Interference of ATP with the fluorescent probes YOYO-1 and YOYO-3 modifies the mechanical properties of intercalator-stained DNA confined in nanochannels

Intercalating fluorescent probes are widely used to visualize DNA in studies on DNA-protein interactions. Some require the presence of adenosine triphosphate (ATP). We have investigated the mechanical properties of DNA stained with the fluorescent intercalating dyes YOYO-1 and YOYO-3 as a function of ATP concentrations (up to 2 mM) by stretching single molecules in nanofluidic channels with a channel cross-section as small as roughly 100?×?100 nm². The presence of ATP reduces the length of the DNA by up to 11 %. On the other hand, negligible effects are found if DNA is visualized with the minor groove-binding probe 4′,6-diamidino-2-phenylindole. The apparent drop in extension under nanoconfinement is attributed to an interaction of the dye and ATP, and the resulting expulsion of YOYO-1 from the double helix.

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Graphical Abstract Nanochannel-stretched DNA (48.5 kbp) stained with YOYO-1 is sensitive to ATP concentration in buffer. Nanochannels with a cross-section of 80?×?80 nm² were used to stretch DNA.