Escherichia coli strain K12 (together with it's plasmids and bacteriophage) has for decades been the most extensively studied bacterium. As a result the genetic and biochemical information available for this strain probably surpasses that for any other living cell. With the new powerful and rapid techniques that have been developed for cloning and sequencing of DNA a complete and exhaustive genetic analysis of this strain has emerged as a desirable and realistic possibility. In this project we propose to begin by dissecting the entire genome into a complete set of ordered clones. The genome of Escherichia coli consists of a circle of about 5 million nucleotide pairs of deoxyribonucleic acid (DNA). The object of this project will be to generate a complete set of about 500 ordered overlapping subfragments, cloned into a bacteriophage vector. We will isolate such a complete collection of ordered subclones, termed an E. coli """"""""gene encyclopedia"""""""", by choosing a subset from a """"""""gene library"""""""" or shotgun collection which is generally understood to refer simply to a random set of clones derived from a genome. The uses to which such a gene encyclopedia could be put include: 1) the rapid mapping of any E. coli gene or mutation; 2) the derivation of clones for over-production of any E. coli gene product; 3) the determination of a complete restriction map of E. coli; 4) to serve as a source of material for the eventual ascertainment of the complete DNA sequence of the genome of E. coli (Project K). Achievement of this later project would provide the first complete specification of the hereditary information needed to define a free living form of life.
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