Bacterial species, in contrast to human and animals, can be highly diverse with diversity both in gene sequences and presence or absence of genetic material. Strains from the same species can differ in gene content (gene presence or absence) by as much as 20% (1,2). Studies routinely draw potential functional inferences from the distribution of genes among epidemiologically different collections, for example, diseased compared to commensal isolates. This is currently accomplished by membrane-based dot blot ? screenings, a low throughput process prone to inconsistency. We propose creating a bacterial library chip/array to replace the current approach. We will array the genomes of a library of bacterial strains on a single chip that then can be probed for the presence or absence of specific genes and for the allelic variations of the genes. This new form of microarray technology, library array, will allow rapid determination of the potential importance and function of genes or gene fragments and, ultimately, increase our understanding of bacterial pathogenesis and transmission. Specially, we will: ? Aim 1. Determine optimal chip fabrication, probe preparation, and array hybridization conditions ? A. Establish procedures for DNA sample preparation and treatment ? B. Establish procedures for array printing and array hybridization ? Aim 2. Optimize and standardize bacterial library chip production ? A. Determine optimal amounts of targets and probes ? B. Establish quality control and assurance ? C. Standardize data collection and normalization ? Aim 3. Create full-scale library chips for genetic screening ? A. Print a chip each with a library of genomic DNA from 1024 E. coli strains ? B. Test applications using bacterial library array chips ? ?
