Liming Ying

Frequency and Voltage Dependence of the Dielectrophoretic Trapping of Short Lengths of DNA and dCTP in a Nanopipette

The study of the properties of DNA under high electric fields is of both fundamental and practical interest. We have exploited the high electric fields produced locally in the tip of a nanopipette to probe the motion of double- and single-stranded 40-mer DNA, a 1-kb single-stranded DNA, and a single-nucleotide triphosphate (dCTP) just inside and outside the pipette tip at different frequencies and amplitudes of applied voltages. We used dual laser excitation and dual color detection to simultaneously follow two fluorophore-labeled DNA sequences with millisecond time resolution, significantly faster than studies to date. A strong trapping effect was observed during the negative half cycle for all DNA samples and also the dCTP. This effect was maximum below 1 Hz and decreased with higher frequency. We assign this trapping to strong dielectrophoresis due to the high electric field and electric field gradient in the pipette tip. Dielectrophoresis in electrodeless tapered nanostructures has potential applications for controlled mixing and manipulation of short lengths of DNA and other biomolecules, opening new possibilities in miniaturized biological analysis.

Non-Arrhenius kinetics for the loop closure of a DNA hairpin

Intramolecular chain diffusion is an elementary process in the conformational fluctuations of the DNA hairpin-loop. We have studied the temperature and viscosity dependence of a model DNA hairpin-loop by FRET (fluorescence resonance energy transfer) fluctuation spectroscopy (FRETfs). Apparent thermodynamic parameters were obtained by analyzing the correlation amplitude through a two-state model and are consistent with steady-state fluorescence measurements. The kinetics of closing the loop show non-Arrhenius behavior, in agreement with theoretical prediction and other experimental measurements on peptide folding. The fluctuation rates show a fractional power dependence (β = 0.83) on the solution viscosity. A much slower intrachain diffusion coefficient in comparison to that of polypeptides was derived based on the first passage time theory of SSS [Szabo, A., Schulten, K. & Schulten, Z. (1980) J. Chem. Phys. 72, 4350–4357], suggesting that intrachain interactions, especially stacking interaction in the loop, might increase the roughness of the free energy surface of the DNA hairpin-loop.

The scanned nanopipette: a new tool for high resolution bioimaging and controlled deposition of biomolecules

The boundary between the physical and biological sciences has been eroded in recent years with new physical methods applied to biology and biological molecules being used for new physical purposes. We have pioneered the application of a form of scanning probe microscopy based on a scanned nanopipette, originally developed by Hansma and co-workers, for reliable non-contact imaging over the surface of a live cell. We have found that the nanopipette can also be used for controlled local voltage-driven application of reagents or biomolecules and this can be used for controlled deposition and the local delivery of probes for mapping of specific species. In this article we review this progress, focussing on the physical principles and new phenomena that we have observed, and then outline the future applications that are now possible.

Room-temperature fluorescence, phosphorescence and crystal structures of 4-acyl pyrazolone lanthanide complexes: Ln(L)3·2H2O

Abstract A series of ternary mixed ligand 4-acyl pyrazolone lanthanide complexes: Ln(L)3·2H2O [where Ln = Tb3+ or Gd3+, L = 1-phenyl-3-methyl-4-acetyl-pyrazolone-5 (PMAP), 1-phenyl-3-methyl-4-propionyl-5-pyrazolone (PMPP), 1-phenyl-3-methyl-4-isobutyryl-5-pyrazolone (PMIP), 1-phenyl-3-methyl-4-neovaleryl-pyrazolone-5 (PMNP) and 1-phenyl-3-methyl-4-benzoyl-pyrazolone-5 (PMBP)] were synthesized and characterized by FT-IR spectra, UV-vis spectra and DTA-TG analysis. Room-temperature phosphorescence was observed from the Gd3+ complexes by excitation of the sample with the fourth harmonic frequency of a Nd: YAG laser beam (γ = 266 nm) and the triplet energies of the pyrazolone ligands were evaluated. Both the fluorescence intensity and fluorescence lifetime of the Tb3+ complexes depend on the structure of the ligands and explanations are presented. The crystal structure [Tb(PMPP)3·2H2O]·EtOH was determined by X-ray diffraction. The structure was refined to R = 0.064 (Rw = 0.073). The complex is mononuclear and the central terbium ion is coordinated by eight oxygen atoms to form a square-antiprism coordination polyhedron, six of which are from the three bidentate pyrazolone ligands and the other two are from the two coordination water molecules.

