DNA Molecules In Aqueous Solution Research Articles

Label-Free Detection of C-T Mutations by Surface-Enhanced Raman Spectroscopy Using Thiosulfate-Modified Nanoparticles.

Recent developments in molecular spectroscopy have widened the scope of surface-enhanced Raman spectroscopy (SERS) for detection of nucleic acids. In order to solve the interference of impurity signals in SERS analysis that hamper the reliable detection of DNA, Ag nanoparticles modified with thiosulfate ions were used to obtain SERS signals of DNA molecules in aqueous solutions, which showed good reproducibility. By using thiosulfate ions and calcium ions as aggregating agents, this method not only eliminated the influence of citrate on DNA signals completely but also obtained the signals for all bases indiscriminately, including the T base that was considered to have low Raman activity. Subsequently, the base stacking rule was used to identify mutations arising from C/T transition. It further identified the mutation sites of single-base C/T transition using this platform for the first time. This method has wide application prospects in DNA analysis, DNA sequencing, and genetic testing.

EIS Biosensor for Detection of Low Concentration DNA Molecules

The results of the theoretical simulation of an electrolyte-insulator-semiconductor (EIS) biosensor made on a silicon nanowire for the detection of low concentration DNA molecules are presented. The equivalent electrical scheme for the EIS structure is constructed. The phenomenon of charge distribution in a depletion layer of semiconductor is taken into account in computations. The behavior and dependence of the total capacitance for the EIS sensor and its capacitive sensitivity on the DNA concentration in aqueous solution are investigated and analyzed. It is shown that the capacitance of the EIS sensor is very sensitive to the presence of the DNA molecules and the sensitivity has comparatively high value at the low concentration of the DNA molecules. The increase of the DNA concentration results in the decrease of the EIS total capacity. It is shown that one can register and determine the number of charged DNA molecules in aqueous solution by measuring the changes in the EIS capacity. The sensitivity increases with the growth of the gate voltage. It is shown that the threshold of sensitivity can be very low and the signal-to-noise ratio can reach high values for the EIS biosensor.

Noises and Signal-to-Noise Ratio of Nanosize EIS and ISFET Biosensors

The results of comparative theoretical analyzes of the behavior of internal low-frequency noises, signal-to-noise ratio and sensitivity to DNA molecules for EIS and ISFET based nanosize biosensors are presented. It is shown that EIS biosensor is more sensitive to the presence of DNA molecules in aqueous solution than ISFET sensor. Internal electrical noises level decreases with the increase of concentration of DNA molecules in aqueous solution. In the frequency range 10−3 - 103 Hz noises level for EIS sensor about in three orders is higher than for ISFET sensor. In the other hand, signal-to-noise ratio for capacitive EIS biosensor is much higher than for ISFET sensor.

ISFET Based DNA Sensor: Current-Voltage Characteristic and Sensitivity to DNA Molecules

Dependency of both source-drain current and current sensitivity of nanosize ISFET biosensor vs. concentration of DNA molecules in aqueous solution theoretically is investigated. In calculations it is carried out effects concerning charge carriers distribution in current channel and concerning carriers’ mobility behavior in high electrical fields in the channel. The influence of DNA molecules on the work of ISFET biosensors is manifested by a change in the magnitude of the gate surface charge. Starting with fairly low concentrations of DNA, ISFET sensors respond to the presence of DNA molecules in an aqueous solution which is manifested by modulation of channel conductance and therefore the source-drain current changes of the field-effect transistor. It is shown that the current sensitivity with respect to concentration of DNA molecules linearly depends on the source-drain voltage and reaches high values.

Naturally occurring branched-chain polyamines induce a crosslinked meshwork structure in a giant DNA

We studied the effect of branched-chain polyamines on the folding transition of genome-sized DNA molecules in aqueous solution by the use of single-molecule observation with fluorescence microcopy. Detailed morphological features of polyamine/DNA complexes were characterized by atomic force microscopy (AFM). The AFM observations indicated that branched-chain polyamines tend to induce a characteristic change in the higher-order structure of DNA by forming bridges or crosslinks between the segments of a DNA molecule. In contrast, natural linear-chain polyamines cause a parallel alignment between DNA segments. Circular dichroism measurements revealed that branched-chain polyamines induce the A-form in the secondary structure of DNA, while linear-chain polyamines have only a minimum effect. This large difference in the effects of branched- and linear-chain polyamines is discussed in relation to the difference in the manner of binding of these polyamines to negatively charged double-stranded DNA.

