Our aim is to study the attachment of the Escherichia coli chromosome to the cell envelopes. We propose to pursue our finding that the chromosome appears to be attached at two classes of sites. One class is sensitive to single strand nucleases such as S1. We will try to determine if these enzymes act at specific sites on the DNA by identification of the fragments released using restriction nucleases and hybridization techniques. If this is the case, we will determine if the fragments are released in a special order, and their chromosomal location. We will attempt to gain understanding of their physiological role by studying cells growing at different rates, treated with specific drugs, and at various times in the cell cycle. The other class of DNA-membrane attachment is at or near the replicative origin. We will attempt to purify this complex and characterize its proteins in wild type and various mutants. We will investigate the temporal and physical relationship of this complex to the zone of peptidoglycan growth. This will need the development of novel techniques. Promising mutants in DNA metabolism will be screened for impairment of DNA attachment to the membrane.