Investigators' Abstract) Escherichia coli strain K12 (together with its plasmids and bacteriophage) has for decades been the most extensively studied bacterium. The genetic and biochemical information available for this strain probably surpasses that for any other living cell. With the advent of rapid and powerful techniques for sequencing and cloning DNA, the desirable goal of analyzing, in detail, physical, structural and organizational characteristics for the complete genome has become a realistic possibility. In this project, the investigators are studying E. coli as a whole by gathering information that spans the entire genome. What the investigators have developed is a complete ordered clone bank and a global strategy using these clones to detect and map changes in mRNA expression. This will be used to study coordinately regulated genes. Response to a variety of environmental stimuli will be determined at the mRNA level and these genes then studied in detail by subcloning, correlation with proteins expressed, and sequencing. The investigators will map the as yet unmapped heat shock proteins of E. coli. The investigators will also study gene expression in mammalian gut by introducing E. coli K12 into germ-free mice and analyzing changes in mRNA expression relative to culture grown E. coli. The significance of this project lies in the ability to detect global changes at the mRNA level, and correlate them to the genetic map and to existing clones. Thus, the global view and the tools for detailed analysis of molecular biology of the individual responding genes are simultaneously achieved.
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