Response of short tandem repeat systems to temperature and sizing methods

In capillary gel electrophoresis (CE), changing run conditions such as temperature can result in minor variations in the size determination of an allele. These effects are caused by secondary structure differences that can occur between the amplified sample and the internal standard. The type of method chosen to generate the sizing curve in STR analysis can influence the relationship between estimated allele size and temperature. To better understand the effects of temperature and sizing method on the reproducibility of DNA migration, two fluorescently labeled allelic ladders, CTTv and Y-PLEX 6 were analyzed using the ABI Prism 310 Genetic Analyzer. The default method on the Genetic Analyzer utilizes an electrophoretic temperature of 60 degrees C and a Local Southern method to generate a sizing curve from the fragment migration times of the internal lane standard. In this work, electrophoresis was conducted at 35-70 degrees C using the commercially available POP 4 buffer at pH 8 and two sizing methods, Global Southern and Local Southern, were compared. The slopes of the regression line between estimated allele size and temperature, using either sizing method, were measured in order to demonstrate the temperature sensitivity of migration time and the importance of the operator-chosen method. Our results indicate that the Global Southern method is a better choice in situations where temperature fluctuations can occur. In addition, the temperature dependence of the DNA size estimates using the POP 4 system were compared to results obtained using an experimental buffer consisting of 3% hydroxyethylcellulose at pH 11. These results demonstrate that secondary structure effects are minimized at an elevated pH, increasing the precision of size estimates obtained.     

Canadian students join struggle for access to treatment

Canadian students have joined the struggle for global access to treatment. This article describes initiatives at McGill University and the University of Toronto.     

Evaluation of a multicapillary electrophoresis instrument for mitochondrial DNA typing

Laser-induced detection of fluorescent labeled PCR products and multi-wavelength detection (i.e., multicolor analysis) enables rapid generation of mtDNA sequencing profiles. Traditionally, polyacrylamide slab gels have been used as the electrophoretic medium for mtDNA sequencing in forensic analyses. Replacement of slab gel electrophoresis with capillary electrophoresis (CE) can facilitate automation of the analytical process. Automation and high throughput can be further enhanced by using multicapillary electrophoretic systems. The use of the ABI Prism 3100 Genetic Analyzer (ABI 3100, Applied Biosystems, Foster City, CA) as well as the ABI Prism 310 Genetic Analyzer (ABI 310, Applied Biosystems, Foster City, CA) were evaluated for mtDNA sequencing capabilities and compared with sequencing results obtained on the platform currently in use in the FBI Laboratory (the ABI Prism 377 DNA Sequencer, ABI 377, Applied Biosystems, Foster City, CA). Various studies were performed to assess the utility of the ABI 3100, as well as the ABI 310 for mtDNA sequencing. The tests included: comparisons of results obtained among the ABI 3100, the ABI 310 and the ABI 377 instruments; comparisons of results obtained within and between capillary arrays; evaluation of capillary length; evaluation of sample injection time; evaluation of the resolution of mixtures/heteroplasmic samples; and evaluation of the sensitivity of detection of a minor component with reduced template on the ABI 3100. In addition, other studies were performed to improve sample preparation; these included: comparison of template suppression reagent (TSR, Applied Biosystems, Foster City, CA) versus formamide; the use of Performa DTR Gel Filtration Cartridges (Edge BioSystems Inc., Gaithersburg, MD) versus Centri-Sep Spin Columns (Princeton Separations, Adelphia, NJ) for product purification after cycle sequencing; and sample stability after denaturation. The data support that valid and reliable results can be obtained using either capillary electrophoresis instrument, and the quality of sequencing results are comparable to or better than those obtained from the ABI 377 instrument.     

Evaluation of extraction techniques for the forensic analysis of human scalp hair using gas chromatography/mass spectrometry (GC/MS)

Preliminary research using on-line supercritical fluid extraction/gas chromatography-mass spectrometry (SFE/GC-MS) has shown that the natural and artificial surface components of human scalp hair are reproducible and differentiable. Therefore, these components may be useful for individualization or determining demographic characteristics or both. However, it is not known how the efficiency and selectivity of on-line SFE/GC-MS compares to other extraction methods. In this study, ultrasound, Soxhlet, and pressurized-fluid extraction were used to extract 1 mg to 1.3 g portions of a composite hair sample taken from an Asian male between the ages of 10 and 18. Percent extractables ranged from 0.9% to 5.6%, depending on the solvent used, and tended to increase with solvent polarity. Chemical analysis using GC/MS showed that the extracts contained large proportions of free fatty acids, squalene, cholesterol, and various wax esters. Finally, comparisons to SFE/GC-MS showed that this method possesses adequate efficiency, no observable differences in selectivity, and greater potential for miniaturization.     

How Has the Law Gone Mad? Reflections on Semiotics and Postomodernism

International Journal for the Semiotics of Law - Revue internationale de Sémiotique juridique -     

Designing Health Service Organization in the UK, 1968 to 1998: from Blueprint to Bright Idea and ‘Manipulated Emergence’

Four sets of reforms of the National Health Service are employed to illustrate the changing character of policy making in this sector over a thirty year period, from the production of a carefully developed technocratic blueprint for its organization to the promulgation of a series of bright ideas accompanied by incentives for local actors to develop them into concrete organizational arrangements consonant with these ideas. We term this latter approach ‘manipulated emergence’ and relate it to the literatures of organizational culture and of post-Fordism. The approach adopted by the 1997 Labour government is largely, though not wholly consistent with this, and it remains to be seen whether the high-water mark of manipulated emergence has passed.     

After Chechnya: At Risk in Dagestan

Population displacements in the aftermath of the Chechen conflict may undermine a complex political balance existing among the extraordinarily numerous ethnic groups in the neighbouring Republic of Dagestan. The destabilisation of Dagestan threatens to bring ethnic conflict to virtually the only state in the Caucasus to have avoided it thus far. This article considers the basis of ethnic accommodation in Dagestan, the policies and migratory patterns that have placed it at risk, and the regional context in which these have occurred.