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Petroleum degrading strain and separation method thereof and petroleum degrading microbial agent and preparation method and application thereof

A petroleum-degrading bacteria and petroleum-degrading technology, applied in the field of petroleum pollution control, can solve the problem of less petroleum pollution

Active Publication Date: 2016-04-20
YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there are relatively few reports on the application of oil-degrading bacteria to the actual environment to control oil pollution.

Method used

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  • Petroleum degrading strain and separation method thereof and petroleum degrading microbial agent and preparation method and application thereof
  • Petroleum degrading strain and separation method thereof and petroleum degrading microbial agent and preparation method and application thereof
  • Petroleum degrading strain and separation method thereof and petroleum degrading microbial agent and preparation method and application thereof

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preparation example Construction

[0067] Please refer to Figure 4 , the preparation method of the above-mentioned petroleum degrading bacterial agent of one embodiment, comprises the following steps:

[0068] S210. Under sterile conditions, take the purified petroleum-degrading bacterial strain CN-3 in a test tube slant medium I, and culture it at 30° C. for 24-48 hours to obtain first-grade seeds.

[0069] Among them, culture medium I was sterilized by high pressure steam at 121°C for 20 minutes. The composition of medium Ⅰ is: 55.1gDifco TM MarineAgar2216 medium, 1000mL distilled water, pH 6-8.

[0070] S220. Under aseptic conditions, the primary seeds are taken and inserted into medium II, placed in a shaker at 160 rpm-200 rpm, and cultured at 30° C. for 24-48 hours to obtain secondary seeds.

[0071] Among them, medium II was sterilized by high pressure steam at 121°C for 20 minutes. The composition of medium II is: 37.4gDifco TM MarineBroth2216 medium, 1000mL distilled water, pH 6-8.

[0072] S230....

Embodiment 1

[0082] 1. Preparation of oil-degrading bacterial strains

[0083] (1) Enrichment of petroleum degrading bacteria

[0084] Weigh 10g of Penglai 19-3 oil spill sediment samples into 100mL BHB medium after high temperature sterilization (121℃ for 20min), add 0.5g of petroleum as the only carbon and energy source. Place in a shaker at 180rpm and culture at 30°C for 7 days.

[0085] (2) Separation and purification of petroleum degrading bacteria

[0086] a. Bacteria were isolated by serial dilution method. Take 1 mL of the enriched cultured bacterial solution and add it into 9 mL of sterile 0.9% (m / v) normal saline, and dilute to 10 mL in turn. -2 、10 -3 、10 -4 For gradient dilutions, three groups of parallels were made for each gradient.

[0087] b. Take 100 μL of bacterial solutions of three different gradients and spread them on the BHA medium plate (containing 0.5% petroleum). Incubate for 24 hours in a constant temperature incubator at 30°C.

[0088] c. Select single c...

Embodiment 2

[0104] 1. Preparation of oil-degrading bacterial strains

[0105] (1) Enrichment of petroleum degrading bacteria

[0106] Weigh 10g of Penglai 19-3 oil spill sediment samples into 100mL BHB medium after high temperature sterilization (121℃ for 20min), add 1g of petroleum as the only carbon and energy source. Place in a shaker at 180rpm and culture at 30°C for 10 days.

[0107] (2) Separation and purification of petroleum degrading bacteria

[0108] a. Bacteria were isolated by serial dilution method. Take 1 mL of the enriched cultured bacterial solution and add it into 9 mL of sterile 0.9% (m / v) normal saline, and dilute to 10 mL in turn. -2 、10 -3 、10 -4 For gradient dilutions, three groups of parallels were made for each gradient.

[0109] b. Take 100 μL of bacterial solutions of three different gradients and spread them on the BHA medium plate (containing 1% petroleum). Incubate for 24 hours in a constant temperature incubator at 30°C.

[0110] c. Select single colo...

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Abstract

The present invention discloses a petroleum degrading strain CN-3, which is preserved in China Center For Type Culture Collection, and the accession number is CCTCC M 2015537. A screening medium and a degradation medium used in experiments both use petroleum as a sole carbon source and sole energy, and the screened-out CN-3 strain is highly efficient in petroleum degradation. The petroleum degrading strain CN-3 has good petroleum degradation effect in the neutral to slightly alkaline environment, can withstand high salinity environment, and is especially suitable for the marine environment, so that the range of application of the petroleum degrading strain CN-3 in marine oil pollution treatment is expanded, and the petroleum degrading strain CN-3 has more application prospects.

Description

technical field [0001] The invention relates to the technical field of petroleum pollution control, in particular to a petroleum-degrading bacterial strain and an isolation method, a petroleum-degrading bacterial agent, a preparation method and application thereof. Background technique [0002] Petroleum, also known as crude oil, is a viscous, dark brown liquid. The different compound components in petroleum can be divided into saturated hydrocarbons (including alkanes and cycloalkanes), aromatic hydrocarbons (unsaturated hydrocarbons), non-hydrocarbons (including oxygen, sulfur, nitrogen compounds) and asphaltenes (carbon, hydrogen, oxygen , sulfur, nitrogen and other elements of the complex structure of polymer compounds). [0003] With the continuous development of society, human's demand for energy is increasing day by day. Petroleum is a valuable energy source and chemical raw material, and its popularization and use has brought great convenience to human production a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20B09C1/10A62D3/02A62D101/20
Inventor 胡晓珂陈卫卫孙向楠李俊德
Owner YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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