Fluorescence resonance energy transfer between a quantum dot donor and a dye acceptor attached to DNA

We show that direct coupling of a dye-labelled DNA (acceptor) to a quantum dot (QD) donor significantly reduces the donor-acceptor distance and improves the FRET efficiency: a highly efficient FRET (approximately 88%) at a low acceptor-to-donor ratio of 2 has been achieved at the single-molecule level.

Two-state model of conformational fluctuation in a DNA hairpin-loop

Stretched exponential kinetics have been observed in the conformational fluctuation of a DNA hairpin-loop under equilibrium conditions. In this paper, we employ a simple multiple-pathway two-state jump model to calculate singlemolecule proximity ratio distributions. The simulation can reasonably reproduce the experimental single-molecule data of the conformational fluctuations in water, indicating that static disorder is dominant. In contrast, there exists significant discrepancy between the two-state simulation and experiment in buAer (2.5 mM Tris-HCl, 250 lM EDTA, 100 mM NaCl), suggesting that both static and dynamic disorder may contribute to the non-exponential kinetics. ” 2001 Elsevier Science B.V. All rights reserved.

Photodissociation of methylazide: Observation of triplet methylnitrene radical

An investigation of the photodissociation of methylazide at wavelengths from 292 nm to 325 nm is presented. Emission spectra and lifetime analysis show the existence of the triplet CH3N radical. A simple kinetic model is proposed to explain the observed fluorescence real time profile, which reflects the vibrational relaxation of hot radical at high pressures. The photodissociation dynamics of methylazide seems complex. The predominant channel is likely to produce CH2NH via concerted 1,2‐hydrogen shift and N2 extrusion process. The triplet CH3N comes from a minor spin‐forbidden channel which involves possibly strong interaction between the low‐lying excited singlet and triplet states of methylazide.

Fluorescence studies of single biomolecules

Single-molecule fluorescence has the capability to detect properties buried in ensemble measurements and, hence, provides new insights about biological processes. Ratiometric methods are normally used to reduce the effects of excitation beam inhomogeneity. Fluorescence resonance energy transfer is widely used but there are problems in inserting the fluorophores in the correct position on the biomolecule, particularly if the structure is not known. We have recently developed two-colour coincidence single-molecule fluorescence that addresses this problem. This method can be used to determine quantitatively the multimerization states of biomolecules, in solution without separation. The future prospects of single-molecule fluorescence as applied to biological molecules are discussed.

Fabrication and their photoelectric conversion properties of two kinds of self-assembled monolayers and Langmuir–Blodgett film of mono-substituted C60-malonic acid

Two kinds of self-assembled monolayer (SAM) and a Langmuir–Blodgett film of fullerene derivative C61(COOH)2 that have the photoelectric conversion property were first reported here. The SAMs were either fabricated directly on hydrophilic indium–tin-oxide (ITO), quartz, and mica by esterification reaction or on the aminopropylsilanated substrates by amide bonds. The SAMs were characterized by contact angle, AFM, UV spectrum and cyclic voltammetry. The film thickness calculated by molecular mechanic with the MM2 method in Molgen 4.0 coincided with that obtained by AFM measurement, indicating the formation of monolayer. All of three systems can generate stable cathodic photocurrent. The separated charge loss process in our systems is monomolecule recombination.

Dissociation dynamics of methylazide on the first excited singlet surface

The photolysis of methylazide around 313 nm is studied experimentally by laser Raman spectroscopy. The emission spectrum with a resolution of 50 cm−1 is reported. Based on our experiments, the photodissociation of methylazide on its first excited singlet surface is proposed to be a concerted N2 elimination and 1,2-H shift process.