This core will provide expertise in the production and use of cDNA and oligonucleotide microarrays and the bioinformatics support required for data storage and analysis. The investigator provides the core laboratory with a minimum of 1 microgram of total RNA from control and test cell samples; core personnel produce the target RNA, the microarrays, the hybridization, the image analysis, and preliminary data analysis. cDNA arrays are produced by printing PCR products from IMAGE consortium cDNA clones onto glass microscope slides using a commercial 3-axis robotic array similar to the Stanford design. Target total RNA (1 microgram) is prepared by RT-PCR using Oligo-dT and random primers in the presence of Aminoallyl-dUTP. The cDNA is dye-labeled by coupling NHS-ester Cys3 or Cy5 dyes to free amines. The labeled cDNA is hybridized to the microarray in slide chambers or in a GeneTAC hybridization workstation and scanned using a scanning laser confocal microscope (SanArray 5000, GSI Luminics, Inc.). The image data files are analyzed using IPlab (Scanalytics, Inc.) And the resulting data transferred to ArrayDB (NHGIR, Oracle version x.y). Investigators retrieve the DNA clones, 9,200 human clones, and approximately 21,000 human ESTs. We are currently producing arrays with approximately 3,600 features/slide. Additional clones and higher density arrays will be produced as they become available and as our expertise increases. The core also has the complete complement of Affymetrix instrumentation and bioinformatics tools available and currently has human """"""""HuGeneFL"""""""", mouse """"""""Mu11K"""""""", and human """"""""Cancer G110"""""""" GeneChips in stock for immediate use. Finally, the core also has access to significant bioinformatics resources in terms of computational hardware, expertise, and software/database resources. This element of the laboratory is critical to the successful use and evaluation of DNA microarray experiments.
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