Figure 1.

Molecular zipper and its denaturation curve. (A) Molecular zipper is a short segment of dsDNA (25 bp), consisting of a positive strand with a quencher at its 5′ end and a negative strand with a fluorophore at its 3′ end. A short ssDNA (23 nt) is attached to the 3′ end of the positive strand, which serves as a reverse primer. The arrow indicates the direction of primer extension. (B) A preannealed molecular zipper with equal amount of negative and positive strands (0.4 μM) were incubated in 25 μl containing 100 mM Tris and 10 mM EDTA (pH 8.3) at 30°C for 10 min and the denaturation curve was determined by measuring the change in fluorescence intensity in a fluorometer (Smart Cycler; Cepheid).