Measuring oxidative DNA damage and DNA repair using the yeast comet assay

Chromosomal DNA damage can be a result of several processes and agents of endogenous or exogenous origin. These cause strand breaks or oxidized bases that lead to strand breaks, which relax the normally supercoiled genomic DNA and increase its electrophoretic mobility. The extent of DNA damage can be assessed by single cell gel electrophoresis, where the chromosomal DNA migration distance correlates with the extent of DNA damage. This technique has been used for a variety of applications with several organisms, but only a few studies have been reported for Saccharomyces cerevisiae. A possible reason for this absence is that low cellular DNA content could hamper visualization. Here we report an optimization of the comet assay protocol for yeast cells that is robust and sensitive enough to reproducibly detect background DNA damage and oxidative damage caused by hydrogen peroxide. DNA repair was observed and quantified as diminishing comet tail length with time after oxidative stress removal in a process well described by first‐order kinetics with a tail length half‐life of 11 min at 37 °C. This is, to our knowledge, the first quantitative measurement of DNA repair kinetics in S. cerevisiae by this method. We also show that diet antioxidants protect from DNA damage, as shown by a three‐fold decrease in comet tail length. The possibility of assessment of DNA damage and repair in individual cells applied to the model organism S. cerevisiae creates new perspectives for studying genotoxicity and DNA repair. Copyright © 2010 John Wiley & Sons, Ltd.     

Batch Tests for Optimisation of Solvent Composition and Process Flexibility of the CHALMEX FS-13 Process

ABSTRACT

Studies have been performed with the purpose of determining the optimal solvent composition of a Chalmers grouped actinide extraction (CHALMEX) solvent for the selective co-extraction of transuranic elements in a novel Grouped ActiNide EXtraction (GANEX) process. The solvent is composed of 6,6’-bis(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-benzo-[1,2,4]-triazin-3-yl)-[2,2’]-bipyridine (CyMe₄-BTBP) and tri-n-butyl phosphate (TBP) in phenyl trifluoromethyl sulfone (FS-13). The performance of the system has been shown to significantly depend on the ratios of the two extracting agents and the diluent to one another. Furthermore, the performance of the determined optimal solvent (10 mM CyMe₄-BTBP in 30% v/v TBP and 70% v/v FS-13) on various simulated PUREX raffinate solutions was tested. It was found that the solvent extracts all transuranic elements with high efficiency and good selectivity with regard to most other elements (fission products/activation products) present in the simulated PUREX raffinate solutions. Moreover, the solvent was found to extract a significant amount of acid. Palladium, silver, and cadmium were co-extracted along with the TRU-radionuclides, which has also been observed in other similar CHALMEX systems. The extraction of plutonium and uranium was preserved for all tested simulated PUREX raffinate solutions compared to experiments using trace amounts.     

Linking entrepreneurial orientation to project success

Entrepreneurial orientation (EO) is a strategic posture of an organization, and it is related to basic policies and practices for the development of entrepreneurial actions looking for creating competitive advantages. This study develops and tests a model of the relationship between entrepreneurial orientation and project success in Brazilian context. As quantitative research, a survey was used to collect data. A sample of 100 valid answers from project practitioners was treated through the structural equation modeling method. As research implications, the main result points out the positive correlation between the entrepreneurial orientation and the project success, contributing to the development of this research subject and helping to minimize the gap in the literature that addresses the relationship between project success and EO. In practical terms, understanding that innovativeness, risk taking, proactiveness, autonomy and competitive aggressiveness (the dimensions of the EO) can contribute to project success and can also indirectly impact on organizational performance, could help organizations get competitive advantage when developing correlate factors. Finally, the results suggest that practices of project management can be aligned to the firm's entrepreneurial orientation to enable firms to attain better results in their projects and generate a competitive advantage. On other hand, given the proportion of the impact of EO on project success (20.3%) identified in this study, it is critical that project management professionals expand their horizon to recognize other factors that affect project success.     

Compressive auto-indexing in femtosecond nanocrystallography

Ultrafast nanocrystallography has the potential to revolutionize biology by enabling structural elucidation of proteins for which it is possible to grow crystals with 10 or fewer unit cells on the side. The success of nanocrystallography depends on robust orientation-determination procedures that allow us to average diffraction data from multiple nanocrystals to produce a three-dimensional (3D) diffraction data volume with a high signal-to-noise ratio. Such a 3D diffraction volume can then be phased using standard crystallographic techniques. “Indexing” algorithms used in crystallography enable orientation determination of diffraction data from a single crystal when a relatively large number of reflections are recorded. Here we show that it is possible to obtain the exact lattice geometry from a smaller number of measurements than standard approaches using a basis pursuit solver.     

