Figure 3.

Detection of unpaired bases after incubation of a stretched and twisted DNA in glyoxal. Extension vs. torsion curves were obtained at a constant stretching force F between 0.2 and 0.3 pN. When no glyoxal was used (control experiment), the extension displayed a pronounced maximum at σ = 0. The formation of plectonemes on positive or negative twisting led to a rapid shortening of the molecule. When a stretched molecule unwound by n_inc = 200 turns was incubated in glyoxal, the subsequent l vs. σ curve (obtained after washing out the glyoxal) displayed a clear plateau. Its width (≈180 turns) was roughly equal to n_inc, which indicates that, beyond a threshold (≈20 turns, i.e., σ ≈ −0.015), every turn applied to a stretched and unwound molecule induced the melting of one turn of the double helix (10.5 base pairs). Molecular modeling suggests that the structure appearing in overwound DNA (P-DNA) also should have unpaired bases. Incubation in glyoxal of a different stretched and overwound molecule confirmed this prediction. After washing out the glyoxal, the l vs. σ curve displays the same plateau as for the underwound case. However, note that, to get the same plateau required three times more turns being applied to the molecule than in the former case. This is consistent with the measurements of σ_d = −1 and σ_p = +3.