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Geobacillus stearothermophilus for generating bioemulsifier and application thereof in high-pour-point oil exploitation

A technology of Geobacillus and biological emulsifier, which is applied in the field of biochemical industry and energy mining, can solve the problems of rapid rise of water content, limited development, low recovery degree, etc., achieve good emulsification performance and reduce production cost

Active Publication Date: 2015-06-10
天津林克森生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Judging from the current development effect of high pour point oil reservoirs, there are many problems: water cut rises rapidly, wax precipitation temperature is high (close to formation temperature), recovery degree is not high, crude oil formation viscosity is low, water flooding efficiency is low, Thermal mining costs are higher, etc.
However, this technology also has certain defects or constraints. The cost of some mining technologies is relatively high, and the economic benefits are thus damaged. For example, the hydraulic piston pump oil recovery technology has complex well pipes, high cost, and difficult to meet the sealing degree, which limits develop
At present, in addition to thermal recovery technology, microbial oil recovery technology is one of the economically feasible three recovery technologies for high pour point oil recovery, but the recovery effect is unstable, and further in-depth research is urgently needed to find out a stable strain system and recovery plan

Method used

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  • Geobacillus stearothermophilus for generating bioemulsifier and application thereof in high-pour-point oil exploitation
  • Geobacillus stearothermophilus for generating bioemulsifier and application thereof in high-pour-point oil exploitation
  • Geobacillus stearothermophilus for generating bioemulsifier and application thereof in high-pour-point oil exploitation

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Experimental program
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Effect test

Embodiment 1

[0040] The construction process and effect verification of the bacterial strain A-2 provided by the present invention.

[0041] 1. Construction of expression vector

[0042] According to the complete gene cluster sequence of glycosyltransferase orf 5703 of Geobacillus thermophilia DM-2, using 6.0 software designed the upstream and downstream primers (see Table 1), and the primers were synthesized by Beijing BGI and purified by PAGE. Use the PCR reaction system and method listed in Table 2 to clone the 1359bp target gene fragment, and connect it to the genetic material preservation center derived from Bacillus after double digestion with SacI and PstI according to the reaction system and method in Table 3 On the pUCG18 vector of (BGSC), an expression vector pUCG18-GT with a size of about 7663bp was constructed, electrotransformed into Escherichia coli DH5α, and the plasmid was extracted.

[0043] Table 1 Primers used in experiments

[0044]

[0045] Table 2 PCR reaction ...

Embodiment 2

[0057] The situation that strain A-2 provided by the present invention utilizes 5 different carbon sources to produce emulsifiers.

[0058] The strain A-2 stored in the glycerol tube was activated, spread on LB plates, and cultured at 60°C overnight. Pick a single colony, inoculate into a 5mL LB test tube, incubate at 60°C, 200rpm for 12h. Then inoculate LB Erlenmeyer flask with 1% inoculation amount as seed solution, and cultivate to OD 600 0.5-0.8, that is, in the logarithmic phase, 5% was inoculated into the fermentation medium for emulsifier production, 60°C, 200rpm, and cultivated for 48h.

[0059] The fermentation broth was centrifuged at 4°C and 11000 rpm for 20 minutes to collect the precipitate, then 3 times the volume of pre-cooled acetone was added to the supernatant, and the refrigerator was placed overnight. Then it was centrifuged at 9000 rpm at 4°C for 10 minutes to collect the precipitate, that is, the coarse emulsifier, dissolved in an appropriate amount of ...

Embodiment 3

[0064] Component analysis of the bioemulsifier produced by the bacterial strain A-2 provided by the present invention.

[0065] 1. Qualitative sugar components

[0066] Carry out qualitative sugar analysis on the emulsifier sample extracted in Example 2 to detect whether sugar is contained in the sample. The specific experiment is as follows: the capillary is spotted on a silica gel plate and air-dried naturally. Evenly spread a thin layer of sulfuric acid-anthrone solution on the silica gel plate, air-dry it naturally, heat it in an oven at 105°C for 2 minutes, and develop the color. Further, the phenol-sulfuric acid method was used to detect the total sugar content in the samples, and glucose was used as a standard to prepare a standard curve. Weigh 10 mg of the sample, dilute to 100 mL with distilled water, prepare a 0.1 mg / mL test solution, measure it several times, and calculate the average value of the sugar content. Then the sample is subjected to acid hydrolysis, hig...

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Abstract

The invention relates to Geobacillus stearothermophilus for generating a bioemulsifier and application thereof in high-pour-point oil exploitation. The invention relates to a Geobacillus stearothermophilus gene engineering bacterium A-2 strain of which the collection number is CGMCC No.10151. The Geobacillus stearothermophilus DM-2 strain is a gene engineering bacterium constructed by introducing an expression vector pUCG18 into a self glycosyltransferase gene. The emulsifier yield of the strain A-2 at 45-70 DEG C by using glucose, sodium acetate, ethanol, cetane and olive oil as carbon sources can reach 3-4 g / L, which is enhanced by 6 times or so as compared with the initial strain DM-2. The generated emulsifier can resist high temperature, salt, acid and alkali, has obvious emulsifying effects on n-hexane, diesel oil, xylene, cetane, olive oil and crude oil petroleum hydrocarbon, and has obvious emulsifying, visbreaking and pour point depressing effects on high-pour-point oil. The field experiment proves that the strain A-2 and bioemulsifier generated by the strain A-2 can be used for high-pour-point oil reservoir exploitation.

Description

technical field [0001] The invention relates to the technical fields of biochemical industry and energy exploitation, in particular to a genetically engineered bacterium with high yield of biological emulsifier and its application in high pour point oil exploitation. Background technique [0002] Biosurfactants are a class of amphiphilic substances produced by animals, plants, and microorganisms, with hydrophobic groups and hydrophilic groups. Biosurfactants can be divided into two categories according to their molecular weight: one is low-molecular-weight surfactants, such as glycolipids, phospholipids, fatty acids, short-chain peptides, etc., and their activity is mainly to reduce the oil-water interfacial tension The other type is mostly biological macromolecules, also known as emulsifiers, which generally do not reduce the oil-water interfacial tension very low, but have a strong affinity for the oil-water interface and can be adsorbed on the surface of dispersed oil dro...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/54C09K8/582C12R1/01
Inventor 马挺周洁芳史忠李国强刘云高配科石慧
Owner 天津林克森生物科技有限公司
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