseudomonas and Bacillus are respectively recorded in application publication numbers CN104371940A and CN102757994A, the growth temperature of the Pseudomonas and the Bacillus which can emulsify crude oil is 30-40 ℃, and the Pseudomonas and the Bacillus cannot be applied to oil reservoirs with higher temperature.
In the process of implementing the invention, the inventor finds that the prior art has at least the following problems: the strain capable of improving the crude oil recovery ratio in the prior art has narrow temperature application range and cannot meet the application of oil reservoirs at higher temperature.
Disclosure of Invention
The invention aims to provide soil composting bacillus and a microbial agent which can meet the requirement of oil reservoir application at higher temperature and application thereof. The technical scheme is as follows:
in a first aspect, the invention discloses a soil composting bacillus (Comptobacillus humi) B L G74. the soil composting bacillus is separated from stratum water of an oil field in North China, is preserved in the general microbiological center of China Committee for Culture Collection of Microorganisms (CCM) 8.10.2015, is named as the soil composting bacillus (Comptobacillus humi) B L G74 with the preservation number of CGMCC No. 11219. the preservation address is No. 3 of Naja district Naja province No.1 of Beijing and the institute of microbiology of China academy of sciences.
According to the invention, a strain of Bacillus compostable soil B L G74 is separated from stratum water of an oil field in North China, and phylogenetic analysis shows that the genetic relationship with the Bacillus composted soil DX-3 is nearest (99.6% similarity), and the Bacillus compostable soil DX-3 represents a new strain of the Bacillus composted soil.
Bacillus agresticola (Comptobacillus humi) B L G74 has the following characteristics:
the soil composting bacillus (Comptobacillus humi) B L G74 grows well on a soybean casein agar culture medium or a solid fermentation culture medium without crude oil, as shown in figure 1, bacterial colonies are in a milky yellow circle shape, are different in size, are 3-4 mm in diameter, are wet in surface and smooth in edge, cells of the strain B L G74 are rod-shaped, are 0.4-0.5 mu m × 2-5 mu m in size, produce spores, are positive in gram staining, grow in a facultative anaerobic manner, the growth temperature of the strain is 37-60 ℃, the pH is 5-9, the NaCl concentration is 0-7% (w/v), and the optimal growth conditions are that the growth temperature is 45 ℃, the pH is 7.5 and the NaCl concentration is 3% (w/v), fructose can be utilized and glucose is not utilized.
The 16S rRNA gene sequence of the bacillus composti (Comptobacillus humi) B L G74 is KT246122 in GenBank accession number, and the gene sequence is shown in a sequence table.
The 16S rRNA gene sequence of the strain B L G74 contains 1387bp, the 16S rRNA gene sequence is aligned in GenBank accession number KT246122 in a GenBank database by a Blast method, MEGA 4 software is adopted to construct a phylogenetic tree of HB-1 and related model strains by an adjacency (Neighbor-Joining) method, as shown in the phylogenetic relationship between the strain B L G74 and adjacent strains in figure 2, the strain B L G74, Compsibacter humi GIESS002 and Compsibacter humi DX-3 are clustered in a phylogenetic branch, the sequence homology is 99.6 percent, and the strain can be determined to be Compsibacter sp, namely Bacillus tumefaciens (Compsibacter humi) B L G74.
It should be noted that the Bacillus soil compost (Compsibacillus humi) is a new genus of high temperature Bacillus reported by Yu et al [ Int J SystEvol Microbiol.2015,65: 346-.
The bacillus composti (Comptobacillus humi) B L G74 can grow and propagate at 37-60 ℃ by taking crude oil as a unique carbon source.
In a second aspect, the present invention discloses a microbial agent comprising any one of the Bacillus agresticola (Comptobacillus humi) B L G74 described above and an activator.
