title: Reading single DNA with DNA polymerase followed by atomic force microscopy.
abstract: The importance of DNA sequencing in the life sciences and personalized medicine is continually increasing. Single-molecule sequencing methods have been developed to analyze DNA directly without the need for amplification. Here, we present a new approach to sequencing single DNA molecules using atomic force microscopy (AFM). In our approach, four surface-conjugated nucleotides were examined sequentially with a DNA polymerase-immobilized AFM tip. By observing the specific rupture events upon examination of a matching nucleotide, we could determine the template base bound in the polymerase's active site. The subsequent incorporation of the complementary base in solution enabled the next base to be read. Additionally, we observed that the DNA polymerase could incorporate the surface-conjugated dGTP when the applied force was controlled by employing the force-clamp mode.
authors.affiliation: School of Interdisciplinary Bioscience and Bioengineering, ‡Department of Chemistry, and §Department of Life Sciences, Pohang University of Science and Technology , San 31 Hyoja-dong, Pohang, 790-784, Korea.
authors.forename: Youngkyu
authors.initials: Y
authors.lastname: Kim
authors.forename: Eung-Sam
authors.initials: ES
authors.lastname: Kim
authors.forename: Yoonhee
authors.initials: Y
authors.lastname: Lee
authors.forename: Joung-Hun
authors.initials: JH
authors.lastname: Kim
authors.forename: Bong Chu
authors.initials: BC
authors.lastname: Shim
authors.forename: Seong Moon
authors.initials: SM
authors.lastname: Cho
authors.forename: Jeong Soo
authors.initials: JS
authors.lastname: Lee
authors.forename: Joon Won
authors.initials: JW
authors.lastname: Park
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism* / Q000648:ultrastructure*; D018625:Microscopy, Atomic Force
chemical_list: D004247:DNA; D004259:DNA-Directed DNA Polymerase