Regensburg 2010 – wissenschaftliches Programm
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BP: Fachverband Biologische Physik
BP 6: Posters: DNA and DNA Enzymes
BP 6.6: Poster
Montag, 22. März 2010, 17:15–20:00, Poster B1
Optical Tweezers Measurements of Threading Individual DNA and DNA-Ligand-Complexes through Solid-State Nanopores — •Andre Spiering1, Andy Sischka1, Tanja Plötz1, Christoph Kleimann2, Katja Tönsing1, Ina Seuffert3, and Dario Anselmetti1 — 1Experimental Biophysics and Applied Nanosciences, Bielefeld University, 33615 Bielefeld, Germany — 2Institut für Bio- und Nanosysteme (IBN-3), Forschungszentrum Jülich, 52425 Jülich, Germany — 3Bioenergetics, Institute of Chemistry, Technische Universität Berlin, 10623 Berlin, Germany
We present a high precision 3D optical tweezers setup, which is incorporated in an optical inverted microscope and uses the detection of backscattered light to measure forces in the sub-pN regime and to manipulate single DNA molecules. With this novel setup, single dsDNA-molecules were threaded into a solid-state nanopore and the electrostatic forces and the ionic currents through the pore were measured simultaneously. In the force-distance diagrams, individual force steps could be observed for each DNA-molecule entering the nanopore and distinct force signals could be identified upon actively withdrawing the single DNA-molecule out of the nanopore. We found that binding of dedicated protein ligands (peroxiredoxin, E. coli RNA-polymerase, and RecA) to dsDNA caused a significant change in the apparent electrostatic forces that are required to thread and unthread the DNA-ligand-complex through the nanopore. Furthermore, we were able to detect the exact position of the binding ligand along the DNA strand with nanometer precision.