The goal of the proposed work is to demonstrate the feasibility of a novel assay that could have major impact in the area of genomic imprinting in the CNS. Imprinting in the CNS is of potential medical importance considering that a number of known imprinted genes play a role in CNS development, and that certain genetic disorders of the CNS display a pattern of inheritance demonstrating that imprinted genes are involved. Despite the potential importance of these genes, there exists no straightforward assay for their detection and/or discovery. An assay is proposed here for the rapid detection of imprinted genes in CNS tissue that takes advantage of recent advances in microarray technology as well as the availability of mice with uniparental duplication for chromosome regions of interest. As a model system, RNA from CNS tissue of mice carrying maternal or paternal duplication of proximal chromosome 7 will be hybridized to high density microarrays containing a large proportion of all expressed mouse genes. Under optimal conditions, genes that are imprinted should show a differential signal, i.e., maternally expressed genes should be expressed only in mice carrying the maternal duplication, while paternally expressed genes should be expressed only in those with the paternal duplication. Prior to the analysis, conditions for the assay will be optimized by use of custom oligonucleotide microarrays containing probes for known imprinted genes expressed in the CNS, as well as other sequences relevant to CNS function. After analysis of CNS tissue is carried out, the applicability of the method for analysis of specific cell populations will be determined, using identified neurons of the hippocampal cell body layer as a model system. This extension of the assay is especially relevant for analysis of the CNS because of the potentially unique expression pattern of specialized neurons and glia.
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