Conductivity control of as-grown branched indium tin oxide nanowire networks.

Branched indium tin oxide (ITO) nanowire networks are promising candidates for transparent conductive oxide applications, such as optoelectronic electrodes, due to their high porosity. However, these branched networks also present new challenges in assessing conductivity. Conventional four-point probe techniques cannot separate the effect of porosity on the long-range conductivity from the intrinsic material conductivity. Here we compare the average nanoscale conductivity within the film measured by terahertz time-domain spectroscopy (THz-TDS) to the film conductivity measured by four-point probe in a branched ITO nanowire network. Both techniques report conductivity increases with deposition flux rate from 0.5 to 3.0 nm s(-1), achieving a maximum of ~ 10 (Ω cm)(-1). Modeling the THz-TDS conductivity data using the Drude-Smith model allows us to distinguish between conductivity increases resulting from morphological changes and those resulting from the intrinsic properties of the ITO. In particular, the intrinsic material conductivity within the nanowires can be extracted, and is found to reach a maximum of ~ 3000 (Ω cm)(-1), comparable to bulk ITO. To determine the mechanism responsible for increasing conductivity with flux rate, we characterize dopant concentration and morphological changes (i.e., to branching behavior, nanowire diameter and nucleation layers). We propose that changes in the electron density, primarily due to changes in O-vacancy concentration at different flux rates, are responsible for the observed conductivity increase. This understanding will assist balancing structural and conductivity requirements in applications of transparent conductive oxide networks.

Kirkpatrick-Baez microscope for hard X-ray imaging of fast ignition experiments.

A Kirkpatrick-Baez X-ray microscope has been developed for use on the Titan laser facility at the Lawrence Livermore National Laboratory in Fast Ignition experiments. It was developed as a broadband alternative to narrow band Bragg crystal imagers for imaging Kα emission from tracer layers. A re-entrant design is employed which allows for alignment from outside the chamber. The mirrors are coated with Pt and operate at a grazing incident angle of 0.5° providing higher resolution than an equal brightness pinhole and sufficient bandwidth to image thermally shifted characteristic Kα emission from heated Cu tracer layers in Fast Ignition experiments. The superpolished substrates (<1 Å rms roughness) had a final visible wavelength roughness of 1.7 Å after coating, and exhibited a reflectivity corresponding to an X-ray wavelength roughness of 7 ± 1 Å. A unique feature of this design is that during experiments, the unfiltered direct signal along with the one-dimensional reflections are retained on the detector in order to enable a live indication of alignment and incident angle. The broad spectral window from 4 to 9 keV enables simultaneous observation of emission from several spectral regions of interest, which has been demonstrated to be particularly useful for cone-wire targets. An experimentally measured resolution of 15 µm has been obtained at the center of the field of view.

Indium tin oxide nanowhisker morphology control by vapour-liquid-solid glancing angle deposition.

A new growth technique for indium tin oxide nanowhiskers with increased control over feature size and spacing is reported. The technique is based on a unique combination of self-catalysed vapour-liquid-solid (VLS) growth and glancing angle deposition (GLAD). This VLS-GLAD technique provides enhanced control over nanowhisker morphology as the effect of typical VLS growth parameters (e.g. flux rate, temperature) is amplified at large deposition angles characteristic of GLAD. Spatial modulation of the collimated growth flux controls trunk width, number and orientation of branches, and overall nanowhisker density. Here we report the influence of growth conditions (including deposition angle, flux rate, nominal pitch and substrate temperature) on nanowhisker morphology, with specific focus on the effect of large deposition angles. Sheet resistance and transmission of the films were measured to characterize their performance as transparent conductive oxides. Hybrid nanostructured films grown in this study include high surface area nanowhiskers protruding from a conductive film, ideal for transparent conductive electrode applications.

In vitro selection of a novel nuclease-resistant RNA phosphodiesterase.

BACKGROUND: Ribonucleotide-based enzymes (ribozymes) that cleave pathological RNAs are being developed as therapeutic agents. Chemical modification of the hammerhead ribozyme has produced nuclease-resistant catalysts that cleave targeted mRNAs in cell culture and exhibit antitumor activity in animals. Unfortunately, stabilizing modifications usually reduce the catalytic rate in vitro. An alternative to rationally designed chemical modifications of existing ribozymes is to identify novel motifs through in vitro selection of nuclease-stable sequence space. This approach is desirable because the catalysts can be optimized to function under simulated physiological conditions. RESULTS: Utilizing in vitro selection, we have identified a nuclease-stable phosphodiesterase that demonstrated optimal activity at simulated physiological conditions. The initial library of 10(14) unique molecules contained 40 randomized nucleotides with all pyrimidines in a nuclease-stabilized 2'-deoxy-2'-amino format. The selection required trans-cleaving activity and base-pairing specificity towards a resin-bound RNA substrate. Initial selective pressure was permissive, with a 30 min reaction time and 25 mM Mg(2+). Stringency of selection pressure was gradually increased until final conditions of 1 mM Mg(2+) and less than 1 min reaction times were achieved. The resulting 61-mer catalyst required the 2'-amino substitutions at selected pyrimidine positions and was stable in human serum (half-life of 16 h). CONCLUSIONS: We demonstrated that it is possible to identify completely novel, nuclease-resistant ribozymes capable of trans-cleaving target RNAs at physiologically relevant Mg(2+) concentrations. The new ribozyme motif has minimal substrate requirements, allowing for a wide range of potential RNA targets.