The effect of a low-frequency electromagnetic field on DNA molecules in aqueous solutions

A chemiluminescence study showed that hepatitis B virus (HBV) and hepatitis C virus (HCV) DNA amplicons are capable of induced radiation when exposed to electromagnetic fields (EMFs) that range from 7.5 to 30 Hz in frequency and from 24 to 40 A/m in field strength. An EMF with a frequency of 9 Hz was shown to exert the greatest effect on aqueous solutions of the hepatitis virus DNA amplicons. The hydration shell of the DNA amplicons was observed to change. The change in the DNA hydration shell on exposure to a low-frequency EMF was presumed to restore hydrogen bonds, to induce crosslinks, and to facilitate DNA repair.

Molecular-scale quantitative charge density measurement of biological molecule by frequency modulation atomic force microscopy in aqueous solutions

Surface charge distributions on biological molecules in aqueous solutions are essential for the interactions between biomolecules, such as DNA condensation, antibody–antigen interactions, and enzyme reactions. There has been a significant demand for a molecular-scale charge density measurement technique for better understanding such interactions. In this paper, we present the local electric double layer (EDL) force measurements on DNA molecules in aqueous solutions using frequency modulation atomic force microscopy (FM-AFM) with a three-dimensional force mapping technique. The EDL forces measured in a 100 mM KCl solution well agreed with the theoretical EDL forces calculated using reasonable parameters, suggesting that FM-AFM can be used for molecular-scale quantitative charge density measurements on biological molecules especially in a highly concentrated electrolyte.

Quaternized Poly[3,5-bis(dimethylaminomethylene)hydroxystyrene]/DNA Complexes: Structure Formation as a Function of Solution Ionic Strength

The formation of complexes between the cationic polyelectrolyte poly[3,5-bis(dimethylaminomethylene)hydroxystyrene] (Q-N-PHOS), bearing two cationic sites per repeating unit, and DNA molecules in aqueous solutions is investigated at pH 7 and at various salt (NaCl) concentrations and DNA/polymer ratios. The structural characteristics of the polyplexes at different DNA/polymer ratios were characterized in terms of mass, size, and charge using static, dynamic, and electrophoretic light scattering and fluorescence spectroscopy. The results indicate that the complexes have a loose spherical morphology with size in the range of 45-100 nm depending on polymer to DNA ratio and ionic strength of the solution. Most interestingly, it is found that the polyplexes' response to changes in the ionic strength of the surrounding solution after complexation depends strongly on the initial solution ionic strength during complex formation. This also points to differences in the nanostructures of polyplexes formed at different ionic strengths.

A simple and effective sample preparation method for atomic force microscopy visualization of individual DNA molecules in situ

A simple, controllable and effective sample preparation method was established for atomic force microscopy (AFM) imaging of individual DNA molecules in aqueous solution. Firstly, magnesium ion (Mg(2+)) at a concentration of 5.0-10.0 mM as a positively charged bridge was transferred onto mica to immobilize DNA molecules. Then Mg(2+)-modified mica was used to investigate DNA molecules in any buffer without magnesium ion by AFM. AFM images demonstrated that DNA molecules can be successfully observed in solution with good resolution, reproducibility, and stability. Further, this DNA sample preparation method makes AFM successful to investigate DNA molecular interaction in situ and DNA/chitosan complex in gene delivery.

Priming with a very low dose of DNA complexed with cationic block copolymers followed by protein boost elicits broad and long-lasting antigen-specific humoral and cellular responses in mice

Cationic block copolymers spontaneously assemble via electrostatic interactions with DNA molecules in aqueous solution giving rise to micellar structures that protect the DNA from enzymatic degradation both in vitro and in vivo. In addition, we have previously shown that they are safe, not immunogenic and greatly increased antigen-specific CTL responses following six intramuscular inoculations of a very low dose (1 μg) of the vaccine DNA as compared to naked DNA. Nevertheless, they failed to elicit detectable humoral responses against the antigen. To gain further insight in the potential application of this technology, here we show that a shorter immunization protocol based on two DNA intramuscular inoculations of 1 μg of DNA delivered by these copolymers and a protein boost elicits in mice broad (both humoral and cellular) and long-lasting responses and increases the antigen-specific Th1-type T cell responses and CTLs as compared to priming with naked DNA. These results indicate that cationic block copolymers represent a promising adjuvant and delivery technology for DNA vaccination strategies aimed at combating intracellular pathogens.

Complexes of Cationic Block Copolymer Micelles with DNA: Histone/DNA Complex Mimetics

The formation of complexes between cationic polymeric micelles of PS-b-PQ2VP amphiphilic block copolymers and DNA molecules in aqueous solutions is investigated at pH = 7. The physicochemical characteristics of the "polyplexes" at different DNA/polymer ratios were characterized in terms of mass, size and charge using static, dynamic and electrophoretic light scattering and AFM. The complexes are spherical and assume their maximum size and mass around the charge stoichiometric ratio. After addition of increased amounts of salt in the solutions, partial dissociation of the systems was observed. The present systems can be considered as mimetics of histone/DNA complexes formed under physiological conditions in living cells.

