Abstract: The present invention describes a comprehensive system for gene discovery using retrovirus that have been engineered to exhibit increased accessibility to genomic DNA, or to mutate and identify the chromosomal target sequences of DNA binding proteins. The strategy employs the combination of retroviral integrase/DNA binding protein fusion constructs and gene-trapping methodologies. This novel technology provides the ability to establish proviral integration at any location within the genome. In addition, it allows for the generation of a collection of eukaryotic cells in which each cell contains a mutation in a target gene or sequence for a known DNA binding protein which also allow for rapid in vivo functional analysis. Sequence information obtained for genes identified using the described methods identify a collection of eukaryotic genes related by, or directly or indirectly regulated by, a given DNA binding protein.