Analysis of mini-F plasmid replication by transposition mutagenesis

Abstract

Derivatives of a mini-F plasmid in which Tn3 is inserted in F deoxyribonucleic acid were obtained, and the sites of insertion for 40 of the derivatives were mapped. Tn3 was found to insert at many sites within mini-F, but most insertions were within the 43.0- to 43.7-kilobase (kb), 44.2- to 44.7-kb, and 45.9- to 46.3-kb segments. Hence, these segments are unnecessary for mini-F replication. Most of the Tn3 derivatives were similar to their parent miniplasmid with respect to copy number, stability, and incompatibility. Insertions at 45.15 kb and near 46.0 kb caused a moderate disruption of copy number control, and insertion at 47.6 kb resulted in unstable maintenance. Deletion derivatives lacking deoxyribonucleic acid between 40.3 and 44.76 kb and between 45.92 and 49.4 kb were obtained. This observation suggests either that mini-F contains a third origin, in addition to those already reported to be at 42.6 and 44.4 kb, or that the reported position of the secondary origin, 44.4 kb, is incorrect and that this origin is between 44.76 and 45.92 kb.