Structural Properties of Oligonucleotide Monolayers on Gold Surfaces Probed by Fluorescence Investigations

Abstract

We present optical investigations on the conformation of oligonucleotide layers on Au surfaces. Our studies concentrate on the effect of varying surface coverage densities on the structural properties of layers of 12- and 24mer single-stranded DNA, tethered to the Au surface at one end while being labeled with a fluorescent marker at the opposing end. The distance-dependent energy transfer from the marker dye to the metal surface, which causes quenching of the observed fluorescence, is used to provide information on the orientation of the DNA strands relative to the surface. Variations in the oligonucleotide coverage density, as determined from electrochemical quantification, over 2 orders of magnitude are achieved by employing different preparation conditions. The observed enhancement in fluorescence intensity with increasing DNA coverage can be related to a model involving mutual steric interactions of oligonucleotides on the surface, as well as fluorescence quenching theory. Finally, the applicability of the presented concepts for investigations of heterogeneous monolayers is demonstrated by means of studying the coadsorption of mercaptohexanol onto DNA-modified Au surfaces.

Independent control of grafting density and conformation of single-stranded DNA brushes

“…the effect to the larger electrostatic and entropic forces that must be overcome to uncoil and pack the longer d(T 25 ) brush strands ( Fig. 1), similar to the trend observed for films of thiolated DNA (15,19,24). As the d(T) length increases, the saturation grafting density is determined not solely by the length of the d(A) blocks, but also by the polyelectrolyte effects in the d(T) brush.…”

“…Passivation prevents nonspecific interactions between the surface and DNA or other biomolecules, which enables widely spaced DNA probes to maintain an upright orientation. In a common implementation of this strategy, films of thiol-anchored DNA can be exposed to a solution containing a competing molecule, such as mercaptohexanol (MCH), which displaces most of the DNA from the surface and forces the remaining DNA strands into an upright conformation (10,(17)(18)(19). Alternatively, grafting densities can be adjusted by coupling functionalized DNA to a bifunctional self-assembled monolayer (6).…”

Nucleobase Orientation and Ordering in Films of Single-Stranded DNA on Gold

“…16 In contrast, the (dT) 25 -SH strands form mostly disordered films similar to polyelectrolyte brushes. 17 In summary, we demonstrate how the structure, orientation, and ordering in ssDNA monolayers can be quantitatively determined by probing electronic and vibrational states of nucleobases. The simplest interpretation of the agreement between structures observed in UHV and ambient is that both must reflect some aspects of the initial structure in solution.…”

Distance-Dependent Emission from Dye-Labeled Oligonucleotides on Striped Au/Ag Nanowires: Effect of Secondary Structure and Hybridization Efficiency

“…Rant, Tornow, and co-workers used this effect to probe conformational changes in 5′ thiolated, 3′ dye labeled 12-and 24-base single-stranded oligonucleotides. 48 Fluorescence increased with increasing DNA surface density, due to steric interactions forcing the strands to "stand up" to a greater extent on the surface.…”