19 research outputs found
Maximizing Mismatch Discrimination by Surface-Tethered Locked Nucleic Acid Probes via Ionic Tuning
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
Regulating the On-Surface LNA Probe Density for the Highest Target Recognition Efficiency
The
recent emergence of on-surface LNA-based assays as potentially
better alternatives over DNA-based approaches, due to enhanced sensitivity
and target specificity, raises the need for the precise identification
of the factors that control the performance of these assays. In this
work, we investigated whether the probe density of fully modified
ssLNA probes on the gold(111) surface could influence the target recognition
capacity of the LNA sensing layer and illustrated simple means to
control it, primarily by adjusting the salt concentration, nature
of the cation, and pH of the immobilization buffer. It was observed
that monovalent Na<sup>+</sup> could more effectively control the
sensor probe density compared to bivalent Mg<sup>2+</sup>, leading
to better target recognition. Interestingly, unlike in the case of
ssDNA sensor probes, the target recognition efficiency of the LNA
layer at the optimum probe density was found to be almost spacer-independent,
probably due to the rigidity of the LNA backbone. The optimized LNA
sensor layer could discriminate single base mismatches, detect a minimum
target DNA concentration of 5 nM, and sense a significant level of
hybridization within a time scale of a few minutes. To our knowledge,
for the first time, we identify the factors that control the on-surface
LNA probe density for maximizing the performance of the LNA sensing
layer
Ordered Self-Assembled Locked Nucleic Acid (LNA) Structures on Gold(111) Surface with Enhanced Single Base Mismatch Recognition Capability
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
A Review on Chemical versus Microbial Leaching of Electronic Wastes with Emphasis on Base Metals Dissolution
There is a growing interest in electronic wastes (e-wastes) recycling for metal recovery because the fast depletion of worldwide reserves for primary resources is gradually becoming a matter of concern. E-wastes contain metals with a concentration higher than that present in the primary ores, which renders them as an apt resource for metal recovery. Owing to such aspects, research is progressing well to address several issues related to e-waste recycling for metal recovery through both chemical and biological routes. Base metals, for example, Cu, Ni, Zn, Al, etc., can be easily leached out through the typical chemical (with higher kinetics) and microbial (with eco-friendly benefits) routes under ambient temperature conditions in contrast to other metals. This feature makes them the most suitable candidates to be targeted primarily for metal leaching from these waste streams. Hence, the current piece of review aims at providing updated information pertinent to e-waste recycling through chemical and microbial treatment methods. Individual process routes are compared and reviewed with focus on non-ferrous metal leaching (with particular emphasis on base metals dissolution) from some selected e-waste streams. Future outlooks are discussed on the suitability of these two important extractive metallurgical routes for e-waste recycling at a scale-up level along with concluding remarks
Enhancing On-Surface Mismatch Discrimination Capability of PNA Probes by AuNP Modification of Gold(111) Surface
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 <i>T</i><sub>m</sub> 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