Size Distribution of the Closed Circular Deoxyribonucleic Acid Molecules of Bacillus megaterium : Sedimentation Velocity and Electron Microscope Measurements

Abstract

The circular deoxyribonucleic acid (DNA) of Bacillus megaterium was fractionated by sedimentation velocity on preparative zonal gradients. The fractions obtained were characterized by analytical sedimentation velocity analysis on neutral and alkaline sucrose gradients, and by contour length measurement by electron microscopy. Each fraction was found to contain circular molecules of one to three discrete sizes, in various combinations of covalently closed and open circular forms. Estimations of the molecular masses of these sizes gave values of 3.9, 6.2, 16.0, 31, and 60 million daltons for the major molecular species. Minor amounts of molecules of 7.6, 11.7, 47, 89, and 112 million daltons were observed in the electron microscope analyses. Length measurements of almost 600 molecules from the various fractions showed that all except six could be placed in distinct size classes. The distribution of molecular sizes in unfractionated circular DNA was shown to consist primarily of the two smallest size classes, although the relative proportions by weight of five of the classes were roughly equivalent.