Specifically, the activating agent in the microbial agent comprises the following components:
100 parts of oil reservoir produced water;
0.08-0.12 parts by mass of ammonium chloride;
0.16-0.22 parts by mass of sodium nitrate;
0.04-0.06 parts by mass of monopotassium phosphate;
0.16-0.22 parts by mass of sodium sulfate;
0.01-0.02 parts by mass of magnesium sulfate;
0.04-0.06 parts by mass of sodium carbonate;
0.01-0.02 parts by mass of sodium acetate;
0.04-0.06 parts by mass of at least one of yeast powder, yeast extract powder or yeast extract; and
0.08-0.12 parts of corn steep liquor by mass;
the pH value is 7.5-8;
or
100 parts by mass of water;
0.08-0.12 parts by mass of ammonium chloride;
0.18-0.22 parts by mass of sodium nitrate;
0.03-0.06 parts by mass of dipotassium hydrogen phosphate;
0.008-0.012 parts by mass of magnesium sulfate;
0.8-1.2 parts by mass of sodium chloride;
0.20-0.30 parts by mass of at least one of yeast powder, yeast extract powder or yeast extract; and
0.3-0.6 part of corn steep liquor;
the pH value is 7.0-7.5;
or
100 parts by mass of water;
0.4-0.6 parts by mass of sodium chloride;
0.4-0.6 parts of soybean peptone; and
casein peptone with the mass part of 0.8-1.2 parts;
the pH value is 7.0-8.0.
Preferably, the water is reservoir produced water.
Specifically, the concentration of the Bacillus Agrobacterium tumefaciens (Comptobacillus humi) B L G74 in the activator is 105～106And each m L.
In a third aspect, the invention discloses the use of Bacillus subtilis B L G74 for degrading crude oil.
In a fourth aspect, the use of the microbial agent described above for degrading crude oil.
In a preferred embodiment, the reservoir temperature of the crude oil is 37-60 ℃.
The technical scheme provided by the embodiment of the invention has the beneficial effects that: the soil composting bacillus provided by the invention can grow under the condition of an oil reservoir at 37-60 ℃ by taking crude oil as a unique carbon source, degrade the crude oil and heavy components, and increase the fluidity, so that the oil recovery rate is improved.
Drawings
In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings needed to be used in the description of the embodiments will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings based on these drawings without creative efforts.
FIG. 1 is a colony and thallus morphology diagram of Bacillus agrestis (Comptobacillus humi) B L G74 provided by the present invention;
FIG. 2 is a phylogenetic tree constructed based on the 16S rRNA gene sequence according to the present invention;
FIG. 3 is a graph showing the emulsifying ability of Bacillus agrestis (Comptobacillus humi) B L G74 in a fermentation medium containing crude oil;
FIG. 4 is a graph showing the emulsifying ability of Bacillus agrestis (Comptobacillus humi) B L G74 in a TSA medium containing crude oil;
FIG. 5 is a graph showing the effect of emulsifying liquid paraffin by a fermentation broth of Bacillus subtilis B L G74 according to the present invention;
FIG. 6 is a gas chromatogram of the carbon component of crude oil before the action of Bacillus subtilis B L G74 provided by the invention;
FIG. 7 is a gas chromatogram of the carbon component of crude oil after the action of Bacillus subtilis B L G74 provided by the invention;
FIG. 8 is a diagram showing the analysis of the carbon content of crude oil before and after the action of Bacillus subtilis B L G74 according to the present invention.
Detailed Description
In order to make the technical solutions and advantages of the present invention clearer, the following will describe embodiments of the present invention in further detail with reference to the accompanying drawings.
The experimental methods in the following examples are all conventional methods unless otherwise specified; the experimental materials used in the following examples, unless otherwise specified, were all conventional biochemical reagents and were commercially available.
Example 1
This example provides an isolation, screening and identification of Bacillus composti (Comptobacillus humi) B L G74, comprising the following steps:
The formulas of the used enrichment culture medium containing crude oil and the enrichment agar culture medium are respectively as follows:
the enrichment medium containing crude oil comprises 100m of distilled water L (namely 100g), 0.12g of ammonium chloride, 0.22g of sodium nitrate, 0.06g of monopotassium phosphate, 0.22g of sodium sulfate, 0.02g of magnesium sulfate, 0.06g of sodium carbonate, 0.02g of sodium acetate, 0.06g of yeast powder, 0.12g of corn steep liquor and 10g of crude oil, wherein the pH value of the enrichment medium is 7.5.
