High throughput approaches to biological problems such as a microarray functional genomics study are often expensive and inherently produce large volumes of data. It is therefore imperative that the studies be carried out as efficaciously and economically as possible. In order to meet those goals, three important phases of a toxicogenomics study must be successfully completed (Fig. 1).
Phase I is the experimental design of the microarray experiment. The successful completion of phase I is realized when the microarray results are relevant to the scientific questions asked. We encourage at least one meeting with the researchers to discuss the critical issues regarding the experimental design.
Phase II is the carrying out of the microarray experiment itself to generate the raw data, and we carry out preferred protocols covering RNA isolation and RNA quality control assays, RNA amplification, complementary DNA (cDNA) synthesis, cDNA and amplified RNA (aRNA) labeling, microarray slide production, microarray hybridization and wash conditions, and microarray slide scanning and data generation.
Phase III is the analyses of the voluminous amounts of data generated by a typical microarray experiment. The data analysis and data management is carried out by the Dartmouth Biostatistics and Bioinformatics Shared Resource Core directed by Dr. Jason Moore.
Information of Note
The spotted DNA sample (known sequence) is referred to as the probe, and the fluorescent-labeled cDNA or cRNA (unknown sequence) is referred to as the target.
To determine experimental variability, it is strongly recommended that a given experiment be carried out at least in triplicate and preferably in quadruplicate. For the same reason, it is also recommended that for one of the slides in the triplicate or for two slides of the quadruplicate, the Cy-3 and Cy-5 labeling are switched for the control and experimental target RNAs.