Theoretical melting curves were calculated for four DNA restriction fragments, 157-257 base pairs (bp), and a series of hypothetical block DNAs with sequences d(C2*AZC&). d(G&T,G&), 5 I n I 40. These DNAs provided a mixture of A-T/G-C sequence distributions with which to investigate the effects of parameters and base-pair changes on the melting of short DNAs. The sensitivity of DNA melting curves to changes in internal loop melting parameters u and k was examined. As expected, theoretical melting curves of short DNAs with a quasirandom base-pair sequence vary little with changes in internal loop parameters. End melting dominates the transition behavior of these molecules. This was also observed for the block DNAs up to x = 22. Beyond this length, melting curves are highly sensitive to the internal loop parameters. Sensitivity is also predicted for a 157-bp fragment with a block distribution of A-T and G-C pairs. These results indicate that accurate evaluation of internal loop parameters is possible with short DNAs (100-200 bp) containing a G-C/A-T/G-C block distribution with at least 22 bp in each block. Duplex-to-single-strands dissociation parameters were reevaluated from experimental melting curve data of eight DNA fragments using a least squares fit approach. This analysis confirmed parameter values previously found with a simpIified dissociation model. A priori predictions are made on the effects of base-pair changes on the melting curves of three characterized DNA restriction fragments. Single base-pair changes are predicted to induce small but measurable changes in the melting curves. The characteristics of the altered melting curves depend on the location of the base-pair change.
E. coli lactose operon control region, pVH51 plasmid DNA, and A-phage Melting curves of longer DNAs, in the range of 4 6 kilobase pairs (kbp), show more detailed features. Five or more peaks can be resolved in the differential transition curves of pBR322, +X174, Col E l , and fd DNAs (Refs. 8-11, and D. K. Howell and R. M. Wartell, unpublished).
Comparisons between calculated and experimental melting curves of fragments 80-301 bp long show good agreement in 0.1M Na+ concentra-ti0n.5,~ Theoretical calculations have not been able to predict consistently