Viruses have protein shells, or capsids, that protect their genomes. During an infection, genomes must be organized, or packaged, into capsids. Genome organization is a hurdle common to all organisms. For example, the human genome is approximately 5 feet long, yet it is organized to fit within a comparatively tiny cell. A similar situation exists with viruses. The project investigates genome organization in single-stranded DNA microviruses. In this virus family, four components ensure proper packaging: the capsid, a small DNA binding protein, a gate keeper protein, and specific genome sequences. The research endeavors to identify how these components operate. In the future, this knowledge could be translated into beneficial applications, such as therapeutic viral vectors. The research contributes to the education of undergraduate, graduate and high-school students. High-school students and teachers conduct 4 to 9-month, data-generating projects, which elicit enthusiasm for science research at an earlier age. Some of the participating schools serve large populations of under-represented groups in science, which further benefits society. Single-stranded viral DNA packaging combines mechanisms found in single-stranded RNA and double-stranded DNA systems. Like double-stranded DNA, single-stranded DNA is packaged into preformed shells but like RNA genomes, they interact extensively with capsids. �X174 packaging is a four-component system: 1) the capsid?s DNA binding pockets, 2) the J protein that guides DNA between pockets, 3) specific DNA sequences, and 4) the A*(star) protein. Structural data suggest that specific coat and J protein residues order the genome into icosahedral symmetry: a hypothesis to be genetically and biochemically tested. To further elucidate protein A* function, the effects of altered protein levels and mutated A* target sites will be determined. Moreover, 60 degenerate, genomic consensus sequences may facilitate genome organization. The consensus sequences will be ?de-optimized? and the resulting effects will be characterized. If ?de-optimization? affects packaging, it may indicate an additional selective pressure operating a single-stranded genomes. The research outcomes will advance our understanding of the basic principles by which single-stranded DNA viruses assemble into infectious particles. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.