This proposal describes technical approaches for efficient and high throughput production of region specific genomic and cDNA microarrays with the ultimate goal of deciphering cancer-associated genes involved in various human neoplasias. The techniques will be applied to the human 5q31-qter region for feasibility studies. This region and its homologous region in mouse have been shown to be critical in mouse and human hepatocellular carcinomas. We will take two independent approaches to reach the goal of identifying genomic regions harboring genetic alterations and the genes encoded by them. For production of genomic microarrays, we will isolate YAC and BAC contigs in the 5q31-qter region and amplify by inter-Alu primers to obtain a fair representation of this region. We will microarray these amplicons and screen with probes obtained from normal and cancerous tissues (each with different fluorochromes). The ratio of fluorescent intensities will be used to identify amplified and deleted regions of the genome. For development of region-specific cDNA arrays, we ill make use of genomic draft sequencing information to select all coding sequences in the 5q31-qter region. We will array these enriched cDNAs and screen them with cDNA probes derived from normal and cancerous tissues. The high throughput production of genomic and cDNA microarrays will facilitate detailed genetic and transcription mapping of cancer associated regions of the human genome.
Commercially available region specific microarrays will be offered to researchers who concentrate on the regions of genome that are critical in human neoplasias. Therefore, the ultimate goal of this project is to generate commercially available region specific, custom made genomic and cDNA microarrays that can be screened with fluorescently labeled genomic and cDNA probes to facilitate gene mapping and discovery efforts.
