Figure 5

LightCycler program and effect of program changes. (A) Quadruplicate runs of antiparallel duplex formation at pH 6, 7 and 8 in sodium phosphate buffers with 0.5 μM of each probe and parallel triplex formation at pH 5, 5.25, 5.5 and 5.75 in sodium acetate buffers with 1.0 μM of each probe. Using the same capillaries for 16 runs. The first run and last run were performed at standard conditions. Empty boxes are standard conditions as written in the grey box. AD is antiparallel duplex and PT is parallel triplex. (B) LightCycler standard program. x = dissociation and annealing speed (°C/second), y = hold time (minutes), z = dissociation speed before measurement (°C/second), k = hold before each measurement (minutes).