Rhodococcus erythropolis T7-3 degrading alkane at low temperature and application thereof

A technology of Rhodococcus erythropolis and strains, which can be applied in the fields of microbial biotechnology and environmental biology, and can solve problems such as oil pollution

Inactive Publication Date: 2013-04-17
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The object of the present invention is to provide a low-temperature hydrocarbon-decomposing Rhodococcus erythoropolis (Rhodococcus erythoropolis) T7-3 suitable for low-tempe

Method used

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  • Rhodococcus erythropolis T7-3 degrading alkane at low temperature and application thereof
  • Rhodococcus erythropolis T7-3 degrading alkane at low temperature and application thereof
  • Rhodococcus erythropolis T7-3 degrading alkane at low temperature and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0028] Example 1: Screening of Rhodococcus erythoropolis T7-3 strain provided by the present invention.

[0029] Take mud samples from the seabed area polluted by oil in the Bohai Sea (latitude coordinates are 118°26′36″ east longitude and 38°50′30″ north latitude), and take 5g mud samples in 100mL inorganic salt culture with 2% liquid paraffin as the only carbon source medium (liquid paraffin content 2%), placed on a 200r / min low-temperature shaker at 15°C for enrichment culture for 7 days, and the mixed flora of 7 strains was obtained by streaking and named T7. Aspirate 5mL of the enriched liquid and transfer it to fresh 100mL liquid paraffin medium for fermentation and culture, the culture conditions are the same as above. After three cycles of enrichment, the main strain T7-3 was finally obtained. Take one ring of T7-3 and put it into a test tube filled with 5 mL of LB culture solution, culture it with shaking at 15°C for 48 hours, and use it as a seed solution. Take 1mL...

Embodiment 2

[0032] Example 2: Morphological and physiological and biochemical characteristics of the Rhodococcus erythoropolis T7-3 strain provided by the present invention.

[0033] Refer to the experimental method of "Bergey's Manual of Systematic Bacteriology" (Vol.Ⅷ) to detect its Gram staining, cell size and shape, presence or absence of flagella and spores, growth temperature, growth pH range, and NaCl tolerance. Catalase, nitrate reduction, urease, escin hydrolysis, 2,3-butanediol, aspartic acid, oxidase, M.R., V-P, indole, nitrite reduction, denitrification, gelatin liquefaction, starch hydrolysis , Adenine hydrolysis experiment and the use of mannitol, inositol experiment.

[0034] The results showed that the strain was Gram-positive, and the strain was short rod-shaped, without flagella, immobile, aerobic, and did not produce spores. The cell size was 2.0-3.0 μm (length) × 0.8-1 μm (width), with a typical figure-of-eight arrangement. The growth temperature is 4-34°C, the growt...

Embodiment 3

[0035] Example 3: PCR amplification and sequence determination of the 16S rRNA gene of Rhodococcus erythoropolis T7-3 strain provided by the present invention.

[0036] Inoculate the strain in LB medium, culture it on a shaker at 15°C (130rpm) for 10 hours, collect the bacteria by centrifugation, resuspend, add lysozyme and SDS to break the wall, extract genomic DNA by phenol-chloroform method, and use normal phase primers 27F (5'-GAGAGTTTGATCCTGGCTCAG-3') and reverse primer 1541R (5'-AAGGAGGTGATCCAGCC-3'), use this pair of primers to perform PCR amplification of its 16S rDNA gene, and send the amplified primers to Beijing Sanbo Company for sequencing . The PCR conditions are: 94°C, 10min; 94°C, 45s, 55°C, 45s, 72°C, 90s, 30 cycles; 72°C, 10min, 4°C storage. The length of the 16S rRNA gene sequence is 1470bp, the accession number in GenBank is DQ855406, and the sequence similarity with Rhodococcus erythoropolis (GenBank accession No. AY168586) is 99.93%. The sequence of its ...

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Abstract

The invention relates to a rhodococcus erythropolis T7-3 degrading alkane at a low temperature and an application thereof. The rhodococcus erythropolis T7-3(CGMCC No. 6104) which is one of the cold-resistant bacteria provided by the invention is obtained from a mud sample collected in a sea-bed area polluted by petroleum in the Bohai Sea, wherein the growth temperature is 4-34 DEG C, optimally 15-25 DEG C, the growth pH range is 5-10, and the sodium chloride tolerance is 0-7.5%. The rhodococcus erythropolis can grow with alkane or crude oil as a unique carbon source. The rhodococcus erythropolis has a very high capacity for degrading petroleum hydrocarbon and can degrade C12-36 n-alkanes and crude oil. The degradation rates for 1g/L alkane or 5g/L crude oil added reach over 70%. The degradation rate is 85% at 25 DEG C after shake cultivation for 7 days with 5g/L diesel oil as the unique carbon source, the degradation rate after 7 days of thin oil with 5g/L crude oil as the unique carbon source is 97.05%, and the degradation rate to thickened oil is 61.08%. Therefore, the rhodococcus erythropolis has a good application prospect in sea bioremediation.

Description

technical field [0001] The invention belongs to the field of microbial biotechnology and environmental biotechnology, and in particular relates to a Rhodococcus erythoropolis T7-3 strain and its application in bioremediation of oil pollution in a low-temperature environment. Background technique [0002] With the rapid development of industry, water pollution is becoming more and more serious, and marine pollution has become a worldwide environmental problem of concern. Among the various pollutants in the ocean, oil pollution is the most common and serious one. Petroleum is the main energy source supporting human life today, and its dominant position will hardly be shaken in the foreseeable future. Therefore, the environmental pollution caused by the storage, transportation and processing of petroleum will become increasingly serious. According to the United Nations International Maritime Organization (IMO) statistics, the world's annual oil and its products flow into the o...

Claims

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Application Information

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IPC IPC(8): C12N1/20C02F3/34C12R1/01
Inventor 马挺黄磊李国强
Owner NANKAI UNIV
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