DNA–Protein Interactions Studied Directly Using Single Molecule Fluorescence Imaging of Quantum Dot Tagged Proteins Moving on DNA Tightropes

  • Luke Springall
  • Alessio V. Inchingolo
  • Neil M. Kad
Part of the Methods in Molecular Biology book series (MIMB, volume 1431)

Abstract

Many protein interactions with DNA require specific sequences; however, how these sequences are located remains uncertain. DNA normally appears bundled in solution but, to study DNA–protein interactions, the DNA needs to be elongated. Using fluidics single DNA strands can be efficiently and rapidly elongated between beads immobilized on a microscope slide surface. Such “DNA tightropes” offer a valuable method to study protein search mechanisms. Real-time fluorescence imaging of these interactions provides quantitative descriptions of search mechanism at the single molecule level. In our lab, we use this method to study the complex process of nucleotide excision DNA repair to determine mechanisms of damage detection, lesion removal, and DNA excision.

Key words

Single molecule imaging DNA tightropes Quantum dots Diffusion DNA repair Search mechanisms Nucleotide excision repair 

Notes

Acknowledgements

This work was supported by the BBSRC [BB/I003460/1] and [BB/M019144/1] to N.M.K., BHF [FS/13/69/30504] to A.V.I., and University of Kent for L.S.

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Luke Springall
    • 1
  • Alessio V. Inchingolo
    • 1
  • Neil M. Kad
    • 1
  1. 1.School of BiosciencesUniversity of KentCanterburyUK

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