Fig. 5: A proof of concept experiment for molecular foot printing.

a Ab-barcode complex with a pre-synthesized oligonucleotide of the complementary strand. Barcode-strand was double-stranded with an oligonucleotide of the complementary sequence of the barcode instead of reverse transcription. b Schematic illustration of the experiment. Protein-G coated magnetic beads binding with the double-stranded Ab-barcode were introduced into a sequencing flow cell and captured on the surface via biotin-avidin interactions (left). After observing the beads, they were washed from the flow cell via a pressure driven flow of 50% formamide solution, while the complementary sequences remained on the surface (middle). Visualization of the complementary sequences remained on the surface by incorporating VTs (right). c Microscopic images corresponding to the schematic illustration shown in b. Blight filed images of protein-G coated beads on the surface (encircled in red, left panel). After washing, the beads were confirmed to have moved from the original position (middle). Fluorescence image of the complementary sequences remained on the surface visualized by VT incorporations (right panel). Scale bars shown in c indicate 5 µm. d Overlaid images of each cycle (green) on the image of the 1st cycle (magenta). Rectangles in orange indicate expected cycles of VT incorporation according to the barcode sequence. Sequencing proceeded from left to right (CTAG) and top to bottom (1Q to 3Q) in this sequential image.