Determinants of ribose specificity in RNA polymerization: effects of Mn2+ and deoxynucleoside monophosphate incorporation into transcripts.

The catalytic specificity of T7 RNA polymerase (RNAP) for ribonucleoside triphosphates vs deoxynucleoside triphosphates {(kcat/Km)rNTP/(kcat/Km)dNTP} during transcript elongation is approximately 80. Mutation of tyrosine 639 to phenylalanine reduces specificity by a factor of approximately 20 and largely eliminates the Km difference between rNTPs and dNTPs. The remaining specificity factor of approximately 4 is kcat-mediated and is nearly eliminated if Mn2+ is substituted for Mg2+ in the reaction. Mn2+ substitution does not significantly affect the Km difference between rNTPs and dNTPs. Mn2+ substitution also enhances the activity of poorly active mutant enzymes carrying nonconservative substitutions in the active site, and its effects are generally consistent with the Mn2+-catalyzed reaction being less restrictive in its requirements for alignment of the reactive groups. In addition to discrimination occurring at the level of nucleoside monophosphate (NMP) incorporation, it is also found that transcripts containing deoxynucleoside monophosphates (dNMPs) are more poorly extended than transcripts of canonical structure, though a severe barrier to transcript extension is seen only when the 3' region of the transcript is heavily substituted with dNMPs. The barrier to extension of transcripts heavily substituted with dNMPs is reduced for sequences known to be amenable to forming A-like helices and is larger for sequences that resist transformation from B-form DNA.DNA structures. The barrier to extension of dNMP-substituted transcripts is also reduced by solution conditions known to destabilize B-form DNA and to stabilize A-form structures. These observations imply a requirement for a non-B-form, possibly A-like, conformation in the transcript.template hybrid that is disrupted when the transcript is of predominantly deoxyribose structure.

Inhibition of vascular smooth muscle cell proliferation by ribozymes that cleave c-myb mRNA.

Proliferation of injured smooth muscle cells contributes to the reocclusion or restenosis of coronary arteries that often occurs following angioplasty procedures. We have identified and optimized nuclease-resistant ribozymes that efficiently cleave c-myb RNA. Three ribozymes targeting different sites in the c-myb mRNA were synthesized chemically and delivered to rat aortic smooth muscle cells with cationic lipids; all three inhibited serum-stimulated cell proliferation significantly. RNA molecules with two base substitutions in the catalytic core that render the ribozyme catalytically inactive had little effect on smooth muscle cell proliferation. Ribozymes with scrambled binding arm sequences also failed to affect cell cycle progression of vascular smooth muscle cells. Furthermore, inhibition of rat smooth muscle cell proliferation correlated with a reduction in intact c-myb mRNA. Efficacy of the chemically-modified ribozyme was compared directly to phosphorothioate antisense oligodeoxynucleotides targeting the same site in the c-myb RNA; the ribozyme had superior efficacy and showed greater specificity than the antisense molecules. Exogenously delivered ribozymes also inhibited porcine and human smooth muscle cell proliferation effectively. Ribozymes targeting c-myb or other regulators of smooth muscle cell proliferation may represent novel therapeutics for the treatment of restenosis after coronary angioplasty.

Directed evolution of an RNA enzyme.

An in vitro evolution procedure was used to obtain RNA enzymes with a particular catalytic function. A population of 10(13) variants of the Tetrahymena ribozyme, a group I ribozyme that catalyzes sequence-specific cleavage of RNA via a phosphoester transfer mechanism, was generated. This enzyme has a limited ability to cleave DNA under conditions of high temperature or high MgCl2 concentration, or both. A selection constraint was imposed on the population of ribozyme variants such that only those individuals that carried out DNA cleavage under physiologic conditions were amplified to produce "progeny" ribozymes. Mutations were introduced during amplification to maintain heterogeneity in the population. This process was repeated for ten successive generations, resulting in enhanced (100 times) DNA cleavage activity.

Minimum secondary structure requirements for catalytic activity of a self-splicing group I intron.

We have completed a comprehensive deletion analysis of the Tetrahymena ribozyme in order to define the minimum secondary structure requirements for phosphoester transfer activity of a self-splicing group I intron. A total of 299 nucleotides were removed in a piecewise fashion, leaving a catalytic core of 114 nucleotides that form 7 base-paired structural elements. Among the various deletion mutants are a 300-nucleotide single-deletion mutant and a 281-nucleotide double-deletion mutant whose activity exceeds that of the wild type when tested under physiologic conditions. Consideration of those structural elements that are essential for catalytic activity leads to a simplified secondary structure model of the catalytic core of a group I intron.