| As the amount of genomic sequence information continues to increase at an incredible pace, so does the need for high throughput global strategies to begin assigning biological meaning to these data. This dissertation describes the development of a high throughput genetic approach for studying bacterial genomes. Specifically, a method has been developed for using custom-designed full genome oligonucleotide micorarrays to analyze pools of Tn5 transposon mutants grown under different conditions. Biotin-labeled RNA transcripts generated from the ends of each transposon insertion are hybridized to high density oligonucleotide microarrays. Transposon insertion locations can be mapped to within 50 bp of their genomic location as determine by sequencing.; Although initially developed in Escherichia coli K-12 and used to study in vitro growth phenotypes in rich and minimal media, this technique has also been applied to an in vivo pathogenesis system in an effort to identify genes in uropathogenic E. coli strain CFT073 that are required for colonization of the mouse urinary tract. In a pilot study, four pools of 48 mutants each have been screened in an ascending urinary tract infection model using Swiss-Webster mice. Nine genes were identified as encoding potential virulence factors. This work shows the development of a high throughput strategy that can be used to study both in vitro and in vivo phenotypes of pooled transposon mutants. |
