DNA chip-based Toxicogenomic Analysis and Species Detection
National Research Laboratory on Environmental Biotechnology
Gwangju Institute of Science and Technology (GIST)
1 Oryong-dong, Buk-gu, Gwangju 500-712, Rep. of Korea
e-mail: mbgu@gist.ac.kr
Recent advances in DNA chip technology offer high-throughput screening abilities for transcriptome analysis or specific species detection. DNA- or oligonucleotide-based chip technology is a powerful genomic tool that allows researcher to view the physiological changes and dynamic molecular identifications. For these reasons, DNA chip platform technology is potentially well suited for 1) discriminating potential toxicity with metabolically changes to target organisms and 2) identifying population of microorganisms in the natural complex environments.
1) Cellular stresses caused by various environmental toxic chemicals on Escherichia coli and Oryzias latipes (Japanese Medaka fish) have been evaluated using both DNA chip and real time PCR analyses. In this study, the transcriptional responses of E. coli were studied after exposure to various toxic chemicals, including paraquat, mitomycin C, H2O2, phenolics, dioxins, and others. For example, paraquat was used as a stress inducer to E. coli. In 35 ppm paraquat induction, the most highly expressed genes (soxS, fumC, inaA, hmp) were selected, which means that these genes are highly related with sensing of those kinds of stresses or chemicals. In addition, transcriptome profilings have been conducted for those many chemicals and clustered and reorganized in order to see chemical-gene, gene-gene, or chemical-chemical relations based upon cellular responses to those chemicals. For a toxicogenomic study for fish, phenol EDCs, and pharmaceutics have been used to study the toxicogenomic responses of Medaka fish using both a newly designed and fabricated fish DNA chip and real time PCR analyses. Similar analysis protocols that have been used for E. coli are implemented for the fish data analysis.
2) A new DNA chip for bacterial species identification in the complex environments such as activated sludge process has been successfully developed. Fifty-one probe sets including 16s rDNAs of each bacterium were immobilized on the amine coated glass slides for total 13 different bacteria as a new species detection DNA chip. The probe sets were generated and spotted after whole genomic DNAs were randomly digested using restriction enzyme sets and libraried. For reference signals, all genomic DNAs are primed with cy5-dCTP and each genomic DNA samples to be detected are primed with cy3-dCTP. For hybridization of primed fragments, a common chip hybridization method has been implemented. This method offers fast and reliable sensing schemes for searching or identifying mixed bacterial communities using unsequenced random genomic DNA based DNA chip.