Quantification of catechol 2,3-dioxygenase genes for monitoring bioremediation

摘要

Current techniques for monitoring pollutant-degrading microorganisms are limited to standard culturing methods despite documented deficiencies. Fortunately, techniques based upon detectionof cellular genetic elements can more sensitively monitor microbial populations without requiring culturing. Aerobic bioremediation of aromatic hydrocarbons generally requires the use of ring atacking and ring cleavage dioxygenase enzymes. Two types of ring cleavage dioxygenases exist, meta and ortho cleavage. Meta cleavage dioxygenases are more extensively characterized. Polymerase chain reaction (PCR) primers have been designed using these characterizations that can detect a subclass of dioxygenase genes encoding enzymes responsible for the degradation of toluene, phenol, xylenes, and the single ring metabolites of biphenyl and naphthalene. These primers were used to quantify meta cleavage dioxygenase genes. In pure cultures this technique has been used to detect and enumerate eight different bacteria with various substrate specificities. Through optimization of PCE conditions as few as 10~3 genes were detected, translating to a detection limit of at least 10~3 cells. Overall this technique proves to be more sensitive than standard culturing methods and can be used to more accurately monitor pollutant-biodegrading microorganisms. Thus, bioremediation inthe field can be optimized to more efficiently use resources based upon the response of the appropriate microorganisms.