Synthetic dendrimers offer multivalent functionality but often require complex chemical synthesis and can exhibit toxicity or poor biocompatibility, limiting their use in biological applications. DNA nanostructures provide a promising alternative, as they are inherently biocompatible and capable of forming higher-order structures. This thesis investigates two methods for assembling large, multivalent DNA nanoparticles: hybridization-driven assembly of DNA-based dendrimers and charge-induced condensation by cationic ligands. For hybridization-driven assembly, tetrahedral DNA nanostructures (TDNs) and Y-shaped DNA were used as monomers. Through complementary sticky-end interactions, monomers were assembled into higher-generation dendrimers, and their structures were characterized using dynamic light scattering (DLS), gel electrophoresis, and atomic force microscopy (AFM). TG1 showed the most consistent assembly, with clear size increases and distinct gel bands. Higher generations (TG2-TG4) displayed predictable size increases by DLS but often showed heterogeneous/smeared bands and partially hybridized generations in the gel, indicating reduced assembly efficiency as structural complexity increases. Y-DNA dendrimers assembled through sticky-end ligation displayed an even lower efficiency, likely due to the short 4-bp sticky-ends and suboptimal ligation conditions. Charged-induced DNA-ligand nanoparticles were explored using the polycations spermine (4+), spermidine (3+), and metformin (1+) at physiological pH. Spermine robustly aggregated TDNs, M13, and rectangular DNA origami at high spermine concentrations and distinct nucleotide-to-spermine ratios, showing consistent thresholds. The presence of Mg2+ suppressed aggregation across all structures, confirming its competitive role in backbone neutralization. Spermine alone was able to show a greater aggregation effect, as the thresholds and spermine concentrations required to form aggregates decreased. In contrast, spermidine and metformin did not reach their thresholds for aggregation under the tested conditions, but metformin instead showed a greater condensation for the long single-stranded scaffold M13. Overall, our results demonstrate that the hybridization of complementary sticky ends can drive the formation of DNA-based dendrimers, but further optimization is required for higher-generation assembly. In contrast, charge interactions between DNA and spermine can form rapid DNA nanoparticle aggregates. By demonstrating two biocompatible routes for constructing multivalent DNA nanoparticles, this work addresses the limitations of synthetic dendrimers, providing a potential alternative for drug delivery, therapeutic applications, and biosensing.
Duessa Red Payumo Bregaudit (Thu,) studied this question.