Essential replication functions of the F plasmid

by Watson, Lynley A.

The work in this thesis concerns the analysis of essential replication genes of the F plasmid of Escherichia coli. The techniques principally employed were the cloning of restriction fragments, transposition mutagenesis, maxicell analysis of proteins, and DNA sequencing. This work led to the following findings. 1. A 1.15kb region of f5 was shown to contain all the functions required for initiation of F replication. Insertions of a transposon within this 1.15kb region abolished mini-F replication. 2. A mini-F plasmid was constructed which constitutes the smallest F derivative so far reported. This plasmid has an elevated copy number, as a result of deletion of the incC region. 3. At least two proteins specified by the essential F replication region were identified in maxicells. The promoter fragments of two mini-F genes were fused to a ?-qalactosidase gene and demonstrated to have low but significant in vivo activities.