Paper BI-TuP15
Asymmetric Hybridization Behavior Exhibited by DNA Probes Containing Surface-Attachment Ligands and Self-Complementary Sequences

Tuesday, November 10, 2009, 6:00 pm, Room Hall 3

We demonstrate the impact that intra-molecular and nucleotide-gold interactions have on conformation of surface-immobilized DNA probes and their hybridization behavior. We take advantage of a method based on the intrinsic affinity of adenine nucleotides for gold (Opdahl et al., PNAS, 104, 9–14, 2007) to immobilize block-oligonucleotides having sequences that follow a d(A_k-T_m-N_n) pattern: a block of k adenines [d(A_k)], followed by a block of m thymines [d(T_m)], and a block of n (arbitrary) nucleotides [d(N_n)]. Such block-oligos assemble on gold via the d(A) blocks, which allow a high degree of control over DNA surface coverage and conformation. We characterize immobilization and hybridization of these DNA probes using x-ray photoelectron spectroscopy (XPS) and surface plasmon resonance (SPR) imaging, specifically to compare two 15-nucleotide N_n sequences: 15 thymines (T15) and a realistic arbitrary sequence (P15). For A15-T5-P15 probes, the hybridization efficiency at the P15 end is enhanced 2-fold compared to that at the A15 end. For A15-T20, which forms a hairpin in solution, we observe a larger asymmetry of hybridization efficiencies when immobilized on gold: while the T15 end is highly accessible for hybridization, the A15 end has virtually no hybridization activity. A thiolated version of the same probe (A15-T20-SH), in contrast, exhibits little asymmetry and overall low hybridization efficiency with either A15 or T15, consistent with an immobilized structure of a stable hairpin. Additional experiments, whereby a displacement thiol is added to reduce DNA-gold interactions, are used to support our inferences about the role played by intra-molecular and surface interactions in immobilization and hybridization of DNA probes.