Site-specific DNA recombination events play central roles in many processes related to human health. Immunoglobulin and T-cell receptor diversity, and antigenic variation in parasites and pathogenic microorganisms all depend on genomic rearrangements. The long-term objective of this project is a detailed examination of a DNA molecule that is excised from its chromosomal position when the gene within which it resides is expressed. This unusual 11 kilobase DNA fragment interrupts a highly conserved nitrogen-fixation gene in most nitrogen-fixing filamentous cyanobacteria. The element contains seven open reading frames including a site-specific recombinase, a cytochrome P450 gene (cyp110), and an open reading frame similar to known alkylation-specific DNA repair enzymes. The element is both widely distributed in three cyanobacterial genera and very stable during decades of laboratory culture. A reasonable hypothesis is that the element confers an as yet unknown selective advantage upon its host.
The specific aims of this project are as follows: (a) Determine conditions inducing expression of cyp110 and the putative DNA repair gene. (b) Determine if the wild-type strain out-performs a strain cured of the element under the inducing conditions. (c) Identify the biochemical activities of these two gene products. Funds are requested to support two undergraduate students and one graduate student. Instruction will be provided in recombinant DNA techniques, the use of gene-fusions to monitor gene expression, and how to perform enzyme assays. These skills will be useful in a wide range of biomedical research settings.
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