DNA demonstrates a remarkable capacity for creating designer nanostructures and devices. A growing number of these structures utilize Förster resonance energy transfer (FRET) as part of the device's functionality, readout or characterization, and, as device sophistication increases so do the concomitant FRET requirements. Here we create multi-dye FRET cascades and assess how well DNA can marshal organic dyes into nanoantennae that focus excitonic energy. We evaluate 36 increasingly complex designs including linear, bifurcated, Holliday junction, 8-arm star and dendrimers involving up to five different dyes engaging in four-consecutive FRET steps, while systematically varying fluorophore spacing by Förster distance (R0). Decreasing R0 while augmenting cross-sectional collection area with multiple donors significantly increases terminal exciton delivery efficiency within dendrimers compared with the first linear constructs. Förster modelling confirms that best results are obtained when there are multiple interacting FRET pathways rather than independent channels by which excitons travel from initial donor(s) to final acceptor.

The growing maturity of DNA-based architectures has raised considerable interest in applying them to create photoactive light harvesting and sensing devices. Toward optimizing efficiency in such structures, resonant energy transfer was systematically examined in a series of dye-labeled DNA duplexes where donor-acceptor separation was incrementally changed from 0 to 16 base pairs. Cyanine dyes were localized on the DNA using double phosphoramidite attachment chemistry. Steady state spectroscopy, single-pair fluorescence, time-resolved fluorescence, and ultrafast two-color pump-probe methods were utilized to examine the energy transfer processes. Energy transfer rates were found to be more sensitive to the distance between the Cy3 donor and Cy5 acceptor dye molecules than efficiency measurements. Picosecond energy transfer and near-unity efficiencies were observed for the closest separations. Comparison between our measurements and the predictions of Förster theory based on structural modeling of the dye-labeled DNA duplex suggest that the double phosphoramidite linkage leads to a distribution of intercalated and nonintercalated dye orientations. Deviations from the predictions of Förster theory point to a failure of the point dipole approximation for separations of less than 10 base pairs. Interactions between the dyes that alter their optical properties and violate the weak-coupling assumption of Förster theory were observed for separations of less than four base pairs, suggesting the removal of nucleobases causes DNA deformation and leads to enhanced dye-dye interaction.

Because of their ease of design and assembly, DNA scaffolds provide a valuable means for organizing fluorophores into complex light harvesting antennae. However, as the size and complexity of the DNA−fluorophore network grows, it can be difficult to fully understand energy transfer properties because of the large number of dipolar interactions between fluorophores. Here, we investigate simple DNA− fluorophore networks that represent elements of the more complex networks and provide insight into the Forster Resonance Energy Transfer (FRET) processes in the presence of multiple pathways. These FRET networks consist of up to two Cy3 donor fluorophores and two Cy3.5 acceptor fluorophores that are linked to a rigid dual-rail DNA scaffold with short interfluorophore separation corresponding to 10 DNA base pairs (∼34 Å). This configuration results in five FRET pathways: four hetero-FRET and one homo-FRET pathway. The FRET properties are characterized using a combination of steady-state and time-resolved spectroscopy and understood using Forster theory. We show that the multiple FRET pathways lead to an increase in FRET efficiency, in part because homo-FRET between donor fluorophores provides access to parallel pathways to the acceptor and thereby compensates for low FRET efficiency channels caused by a static transition dipole distribution. More generally, the results show that multiple pathways may be used in the design of artificial light harvesting devices to compensate for inhomogeneities and nonideal ensemble effects that degrade FRET efficiency.