On the production of novel zirconia-reinforced bioactive glass porous structures for bone repair

Journal of Materials Science - The objective of this study was to develop a replica method for producing zirconia-reinforced bioactive glass (ZRBG) porous structures for bone...     

Structural and Catalytic Studies of a Trimethyltin Vanadate Coordination Polymer

The organotin vanadate [Me₃SnVO₃] (1) has been prepared and characterised in the solid state by powder X-ray diffraction (XRD), thermogravimetric analysis, multinuclear magic-angle spinning (MAS) NMR, IR and Raman spectroscopy. The phase purity and structure of microcrystalline 1 were confirmed by carrying out a full Rietveld structural refinement at ambient temperature and from conventional powder XRD. ⁵¹V and ¹¹⁹Sn MAS NMR data for compound 1 were in agreement with the predicted structure, showing two equally-abundant, nonequivalent Me₃Sn groups and two equally-abundant, nonequivalent vanadium atoms. The compound was applied as a catalyst for the liquid-phase epoxidation of olefins at 55 °C using tert-butyl hydroperoxide (tBuOOH) as the oxidant. The reaction rate for the different substrates followed the order cis-cyclooctene > (R)-(+)-limonene ≅ trans-2-octene > cyclododecene > styrene > 1-octene; the corresponding epoxides were the only observed products. Leaching tests indicated that the catalytic epoxidation of cyclooctene was mainly heterogeneous in nature. This paper is dedicated to Professor Ian Manners and his scientific accomplishments.     

Thermal-responsive hydrogels for sublingual administration of Ondansetron™

Thermo-sensitive hydrogels were prepared by grafting N-isopropylacrylamide (NIPAAm) onto chitosan and dextran. The grafting reactions were performed during variable periods of time and different films were obtained. To prepare a drug-delivery system for sublingual delivery of Ondansetron?, this drug was entrapped in the films. Elemental analysis results showed that the amount of poly(NIPAAm) in the final films increased with reaction time, influencing their lower critical solution temperature and swelling behavior. Also, drug release profile depended on grafting extension as well as on the temperature, as the release of Ondansetron? presented a more sustained-like profile at physiologic temperature.     

Online automatic identification of the modal parameters of a long span arch bridge

The “Infante D. Henrique” bridge is a concrete arch bridge, with a span of 280 m that crosses the Douro River, linking the cities of Porto and Gaia located in the North of Portugal. This structure is being monitored by a recently installed dynamic monitoring system that comprises 12 acceleration channels. This paper describes the bridge structure, its dynamic parameters identified with a previously developed ambient vibration test, the installed monitoring equipment and the software that continuously processes the data received from the bridge through an Internet connection. Special emphasis is given to the algorithms that have been developed and implemented to perform the online automatic identification of the structure modal parameters from its measured responses during normal operation. The proposed methodology uses the covariance driven stochastic subspace identification method (SSI-COV), which is then complemented by a new algorithm developed for the automatic analysis of stabilization diagrams. This new tool, based on a hierarchical clustering algorithm, proved to be very efficient on the identification of the bridge first 12 modes. The results achieved during 2 months of observation, which involved the analysis of more than 2500 datasets, are presented in detail. It is demonstrated that with the combination of high-quality equipment and powerful identification algorithms, it is possible to estimate, in an automatic manner, accurate modal parameters for several modes. These can then be used as inputs for damage detection algorithms.     

Multi-photon imaging of amine-functionalized silica nanoparticles

A convenient and simple strategy for preparing water soluble, photoluminescent functionalized silica nanoparticles (M-dots) in the absence of fluorophores or metal doping is demonstrated. These M-dots can be used for bioimaging using one and two-photon microscopy. Because of their high photostability, low toxicity and high biocompatibility compared with Lumidot? CdSe/ZnS quantum dots, functionalized silica particles are superior alternatives for current bioimaging platforms. Moreover, the presence of a free amine group at the surface of the M-dots allows biomolecule conjugation (e.g. with antibodies, proteins) in a single step for converting these photoluminescent SiO(2) nanoparticles into multifunctional efficient vehicles for theragnostics.