Marian Groot Koerkamp, Martijn Rep, Guy Hardy, Adri Mul and Henk Tabak.
Department of Biochemistry, University of Amsterdam, Academic Medical Centre, Meibergdreef 15, 1105AZ Amsterdam.
Genome-wide expression analysis in our lab started with SAGE (Serial Analysis of Gene Expression), a technique that allows estimation of the abundance of each mRNA in a cell culture. SAGE was applied to yeast, comparing expression profiles of yeast grown on different carbon sources (Kal et al 1999). SAGE is now being used by other departments at the AMC to investigate human diseases.
Presently, we are setting up a facility for the production and the analysis of DNA-microarrays, in collaboration with Frank Baas (Neurozintuigen) and Antoine van Kampen (Bioinformatics) at the AMC. Our arrayer is home-built (see http://cmgm.stanford.edu/pbrown/array.html) and can spot with different types of printheads onto filters as well as glass (different needles are used for the two applications). We are now in the process of producing the complete set of yeast ORFs for the production of yeast genome arrays.
Awaiting our home-made filters, we are currently using genome filters produced at the DKFZ in Heidelberg (group of Hoheisel) to compare expression profiles of glucose-grown and oleate-grown yeast cells. Filters are hybridized to 33P-labelled cDNA probes and can be reused easily. The results are scanned on a Storm 860 imager; images are loaded in the Visage HDG-Analyzer program (Bioimage) to run a spot-finding algorithm and quantify spot-intensities. A platform-independent web-interface is being developed to normalize and interrogate the data.
A.J. Kal et al (1999) Mol Biol Cell 10:1859-1872