The agar-rich medium comprises 100m of distilled water L (namely 100g), 0.08g of ammonium chloride, 0.16g of sodium nitrate, 0.04g of monopotassium phosphate, 0.16g of sodium sulfate, 0.01g of magnesium sulfate, 0.04g of sodium carbonate, 0.01g of sodium acetate, 0.04g of yeast powder, 0.08g of corn steep liquor and 2.0g of agar, and the pH value of the agar-rich medium is 7.5.
And 2, respectively inoculating the bacterial strain which is obtained in the step 1 and takes the crude oil as a carbon source into a fermentation culture medium and a soybean casein (TSA) culture medium which contain the crude oil and are sterilized for 30min at 105 ℃, wherein the liquid loading amount of the culture medium is 250m L, the liquid loading amount of a triangular flask is 100m L, respectively culturing for 72 hours and 7 days (d) in a shaking table with the temperature of 58 ℃ and the rotating speed of 200rpm (revolutions per minute), measuring the surface tension of a fermentation liquid and the viscosity of the crude oil, and obtaining a bacterial strain B L G74 with better emulsified crude oil through detecting the crude oil emulsification and viscosity reduction capability of the bacterial strain, wherein the surface tension of the fermentation liquid is 32.02-34.51 mN/m, and the viscosity reduction rate of the crude oil is 35.6-39.
The formula of the fermentation medium is as follows:
100m of distilled water L (namely 100g), 0.12g of ammonium chloride, 0.22g of sodium nitrate, 0.06g of dipotassium phosphate, 0.012g of magnesium sulfate, 1.2g of sodium chloride, 0.3g of yeast powder and 0.6g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
The formula of the soybean casein (TSA) culture medium is as follows:
100m of distilled water L (i.e., 100g), NaCl 0.6g, soybean peptone 0.6g, casein peptone 1.2g, pH 7.5.
The screened soil composting bacillus (Comptobacillus humi) B L G74 grows well on a soybean casein agar culture medium or a solid fermentation culture medium without crude oil, bacterial colonies are creamy yellow and round, have different sizes, have diameters of 3-4 mm, are wet in surface and smooth in edges, the cells of the bacterial strain B L G74 are rod-shaped, have the size of 0.4-0.5 mu m and × 2-5 mu m, produce spores (the bacterial colonies and the bacterial morphologies are shown in figure 1), are positive in gram staining and grow in a facultative anaerobic manner, the growth temperature of the bacterial strain is 37-60 ℃, the pH value is 5-9, the NaCl concentration is 0-7% (w/v), the optimal growth condition is 45 ℃, the pH value is 7.5, and the NaCl concentration is 3% (w/v).
Example 2
The embodiment provides a method for extracting a bacillus gene of soil compost and constructing a 16S rRNA phylogenetic tree, which comprises the following steps:
reservoir produced water 100ml (i.e. 100 g); 0.1g of ammonium chloride; 0.2g of sodium nitrate; dipotassium phosphate 0.05 g; magnesium sulfate 0.01 g; 0.1g of sodium chloride; 0.25g of yeast powder; 0.5g of corn steep liquor; the pH of the fermentation medium was 7.5.
And 3, performing PCR amplification on the bacterial genes in the step 2, performing quantitative and quality monitoring on the extracted DNA by using a Qubit double-stranded DNA quantitative detection kit (Invitrogen, Carlsbad, CA, USA), and amplifying the V4-V5 region of the 16S rRNA gene by using a primer forward chain (5 '-GTGYCAGCMGCCGCGGTA-3') and a primer backward chain (5 '-CTTGTGCGGKCCCCCGYCAATTC-3').
