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Realising quantitative dynamic atomic force microscopy to probe
transactions of DNA at the single molecule level
Neil H Thomson1,2
1
2
Department of Oral Biology, School of Dentistry
Molecular and Nanoscale Physics Group,
School of Physics and Astronomy, University of
Leeds, LS2 9JT, United Kingdom.
The dynamic modes of atomic force microscopy (dAFM), where the force sensing cantilever is oscillated at or
close to resonance, have been essential for atomic force microscopy to reliably image and measure soft
biological samples, from single molecules to cells. Extracting quantitative information has been hampered by a
number of problems, including understanding the dynamics of the oscillating cantilever in the non-linear tipsample potential and the response of the cantilever from the integration of forces of different origin (sign and
length scale) by the AFM tip.
The first part of the seminar will summarise a number of key advances our group has made towards making
AFM measurements quantitative. It will demonstrate that loss of height at the nanoscale is a consequence of
the intrinsic convolution between the force fields of the tip and sample [1]. We show for DNA that on
hydrophilic surfaces such as mica, even in ambient conditions, there is sufficient water present to retain
structural hydration of biomolecules [2]. Building on these two outcomes, we have been able to measure the
hydrophilicity of individual DNA molecules [3]. Furthermore, we have implemented a new small amplitude
method that maximises resolution and can resolve the right-handed DNA double helix [4].
The second part of the seminar will concentrate on the use of amplitude modulation (AM AFM) imaging
applied to biomolecular samples in ambient conditions, in particular, DNA-protein complexes. We have studied
transcription of DNA by AFM for a number of years [5] and are investigating dual RNA polymerase transactions
on single DNA templates in the context of the twin supercoiling domain paradigm. The outcomes have
implications for compressed genetic structures found in vivo and are giving foundation to understanding the
“rules of the road” for these molecular motors that mediate gene expression.
Keywords: DNA, RNA polymerase, transcription, twin supercoiling domain model, gene expression, nested
genes, AFM, mica, humidity, polymer chain statistics
[1] Santos S., Barcons V., Christenson H.K., Font J. and Thomson N.H. (2011)
“The intrinsic resolution limit in the atomic force microscope: implications for heights of nanoscale features.”
PLoS ONE 6 (8): e23821.
[2] Billingsley D.J., Kirkham J., Bonass W.A, and Thomson N.H. (2010)
“Atomic force microscopy at high humidity: irreversible conformational switching of supercoiled DNA molecules.”
Phys. Chem. Chem. Phys. 12 (44), 14727 - 14734
[3] Santos S., Stefancich M., Hernandez H., Chiesa M. and Thomson N.H. (2012)
“The hydrophilicity of a single DNA molecule.” J. Phys. Chem. C 116 (4) 2807-2818.
[4] Santos S., Barcons V., Christenson H.K., Billingsley D.J., Bonass W.A, Font J. and Thomson N.H. (2013)
“Stability, resolution and ultra-low wear amplitude modulation atomic force microscopy of DNA: small amplitude small
set-point imaging.” Applied Physics Letters 103, 063702.
[5] Billingsley D.J., Bonass W.A, Crampton N., Kirkham J. and Thomson N.H. (2012)
“Single molecule studies of DNA transcription using atomic force microscopy.” Physical Biology 9, 021001.