Dr. Daniel Hern Paik Profile

Dr. Daniel Hern Paik

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Proceedings Article | 10 October 2012 Paper

Force spectroscopy of DNA: there is still a lot to learn

D. Paik, Thomas Perkins

Proceedings Volume 8458, 845817 (2012) https://doi.org/10.1117/12.945812

KEYWORDS: Molecules, Molecular spectroscopy, Spectroscopy, Single molecule spectroscopy, Optical tweezers, Single mode fibers, Ions, Forward error correction, Biology, Visualization

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Single-molecule studies of the mechanical properties of individual double-stranded DNA have excited interest across many scientific disciplines because of DNA’s fundamental role in biology and DNA’s remarkable overstretching transition at higher forces. Here, we discuss a recent result on the overstretching transition of DNA and on the dynamics of dye molecules intercalating into DNA under tension. Overstretching DNA is mechanical transition whereby DNA’s extension increases by 70% at 65 pN. Notwithstanding more than a decade of experimental and theoretical studies, there remains significant debate on the nature of overstretched DNA. We developed a topologically closed but torsionally unconstrained DNA assay that contains no nicks or free ends. DNA in this assay exhibited the canonical overstretching transition at 65 pN but without hysteresis upon retraction. Controlled introduction of a nick led to hysteresis in the force extension curve. Moreover, the degree of hysteresis increased with the number of nicks. In the second study, we isolated the effects of binding and intercalation of a DNA staining dye, by combining single molecule force spectroscopy with simple buffer exchange. We showed that force-enhanced intercalation can occur from a reservoir of bound dye that was not bis-intercalated, yet remained out of equilibrium with free dye for long periods (<5 min for YOPRO and <2 hr for YOYO). Our work highlights that binding/unbinding and intercalation/de-intercalation are distinct processes that can occur on very different time scales. Taken together, these works highlight ongoing discoveries based on a twenty year old technique, force spectroscopy of single DNA molecules.

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