Development of an oligonucleotide microarray to detect di- and monooxygenase genes for benzene degradation in soil.
Diverse environmental genes have been identified recently. To characterize their functions, it is necessary to understand which genes and what combinations of those genes are responsible for the biodegradation of soil contaminants. In this article, a 60-mer oligonucleotide microarray was constructed to simultaneously detect di- and monooxygenase genes for benzene and related compounds. In total, 148 probes were designed and validated by pure-culture hybridizations using the following criteria to discriminate between highly homologous genes: < or =53-bp identities and < or =25-bp continuous stretch to nontarget sequences. Microarray hybridizations were performed using PCR products amplified from five benzene-amended soils and two oil-contaminated soils. Six of the probes gave a positive signal for more than six soils; thus, they may represent key sequences for benzene degradation in the environment. The microarray developed in this study will be a powerful tool for the screening of key genes involved in benzene degradation and for the rapid profiling of benzene oxygenase gene diversity in contaminated soils.
Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo, Tokyo, Japan. siwai@msu.edu
Shoko
S
Iwai
Futoshi
F
Kurisu
Hidetoshi
H
Urakawa
Osami
O
Yagi
Ikuro
I
Kasuga
Hiroaki
H
Furumai
D001419:Bacteria / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001554:Benzene / Q000378:metabolism*; D001673:Biodegradation, Environmental; D017931:DNA Primers / Q000235:genetics; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010802:Phylogeny; D012988:Soil Microbiology; D012989:Soil Pollutants / Q000378:metabolism*
D001426:Bacterial Proteins; D017931:DNA Primers; D012989:Soil Pollutants; D006899:Mixed Function Oxygenases; D001554:Benzene