Transposition of a plasmid deoxyribonucleic acid sequence that mediates ampicillin resistance: independence from host rec functions and orientation of insertion

Abstract

Insertion of the transposable DNA sequence that specifies the TEM .beta.-lactamase (TnA) occurred in at least 19 sites on the 5.5 .times. 106-dalton plasmid RSF1010. There was no significant difference in the frequency of transposition or in the distribution of TnA insertion sites for recombinant plasmids isolated from recombination-proficient (rec+) or recombination-deficient (rec-) bacterial [Escherichia coli] host cells. The site and orientation of TnA insertions were determined by heteroduplex analysis and enzymatic digestion with [Eco RI, B am HI and Hin II] restriction endonucleases. Insertions in the gene encoding for sulfonamide resistance occurred without circular permutation in 1 or the other of 2 distinct orientations. Insertions in orientation P were strongly polar on distal gene expression, but insertions in orientation M were mutagenic but not polar. TnA elements from different R plasmids show fine structural heterogeneity, and TnA insertion at a site adjacent to the origin of replication causes an increase in plasmid copy number.