Project description:Previous analysis of gene transcript levels of Geobacter species in groundwater during in situ bioremediation of a uranium-contaminated aquifer detected expression of genes encoding superoxide dismutase (sodA) and cytochrome d ubiquinol oxidase (cydA), proteins known to be involved in the response to oxidative stress in other microorganisms. In order to further elucidate gene expression patterns that could be attributed to oxygen exposure, G. uraniumreducens was grown with acetate as the electron donor and fumarate as the electron acceptor in the presence of oxygen and compared to non-oxygen treated cultures.
Project description:HiSpOD is a new efficient functional microarrays probe design algorithm especially dedicated for the microbial ecology and environmental studies. It was used to design 3392 probes targeting 21 genes involved in chlorinated solvent biodegradation pathways and synthesized on a nimblegen microarray. In order to test the probe specificity, the microarray was firstly hybridized to 6 M-BM-5g of labelled aRNA from sheep rumen content (background aRNA). Secondly, hybridization of 1011 copies of labelled aRNA derived from in vitro transcription of three synthetic genes (mmoC, vcrA and tceA) and mixed with 6 M-BM-5g of the same complex background material were performed to test their sensibility. Finally, the expression analysis of a contaminated groundwater sample was performed. A 3 chip study was realized. The first one is a negative control performed with a complex background material (labelled antisense mRNA from sheep rumen content). The second one is a positive control realized with labelled antisense RNA derived from in vitro transcription of three synthetic genes mixed the same complex background material. The third consists in the hybridization of antisense mRNA retrieved from a contaminated groundwater. Each probe (3392) was synthetized in triplicate, and a total of 8,863 random probes was used to determine the background noise.