And 4, determining the gene sequence of the 16S rRNA in the step 2 by utilizing an Illumina MiSeq high-throughput measurement technology, wherein the gene sequence of the 16S rRNA of the strain B L G74 contains 1387bp, the GenBank accession number KT246122 is obtained, and the gene sequence of the 16S rRNA is compared with the sequence in the GenBank to find that the gene sequence has higher sequence similarity with the members of the composting bacillus, wherein the phylogenetic tree (shown in figure 2) constructed by utilizing a Neighbor-Joining method is shown to be a new member of the composting bacillus, has 99.6 percent of similarity with the 16S rRNA (JX274434) of the bacillus composter (composting humi) DX-3, shows that the two are homologous, is a new strain different from the bacillus composter DX-3, and is named as the bacillus composter (composting humi) B L G74.
Example 3
This example provides the effect of a Bacillus soil composter (Comptobacillus humi) B L G74 crude oil emulsification viscosity reduction test comprising the steps of:
100m of distilled water L (namely 100g), 0.08g of ammonium chloride, 0.18g of sodium nitrate, 0.03g of dipotassium phosphate, 0.008g of magnesium sulfate, 0.8g of sodium chloride, 0.2g of yeast powder and 0.3g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
After 3d of incubation at 58 ℃, as shown in fig. 3, the crude oil was significantly emulsified, whereas the crude oil was not emulsified in the control medium of the non-cultured strain B L G74, and the surface tension of the emulsion after the action of the microorganism was measured, and the results are shown in table 1.
100m of distilled water L (i.e., 100g), NaCl 0.4g, soybean peptone 0.4g, casein peptone 0.8g, pH 7.5.
After 3d of incubation at 58 ℃, as shown in fig. 4, the crude oil was significantly emulsified, while the crude oil was not emulsified in the control medium of the non-inoculated strain, and the surface tension and emulsifying ability of the emulsion were measured, and the results are shown in table 1.
Step 4, inoculating the mother liquid of strain B L G74 in an amount of 1% (w/v) to 100m L fermentation medium and soybean casein (TSA) medium, respectively, to which 50G of crude oil was added and sterilized at 105 ℃ for 20min, and measuring the viscosity of the crude oil after culturing at 58 ℃ for 7 days, the results are shown in Table 1.
The surface tension was measured by using JYW-200 surface tension meter (Dingsheng testing machine, Chengde, Hebei province) and removing crude oil from the cultured sample for 72 hours, and measuring the surface tension with the fermentation liquid.
The emulsifying capacity of the culture solution is measured, at room temperature of 25 ℃, the fermentation medium 4m L culture solution after being cultured for 72 hours and 1m L liquid paraffin are placed in a 10m L centrifuge tube, and are vigorously shaken for 1min and then are kept stand for 24 hours, the emulsifying layer is slightly layered (see figure 5), and the emulsifying capacity is more than 90%.
The viscosity of the crude oil is measured by adopting a DV-III + pro Brookfield rotational viscometer, the crude oil is heated to 50 ℃ to be fully melted, a centrifuge is used for centrifuging for 20min at 2000rpm to remove free water, and the temperature is kept constant at 50 ℃ and 6.0rpm for 10min, and then the viscosity of the crude oil is measured.
TABLE 1 results of the emulsification and viscosity reduction experiments
As can be seen from example 3, the present invention provides a Bacillus soil composter (Comptobacillus humi) B L G74 having the effects of emulsifying and viscosity reducing crude oil.
Example 4
This example provides the experimental results of a Bacillus soil composter (Comptobacillus humi) B L G74 for degrading crude oil, comprising the steps of:
100m of distilled water L (namely 100g), 0.1g of ammonium chloride, 0.2g of sodium nitrate, 0.05g of dipotassium phosphate, 0.01g of magnesium sulfate, 1.0g of sodium chloride, 0.25g of yeast powder and 0.5g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
Distilled water 100m L (i.e., 100g), NaCl 0.5g, Soytone 0.5g, peptone 1.0g, and the pH of the TSA medium was 7.5.
