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.
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 µg 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 µg of the same complex background material were performed to test their sensibility. Finally, the expression analysis of a contaminated groundwater sample was performed.
Project description:RNA from stem containing first node and internodes during dough stage was considered for the above purpose. Satabdi, (popularly known as Minikit) most popular cultivar of West Bengal and Palman, another high yielding variety were considered for microarray analysis for comparison to each other. Two rice genotypes were grown in identical conditions (same field with contaminated ground water) to obtain the most meaningful conclusion. Dough stage was considered as variation in arsenic accumulation among the genotypes starts during dough stage to maturity. Plant tissues were collected from stem for transcriptomic analysis as expression profiling of such tissue in response to arsenic would be most useful as arsenic transport/loading in grain was assumed to be controlled by arsenic translocation behavior at inter vascular level or from xylem to phloem. Satabdi accumulates more than twice of arsenic in brown rice (0.346 mg/kg) than that of Palman (0.156 mg/kg) whereas accumulation in straw (2.07 mg/kg) was approximately less than half of the Palman (4.491 mg/kg)
Project description:A functional microarray targeting 24 genes involved in chlorinated solvent biodegradation pathways has been developed and used to monitor the gene expression in a contaminated site (site B) under ERD (enhanced reductive dechlorination) treatment. The microarray format provided by NimbleGen and used in this study is 12x135K. 4 M-BM-5g of labelled antisense mRNA from 3 groundwater samples were hybridized on the microarray. A 3-chip study was performed, each corresponding to hybridization with 4 M-BM-5g of labelled antisense mRNA retrieved from a monitoring well of a contaminated site (site B). Each probe (760nt) on the microarray was synthesized in eight replicates, and a total of 5,707 random probes was used to determine the background noise. Groundwater samples were collected from a contaminated site (site B) from three monitoring wells (P1, P2 and P3). P1: well located upstream to the contamination source. P2: well in the contamination source. P3 : well located downstream to the contamination source.
Project description:A functional microarray targeting 24 genes involved in chlorinated solvent biodegradation pathways has been developed and used to monitor the gene diversity present in four trichloroethylene (TCE) contaminated sites under ERD (enhanced reductive dechlorination) treatment. The microarray format provided by NimbleGen and used in this study is 12x135K. 2 µg of labelled gDNA from 30 groundwater samples were hybridized on the microarrays.
