Rapid microscale procedure for visualizing intracellular plasmid DNA by electron microscopy

A rapid microscale procedure is described that releases plasmid DNA in situ from the bacterial cell and that allows selective observation of the plasmid bound to cellular components. The released plasmid DNA was adsorbed preferentially on mica in a divalent cation-free medium then processed for electron microscopy. The plasmid DNAs studied were pAO3 (1683 base pairs (bp²)), λdv021 (3505 bp), pTSO118 (4000 bp), pAO65 (4786 bp), ColE1 (6500 bp), and RSF2124 (11,400 bp). These DNAs were seen as supercoiled circles or as relaxed circles of corresponding length. Occasionally an internal loop of replicating DNA was present. One micrometer of measured length corresponded to 3100 bp.