And 4, culturing the experimental sample in the step 3 for 20 days, adding 20m L petroleum ether into the culture solution, sufficiently shaking, transferring to a separating funnel, standing, collecting the upper solution, and placing the upper solution into a weighed 50m L centrifuge tube.
And 5, taking 10m L petroleum ether to wash the triangular flask, transferring the washing liquid into a separating funnel, standing, collecting the upper-layer solution, and placing the upper-layer solution into a 50m L centrifugal tube in the step 4.
And 6, washing the separating funnel by using 10m L petroleum ether, and placing the washing liquid into a 50m L centrifugal tube in the step 5.
And 7, putting the weighed solution in the 50m L centrifuge tube in the step 6 at 68 ℃ to evaporate the petroleum ether until the residual petroleum component is dried to constant weight, weighing, and calculating the degradation rate of petroleum according to the formula of H ═ (m ═ m-1－m2)/m1×100％
In the formula: h is the degradation rate (%) of the degrading bacteria to petroleum; m is1Residual oil mass (g) for the control sample; m is2The mass (g) of the degraded residual oil.
Example 5
This example provides the effect of decomposing normal paraffins in crude oil by the action of Bacillus licheniformis (Comptobacillus humi) B L G74 with crude oil, comprising the following steps:
100m of distilled water L (namely 100g), 0.1g of ammonium chloride, 0.2g of sodium nitrate, 0.05g of dipotassium phosphate, 0.01g of magnesium sulfate, 1.0g of sodium chloride, 0.25g of yeast powder and 0.5g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
The culture was incubated at 58 ℃ for 7d and a sample without added inoculum was prepared as a blank.
And 3, measuring and analyzing crude oil components before and after the action of the microorganisms by using a GC-112A gas chromatograph.
The chromatographic column is PEG-20M capillary chromatographic column with 30M × 0.25 and 0.25mm × 0.33 and 0.33 mu M.
The gas chromatography conditions comprise 99.99% nitrogen gas as carrier gas, 300 deg.C of injection port, 40 deg.C of column box temperature, 2 deg.C/min of heating rate, 6 deg.C/min of heating rate after the temperature reaches 200 deg.C, 290 deg.C of heating rate, 20min of holding, split-flow injection with split-flow ratio of 10:1, 1 μ L of injection amount, and N carrier gas2The flow rate was 1.0m L/min.
The analysis results of crude oil components before and after the action of the strain B L G74 are shown in a figure 6 and a figure 7, the relative content of long-chain hydrocarbons (C20-C34) (heavy components) of the crude oil is reduced after the action of the strain B L G74, light components are increased (shown in a figure 8), and the fluidity of the crude oil can be improved after the action of the strain B L G74.
Example 6
This example provides a bacillus soil composter (composibacillus humi) B L G74 biogas assay comprising the steps of:
100m of distilled water L (namely 100g), 1.0g of ammonium chloride, 0.2g of sodium nitrate, 0.05g of dipotassium phosphate, 0.01g of magnesium sulfate, 1.0g of sodium chloride, 0.25g of yeast powder and 0.5g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
Culturing at 58 deg.C for 7d, collecting and measuring the generated gas with gas collecting bag during culturing process, and collecting gas 280m L.
And 3, carrying out gas component analysis on the gas collected in the step 1 by using an HP6890Plus four-valve five-column gas chromatograph. The results of the gas composition measurements are shown in Table 2.
The measurement conditions were: the column temperature is 50 ℃, the injection port temperature is 100 ℃, and the carrier gas is nitrogen.
TABLE 2 gas composition analysis results
The biogas generated by the strain B L G74 can increase the formation pressure, part of the gas is dissolved in the crude oil to swell, the viscosity of the crude oil is reduced, and the fluidity of the crude oil is improved, so that the recovery rate of the crude oil is improved.
Example 7
Taking the mother liquor obtained in step 1 of example 3 as an example, this example provides a Bacillus licheniformis (Comptobacillus humi) B L G74 indoor model test, which can grow and metabolize crude oil as a sole carbon source, degrade crude oil and improve the recovery rate of crude oil, and comprises the following steps:
And 2, firstly carrying out a water flooding test by using the core prepared in the step 1 until the water content is 98 percent, and determining that the water flooding recovery rate is 42.35 percent.
