THE BUOYANT BEHAVIOR OF VIRAL AND BACTERIAL DNA IN ALKALINE CsCl

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Abstract
Denatured DNA increases in buoyant density by about 0.045 density units in alkaline CsCl solution because of the titration of the N[long dash]H protons of the thymine and guanine residues. The pH at the midpoint and the narrow width of the titration curve indicate considerable residual hydrogen bonding in the denatured state at neutral pH. The density of native DNA is not affected until a critical pH> 11 is reached, where the DNA abruptly denatures and increases in density. The change in buoyant density in going from the native material at neutral pH to the fully titrated denatured form at alkaline pH is constant and independent of the base composition. Density gradient centrifugation in alkaline CsCl has been demonstrated to be a useful method for the separation of native and denatured DNA. It may also be useful for the separation of complementary strands. A neutral formaldehyde solution has been recommended as a medium for the study of single stranded DNA''s obtained by denaturation without complications due to reannealing. Alkaline solutions (without formaldehyde) are versatile in that at a pH somewhat below the critical pH for denaturation they are good reannealing media; at a higher pH, they are probably suitable for the study of independent separated strands.