Locked nucleic acid (LNA) is a conformationally restricted nucleic acid analogue, which is potentially a better alternative than DNA for application in the nucleic acid based biosensor technologies, due to its efficient and sequence-specific DNA/RNA detection capability and lack of molecule-surface interaction on solid surfaces, compared to DNA. We report, for the first time, a straightforward way (based on simple immersion method) of generating an ordered self-assembled LNA monolayer, which is bioactive, onto a gold(111) surface. This layer is capable of giving rise to a stronger DNA recognition signal (4-4.5 times) than its DNA counterpart, and importantly, it can differentiate between a fully complementary DNA target and that having a single base mismatch, where the mismatch discrimination ratio is almost two times compared to the ratio relevant in case of DNA-based detection. We have presented high-resolution atomic force microscopy (AFM) topographs of the well-defined one-dimensional LNA molecular ordering (few hundred nanometers long) and of the two-dimensional ordered assembly formed over a large area (7 μm × 7 μm) due to parallel positioning of the one-dimensional ordered arrangements. The effects of different parameters such as LNA concentration and incubation time on LNA self-assembly have been investigated. Further, reflection absorption infrared (RAIR) spectroscopy has been applied to obtain information about the orientation of the surface-immobilized LNA molecules for the first time. It has been found that the LNA molecules undergo an orientational transition from the "lying down" to the "upright" configuration in a time scale of few hours.

Several investigations on DNA-based nucleic acid sensors performed in the past few years point toward the requirement of an alternative nucleic acid that can detect target DNA strands more efficiently, i.e., with higher sensitivity and selectivity, and can be more robust compared to the DNA sensor probes. Locked nucleic acid (LNA), a conformationally restricted DNA analogue, is potentially a better alternative than DNA, since it is nuclease-resistant, it can form a more stable duplex with DNA in a sequence-specific manner, and it interacts less with substrate surface due to presence of a rigid backbone. In this work, we probed solid-phase dehybridization of ssDNA targets from densely packed fully modified ssLNA probes immobilized onto a gold(111) surface by fluorescence-based measurement of the "on-surface" melting temperatures. We find that mismatch discrimination can be clearly improved by applying the surface-tethered LNA probes, in comparison to the corresponding DNA probes. We show that concentration as well as type of cation (monovalent and polyvalent) can significantly influence thermal stability of the surface-confined LNA-DNA duplexes, the nature of concentration dependence contradicting the solution phase behavior. Since the ionic setting influenced the fully matched duplexes more strongly than the singly mismatched duplexes, the mismatch discrimination ability of the surface-confined LNA probes could be controlled by ionic modulations. To our knowledge, this is the first report on ionic regulation of melting behavior of surface-confined LNA-DNA duplexes.

Solvation thermodynamic data of dl-serine and dl-phenylalanine in aqueous mixtures of dimethylsulfoxide at 298.15 K were determined from solubility measurement.

No abstract

Unambiguous identification of single base mismatches in nucleic acid sequences is of great importance in nucleic acid detection assays. However, ambiguities are often encountered with, and therefore, a strategy for attaining substantially large enhancement of mismatch discrimination has been worked upon in this study. Short single-stranded peptide nucleic acid (PNA) and deoxyribonucleic acid (DNA) sensor probes that are immobilized onto gold nanoparticle (AuNP) modified Au(111) surface have been applied for target DNA detection. It will be shown that while both PNA and the analogous DNA probes exhibit generally better target detection abilities on the AuNP-modified Au(111) surface (elicited from fluorescence-based measurement of on-surface Tm values), compared to the bare Au(111) surface, PNA supersedes DNA, for all sizes of AuNPs (10, 50, and 90 nm) applied, with the difference being quite drastic in the case of the smallest 10 nm AuNP. It is found that while the AuNP curvature plays a pivotal role in target detection abilities of the PNA probes, the changes in the surface roughness caused by AuNP treatment do not exert any significant influence. This study also presents a means for preparing PNA-AuNP hybrids without altering PNA functionality and without AuNP aggregation by working with the surface-affixed AuNPs.