And 3, performing microbial working solution flooding, namely inoculating 1% (w/v) of mother liquor (from the mother liquor obtained in the step 1 in the example 3) of the strain B L G74 into a fermentation medium sterilized at 105 ℃ for 30min, and performing shaking culture in a shaking table at 58 ℃ and a rotating speed of 120rpm for 36 hours to obtain the microbial working solution, wherein the formula of the fermentation medium is as follows:
100m of L (namely 100g) of oil reservoir produced water, 0.1g of ammonium chloride, 0.2g of sodium nitrate, 0.05g of dipotassium phosphate, 0.01g of magnesium sulfate, 1.0g of sodium chloride, 0.25g of yeast powder and 0.5g of corn steep liquor, wherein the pH value of the fermentation medium is 7.5.
The injection amount of the microbial working solution was 1.0 PV.
And 4, placing the core subjected to the micro-flooding at a constant temperature of 60 ℃ for 7 days, performing water flooding until the water content is 98%, and determining the recovery ratio to be 53.78%, wherein the recovery ratio can be increased by 11.43% on the basis of the water flooding by utilizing Bacillus composti (Compsibacter humi) B L G74.
The soil composting bacillus (Compsibacter humi) B L G74 provided by the embodiment of the invention can grow and propagate by taking crude oil as a unique carbon source, metabolizes at the temperature of 37-60 ℃ to generate lipopeptide biosurfactant, biogas such as carbon dioxide, nitrogen and methane, and small molecular organic acid such as acetic acid, propionic acid and butyric acid, the emulsifying capacity of the strain on a fermentation medium reaches above 90%, the degradation rate of the crude oil is 40.8% after the fermentation medium containing 10G of the crude oil is cultured for 20 days, the degradation rate of the crude oil is 61.2% after the TSA medium containing 10G of the crude oil is cultured for 20 days, the surface tension and the crude oil viscosity are respectively reduced by 46.94% and 35.6% after the fermentation medium and the soybean casein medium added with the crude oil are respectively cultured for 72h and 7d, the recovery ratio of the crude oil can be improved by utilizing the soil composting bacillus (Compsibacter humi) B L G54 to improve the recovery ratio on the basis of water flooding, the oil field by utilizing the bacillus (Compsibacter humi) B L, the microbial strain of the invention is a strain which belongs to be a new carbon source of oil field, and the oil produced by adopting the bacterial strain which is screened, and the soil composting strain which can improve the oil field viscosity and the oil field, and the recovery ratio of the soil composting strain which is favorable for improving the oil field.
The above description is only for facilitating the understanding of the technical solutions of the present invention by those skilled in the art, and is not intended to limit the present invention. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
SEQUENCE LISTING
<110> institute of economic and technological of China Petroleum group
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gcggcgcaat agctagttgg tggggtaacg gcctaccaag gcgacgatgc gtagccgacc 240
tgagagggtg atcggccaca ctgggactga gacacggccc agactcctac gggaggcagc 300
agtagggaat cttccgcaat ggacgaaagt ctgacggagc aacgccgcgt gagcgaagaa 360
ggtcttcgga tcgtaaagct ctgttgtcag ggaagaacaa gtacgagagt aactgctcgt 420
accttgacgg tacctgacca gaaagccacg gctaactacg tgccagcagc cgcggtaata 480
cgtaggtggc aagcgttgtc cggaattatt gggcgtaaag cgcgcgcagg cggtccttta 540
agtctgatgt gaaagcccgc ggcttaaccg cggagggtca ttggaaactg gaggacttga 600
