Separating and screening method for saline-alkaline-resistant bacteria degrading petroleum hydrocarbon

A technology for separation and screening of petroleum hydrocarbon degrading bacteria, which is applied in the field of environmental remediation, can solve the problems of petroleum hydrocarbon degrading bacteria not adapting to the saline-alkali soil environment, affecting the degradation effect of growth active agents, etc., so as to improve the quality of the micro-ecological environment and improve the The effect of micro-ecological environment quality and low treatment cost

Inactive Publication Date: 2015-09-02
NANKAI UNIV
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

Under normal circumstances, petroleum hydrocarbon-degrading bacteria are not well adapted to saline-alkaline soil environment, and saline-alkali stress affects the degradation effect of their growth active agents.

Method used

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  • Separating and screening method for saline-alkaline-resistant bacteria degrading petroleum hydrocarbon
  • Separating and screening method for saline-alkaline-resistant bacteria degrading petroleum hydrocarbon
  • Separating and screening method for saline-alkaline-resistant bacteria degrading petroleum hydrocarbon

Examples

Experimental program
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Effect test

Embodiment

[0020] (1) Strain screening

[0021] Weigh 10g of petroleum-contaminated soil and oil sludge (both collected from Dagang Oilfield), inoculate them into a sterilized 250ml Erlenmeyer flask filled with 90ml of inorganic salt medium by aseptic operation, shake at 30°C and 160rpm Cultivate for 7 days, then inoculate 10 mL of the enrichment solution into fresh crude oil inorganic salt medium, and culture continuously for 4 to 5 times under the same conditions. During the acclimatization process, the pH, salinity and crude oil content of the medium gradually increase. as table 1 shown.

[0022] After the acclimatization was completed, the culture solution was diluted in gradient and spread on LB-agar plates. After the colonies grew out, single colonies of different shapes were picked and purified by streaking to isolate single bacteria. Observe the shape of bacteria under a microscope, Such as figure 1 shown.

[0023] Then pick the single bacteria after separation and purifica...

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Abstract

The invention provides a method for separating and screening a saline-alkaline-resistant pure bacterial strain degrading petroleum hydrocarbon from oily soil. The method comprises the steps: firstly collecting soil polluted by salinized petroleum hydrocarbon from an oil field, adding the collecting soil according to an amount of 10% into an inorganic salt culture medium which takes crude oil as a unique carbon source, and carrying out enrichment culture in a constant temperature shaker; then inoculating a 10% enriching liquid into a fresh crude oil inorganic culture medium and continuously carrying out habituated culture under a same condition for 4-5 times, wherein the pH, salinity and crude oil content of the culture medium in the domestication process are gradually raised; after diluting the culture liquid after domestication in a gradient manner, coating the culture liquid to a LB-agar panel, after bacterial colonies grow, picking up single colonies in different shapes, and lineating and purifying and separating single bacteria; and finally, spreading the separated single bacteria to a crude oil inorganic salt-agar panel and detecting whether the bacterial colonies grow or not, if so, the bacterial strain is the bacteria degrading petroleum hydrocarbon; determining the petroleum hydrocarbon degradation rate of the single bacterial strain and carrying out 16Sr DNA molecular identification on the bacterial strain so as to determine the species relationship of the bacterial strain.

Description

technical field [0001] The present invention relates to a microbial separation and screening method for repairing salinized soil polluted by petroleum hydrocarbons, in particular to a technical method for screening saline-alkaline-resistant petroleum hydrocarbon-degrading bacteria for repairing saline-alkali petroleum-contaminated environments, which belongs to environmental restoration technology field. Background technique [0002] Due to the specific geological conditions and unreasonable human activities, the soils of Dagang Oilfield, Daqing Oilfield, Shengli Oilfield and other oilfields in my country have different degrees of salinization. During oil exploitation and oil well construction operations, it is easy to cause oil leakage and pollute the soil. Saline-alkali soil itself has poor physical and chemical properties, and oil pollution further deteriorates its physical and chemical properties, causing significant stress to soil microorganisms and interfering with so...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12Q1/68C12Q1/04B09C1/10C12R1/01
Inventor 唐景春张海荣孙克静
Owner NANKAI UNIV
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