Long-term preservation of DNA in aqueous solutions in the presence of the chemical analogues of microbial autoregulators

The fact of long-term preservation of the physicochemical properties of DNA molecules in aqueous solutions in complexes with methylresorcinol, hexylresorcinol, and tyrosol, the chemical analogues of microbial autoregulators (d1 factors) from the group of alkylhydroxybenzenes (AOB), was established. Compared to the control variants of storage of aqueous DNA solutions, the AOB influence consisted in the sum of correlating effects: the prevention of DNA degradation (according to spectrophotometric parameters) and the preservation of its viscous characteristics and electrophoretic mobility. The initial DNA properties were preserved to the greatest degree in the presence of hexylresorcinol, the compound with the longest alkyl radical. Possible mechanisms of the protective action of alkylhydroxybenzenes in relation to DNA are discussed, namely, the prevention of its hydrolysis due to isolation from the aqueous environment and maintaining DNA stability in the dormant forms of microorganisms.

Molecular dynamics study of single-stranded DNA in aqueous solution confined in a nanopore

Molecular dynamics simulations have been carried out to study the molecular behaviours of single-strand DNA molecules in aqueous solution in a cylindrical pore of 2–3 nm diameter. We examined the solvent and the ion distribution in the nanopore. We studied the conformational properties of the single-strand DNA and the dependence of the conformation on the diameter of the nanopore and on the length of the single-stranded DNA, and the interaction between the single-stranded DNA and the ions. We found that the ions in the nanopore show a preference to the centre of the pore. In addition, the sodium ions exhibit an oscillatory density distribution in the radial direction correlated with the density distribution of the water molecules. The end-to-end distance, and the orientation of the end-to-end vector of the single-stranded DNA are strongly affected by the confinement. The counter-ions adsorb strongly to the charged sites on the single-stranded DNA, most dominantly the backbone phosphate groups. There exists, however, substantial freedom for the counter-ion to desorb from the DNA.

Ion-beam sculpting at nanometre length scales.

Manipulating matter at the nanometre scale is important for many electronic, chemical and biological advances, but present solid-state fabrication methods do not reproducibly achieve dimensional control at the nanometre scale. Here we report a means of fashioning matter at these dimensions that uses low-energy ion beams and reveals surprising atomic transport phenomena that occur in a variety of materials and geometries. The method is implemented in a feedback-controlled sputtering system that provides fine control over ion beam exposure and sample temperature. We call the method "ion-beam sculpting", and apply it to the problem of fabricating a molecular-scale hole, or nanopore, in a thin insulating solid-state membrane. Such pores can serve to localize molecular-scale electrical junctions and switches and function as masks to create other small-scale structures. Nanopores also function as membrane channels in all living systems, where they serve as extremely sensitive electro-mechanical devices that regulate electric potential, ionic flow, and molecular transport across cellular membranes. We show that ion-beam sculpting can be used to fashion an analogous solid-state device: a robust electronic detector consisting of a single nanopore in a Si3N4 membrane, capable of registering single DNA molecules in aqueous solution.

A comparative pulse radiolysis study on DNA's of various molecular masses under anoxic and salt-free conditions, based on conductivity measurements

OH radical attack on various molecular mass DNA molecules in aqueous solutions in O2-free, N2O-saturated solutions at room temperature and with lower doses have been studied. The two phenomena, resulting in a negative and a positive conductivity build-up under a short electron pulse (0.4–1 μs), are discussed in the light of their dependence on the pH, dose rate, concentration and temperature. The positive conductivity build-up observed at lower concentrations of DNA and higher doses is attributed to a degradation process resulting in ssb formation and liberation of counter ions, whereas the negative conductivity build-up at the higher concentrations of DNA and relatively lower doses is attributed to an intermolecular reaction, resulting in cross-links and condensation of the counter ions. The dependence of the applied potential (20–100 V) on the rate constant and conductivity build-up for both processes are shown.

A simple model of DNA superhelices in solution.

Closed circular DNA molecules in aqueous solution take the form of interwound superhelices over a wide range of superhelix densities. We describe a very simple model of such a superhelix in which twisting and bending forces are in balance, subject both to topological constraints and to a limitation on the distance of closet approach of the interwound duplexes of the superhelix. The model is consistent with some of the observed physical properties of closed circular DNA, and suggests that there may be severe limits to the range of allowable geometries for the superhelix structure of minimum energy.