gtgcagaaga ggagagtgga attccacgtg tagcggtgaa atgcgtagag atgtggagga 660
acaccagtgg cgaaggcgac tctctggtct gtaactgacg ctgaggcgcg aaagcgtggg 720
gagcaaacag gattagatac cctggtagtc cacgccgtaa acgatgagtg ctaggtgtta 780
gggggtttcc gccccttagt gctgaagtta acgcattaag cactccgcct ggggagtacg 840
gccgcaaggc tgaaactcaa aagaattgac gggggcccgc acaagcggtg gagcatgtgg 900
tttaattcga agcaacgcga agaaccttac caggtcttga catcctctga ccaccctaga 960
gatagggctt tcccttcggg gacagagtga caggtggtgc atggttgtcg tcagctcgtg 1020
tcgtgagatg ttgggttaag tcccgcaacg agcgcaaccc ttgagattag ttgccagcat 1080
tcagttgggc actctagtct gactgccgca gacaatgcgg aggaaggtgg ggatgacgtc 1140
aaatcatcat gccccttatg acctgggcta cacacgtgct acaatggatg gtacaaaggg 1200
cagcgaaacc gcgaggtgga gcaaatccca caaaaccatt ctcagttcgg attgcaggct 1260
gcaactcgcc tgcatgaagc cggaatcgct agtaatcgcg gatcagaatg ccgcggtgaa 1320
tacgttcccg ggccttgtac acaccgcccg tcacaccacg agagttggta acacccgaag 1380
tcggtga 1387
Claims (9)
1. Soil composting bacillus (B)Compostibacillus humi) B L G74 with the preservation number of CGMCCNo.11219.
2. The Bacillus agresticola of claim 1, wherein the Bacillus agresticola can grow and propagate at 37-60 ℃ by using crude oil as a sole carbon source.
3. A microbial agent comprising the bacillus composter according to any one of claims 1 to 2 and an activator.
4. The microbial inoculant according to claim 3, wherein said activator comprises the following components:
100 parts by mass of water;
0.08-0.12 parts by mass of ammonium chloride;
0.16-0.22 parts by mass of sodium nitrate;
0.04-0.06 parts by mass of monopotassium phosphate;
0.16-0.22 parts by mass of sodium sulfate;
0.01-0.02 parts by mass of magnesium sulfate;
0.04-0.06 parts by mass of sodium carbonate;
0.01-0.02 parts by mass of sodium acetate;
0.04-0.06 parts by mass of at least one of yeast powder, yeast extract powder or yeast extract; and
0.08-0.12 parts of corn steep liquor by mass;
or
100 parts by mass of water;
0.08-0.12 parts by mass of ammonium chloride;
0.18-0.22 parts by mass of sodium nitrate;
0.03-0.06 parts by mass of dipotassium hydrogen phosphate;
0.008-0.012 parts by mass of magnesium sulfate;
0.8-1.2 parts by mass of sodium chloride;
0.20-0.30 parts by mass of at least one of yeast powder, yeast extract powder or yeast extract; and
0.3-0.6 part of corn steep liquor;
or
100 parts by mass of water;
0.4-0.6 parts by mass of sodium chloride;
0.4-0.6 parts of soybean peptone; and
casein peptone with the mass portion of 0.8-1.2 parts.
5. The microbial inoculant according to claim 4, wherein said water is reservoir produced water.
6. The microbial agent according to claim 3, wherein the concentration of said Bacillus agrestis in said activator is 105~106And each m L.
7. Use of a bacillus composter according to any one of claims 1 to 2 for degrading crude oil.
8. Use of the microbial agent according to any one of claims 3 to 6 for degrading crude oil.
9. The application of the microbial agent in degrading crude oil according to claim 8, wherein the oil reservoir temperature of the crude oil is 37-60 ℃.
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| CN106811426A (en) * | 2015-11-30 | 2017-06-09 | 中国石油天然气股份有限公司 | Thermal reactor fertilizer Bacillus strain, cultural method and application for emulsified crude oil |
Non-Patent Citations (1)
| Title |
|---|
| Compostibacillus humi gen.nov.,sp.nov.,a member of the family Bacillaceae, isolated from sludge compost;Zhen Yu等;《International Journal of Systematic and Evolutionary Microbiology》;20051231(第65期);第346-352页 * |
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