Publication

PCR-DGGE method to assess the diversity of BTEX mono-oxygenase genes at contaminated sites

Hendrickx, B., Dejonghe, W., Faber, F., Boenne, W., Bastiaens, L., Verstraete, W., Top, EM. & Springael, D., Feb-2006, In : FEMS Microbial Ecology. 55, 2, p. 262-273 12 p.

Research output: Contribution to journalArticleAcademicpeer-review

  • B Hendrickx
  • W Dejonghe
  • F Faber
  • W Boenne
  • L Bastiaens
  • W Verstraete
  • EM Top
  • D Springael

tmoA and related genes encode the alpha-subunit of the hydroxylase component of the major group (subgroup 1 of subfamily 2) of bacterial multicomponent mono-oxygenase enzyme complexes involved in aerobic benzene, toluene, ethylbenzene and xylene (BTEX) degradation. A PCR-denaturing gradient gel electrophoresis (DGGE) method was developed to assess the diversity of tmoA-like gene sequences in environmental samples using a newly designed moderately degenerate primer set suitable for that purpose. In 35 BTEX-degrading bacterial strains isolated from a hydrocarbon polluted aquifer, tmoA-like genes were only detected in two o-xylene degraders and were identical to the touA gene of Pseudomonas stutzeri OX1. The diversity of tmoA-like genes was examined in DNA extracts from contaminated and non-contaminated subsurface samples at a site containing a BTEX-contaminated groundwater plume. Differences in DGGE patterns were observed between strongly contaminated, less contaminated and non-contaminated samples and between different depths, suggesting that the diversity of tmoA-like genes was determined by environmental conditions including the contamination level. Phylogenetic analysis of the protein sequences deduced from the amplified amplicons showed that the diversity of TmoA-analogues in the environment is larger than suggested from described TmoA-analogues from cultured isolates, which was translated in the DGGE patterns. Although different positions on the DGGE gel can correspond to closely related TmoA-proteins, relationships could be noticed between the position of tmoA-like amplicons in the DGGE profile and the phylogenetic position of the deduced protein sequence.

Original languageEnglish
Pages (from-to)262-273
Number of pages12
JournalFEMS Microbial Ecology
Volume55
Issue number2
Publication statusPublished - Feb-2006

    Keywords

  • BTEX biodegradation, catabolic gene, tmoA, PCR detection, DGGE analysis, GRADIENT GEL-ELECTROPHORESIS, 16S RIBOSOMAL-RNA, CATECHOL 2,3-DIOXYGENASE GENE, AROMATIC-HYDROCARBONS, ENVIRONMENTAL-SAMPLES, TOLUENE DEGRADATION, FUNCTIONAL-ANALYSIS, PHENOL HYDROXYLASE, OXIDIZING BACTERIA, SOIL

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