Fig. 1. Schematic of experimental setup: (A) diagram illustrating the DNA construct (not to scale). One end of the main DNA unzipping segment (blue) was ligated to complementary DNA oligonucleotides bearing the receptor (HR, dark green) and the cyanuric acid (cy, purple). The other end of the unzipping segment was ligated through a short DNA linker (red) to a dsDNA (2686 bp) labelled with digoxigenin (Dig, green). The 5′ end of the linker was labelled with biotin. The insert shows the structure of the HR–cy couple and the sequence of the complementary DNA oligonucleotides. (B) A single DNA construct was tethered to functionalized beads via Dig-antiDig (green) and biotin-streptavidin (red) connections: one strand of the unzipping segment was attached to a bead (grey sphere) held in the laser trap (dark yellow beams) and the complementary strand to a bead on top of a mobile micropipette (light grey cone). The dsDNA spacer (green) provides separation (∼1 μm) between the two attachment points. Pulling the beads in opposite directions promotes the unzipping of the DNA (Δx) and allows applying controlled, axial mechanical force (F) to the host–guest couple located at one end of the unzipping segment.