A spectrophotometric study of the denaturation of deoxyribonucleic acid in the presence of urea or formaldehyde and its relevance to the secondary structure of single-stranded polynucleotides

1. The thermal denaturation of DNA from rat liver was studied spectrophotometrically. In sodium phosphate buffers denaturation led to a single-stranded form having, at 25°, about 25% of the hypochromism of the intact double helix. 2. The hypochromism of the denatured form was the same in 1mm- as in 10mm-sodium phosphate buffer and was scarcely affected by reaction with formaldehyde. The hypochromism was decreased by about 40% in the presence of 8m-urea. 3. The hypochromism of denatured DNA at low ionic strengths was about the same as that of fragments of reticulocyte ribosomal RNA that were too short to form double-helical secondary structure and about the same as that of RNA after reaction with formaldehyde. 4. The spectrum of DNA was slightly affected by the presence of 8m-urea or 4m-guanidinium chloride. The differences in the spectrum of the native and denatured forms of DNA in 0·1m-sodium phosphate buffer, in 8m-urea–10mm-sodium phosphate buffer and in 4m-guanidinium chloride–10mm-sodium phosphate buffer, pH7·6, were similar but not identical. 5. Denatured rat liver DNA appears to have no double-helical character at 25° in 10mm-sodium phosphate buffer, pH7·6; increasing the buffer concentration to 0·1m leads to a more compact form in which about 40% of the residues form base pairs.