The Sister-Chromatid Exchange Assay in Human Cells

The semiconservative nature of DNA replication allows the differential labeling of sister chromatids that isthe fundamental requirement to perform the sister-chromatid exchange (SCE) assay. SCE assay is apowerful technique to visually detect the physical exchange of DNA between sister chromatids. SCEscould result as a consequence of DNA damage repair by homologous recombination (HR) during DNAreplication. Here, we provide the detailed protocol to perform the SCE assay in cultured human cells. Cellsare exposed to the thymidine analog 5-bromo-20-deoxyuridine (BrdU) during two cell cycles, resulting inthe two sister chromatids having differential incorporation of the analog. After metaphase spreads prepara-tion and further processing, SCEs are nicely visualized under the microscope

Detection of known haemophilia B mutations and carrier testing by microarray

The molecular basis of haemophilia B is heterogeneous and many mutations of the Factor IX (FIX) gene have been characterised. Using the allele-specific arrayed primer extension (AS-APEX) technology, we have designed a FIX array to simultaneously analyse 69 mutations found in British, Thai and Chinese patients. This technology overcomes the problem of multiple reverse dot-blot analysis and has a 100% accuracy in the detection of both affected subjects and carriers in families with known mutations. In seven unknown mutations from Thailand, the array could detect the specific mutation in five and in the remainders the normal primer at specific spots failed to extend due to a mutation a few nucleotides upstream, thus allowing their identification. Hence this FIX array can detect 53% of the 2891 mutation entries in the FIX database. Each of the microarray slide can be used for three different test samples and would be useful for carrier testing for common mutations and prenatal diagnosis. It is simpler and more cost effective than genome sequencing and would be particularly useful in laboratories with limited technical capabilities. © 2005 Schattauer GmbH, Stuttgart.published_or_final_versio

Irreversible reorganization in a supercooled liquid originates from localised soft modes

The transition of a fluid to a rigid glass upon cooling is a common route of transformation from liquid to solid that embodies the most poorly understood features of both phases1,2,3. From the liquid perspective, the puzzle is to understand stress relaxation in the disordered state. From the perspective of solids, the challenge is to extend our description of structure and its mechanical consequences to materials without long range order. Using computer simulations, we show that the localized low frequency normal modes of a configuration in a supercooled liquid are causally correlated to the irreversible structural reorganization of the particles within that configuration. We also demonstrate that the spatial distribution of these soft local modes can persist in spite of significant particle reorganization. The consequence of these two results is that it is now feasible to construct a theory of relaxation length scales in glass-forming liquids without recourse to dynamics and to explicitly relate molecular properties to their collective relaxation.Comment: Published online: 20 July 2008 | doi:10.1038/nphys1025 Available from http://www.nature.com/nphys/journal/v4/n9/abs/nphys1025.htm

Confronting Standard Models of Proto–Planetary Disks With New Mid–Infrared Sizes from the Keck Interferometer

This is the final version of the article. Available from American Astronomical Society via the DOI in this record.The accepted author manuscript is in ORE at http://hdl.handle.net/10871/21611We present near- and mid-infrared (MIR) interferometric observations made with the Keck Interferometer Nuller and near-contemporaneous spectro-photometry from the infrared telescope facilities (IRTFs) of 11 well-known young stellar objects, several of which were observed for the first time in these spectral and spatial resolution regimes. With au-level spatial resolution, we first establish characteristic sizes of the infrared emission using a simple geometrical model consisting of a hot inner rim and MIR disk emission. We find a high degree of correlation between the stellar luminosity and the MIR disk sizes after using near-infrared data to remove the contribution from the inner rim. We then use a semi-analytical physical model to also find that the very widely used "star + inner dust rim + flared disk" class of models strongly fails to reproduce the spectral energy distribution (SED) and spatially resolved MIR data simultaneously; specifically a more compact source of MIR emission is required than results from the standard flared disk model. We explore the viability of a modification to the model whereby a second dust rim containing smaller dust grains is added, and find that the 2-rim model leads to significantly improved fits in most cases. This complexity is largely missed when carrying out SED modeling alone, although detailed silicate feature fitting by McClure et al. recently came to a similar conclusion. As has been suggested recently by Menu et al., the difficulty in predicting MIR sizes from the SED alone might hint at "transition disk"-like gaps in the inner au; however, the relatively high correlation found in our MIR disk size versus stellar luminosity relation favors layered disk morphologies and points to missing disk model ingredients instead.M.S. was supported by NASA ADAP grant NNX09AC73G. R.W.R. was supported by the IR&D program of The Aerospace Corporation

Characterisation of LV myocardial exercise function by 2-D strain deformation imaging in elite adolescent footballers

This is the final version. Available on open access from Springer via the DOI in this recordPurpose: Few data exist on the descriptions of LV myocardial mechanics and reserve during dynamic exercise of adolescent athletes. The aim of this study was to describe the LV myocardial and cardiopulmonary changes during exercise using 2-D strain deformation imaging. Methods: Elite adolescent male football players (n = 42) completed simultaneous cardiopulmonary exercise testing (CPET) and exercise echocardiography measurement of LV myocardial deformation by 2-D strain imaging. LV longitudinal and circumferential 2-D strain and strain rates were analyzed at each stage during incremental exercise to a work rate of 150 W. Additionally, exercise LV myocardial deformation and its relation to metabolic exercise parameters were evaluated at each exercise stage and in recovery using repeated measures ANOVA, linear regression and paired t tests. Results: LV peak systolic baseline 2-D strain (longitudinal: − 15.4 ± 2.5%, circumferential: − 22.5 ± 3.1%) increased with each exercise stage, but longitudinal strain plateaued at 50 W (mean strain reserve − 7.8 ± 3.0) and did not significantly increase compared to subsequent exercise stages (P > 0.05), whilst circumferential strain (mean strain reserve − 11.6 ± 3.3) significantly increased (P < 0.05) throughout exercise up to 150 W as the dominant mechanism of exercise LV contractility increase. Regression analyses showed LV myocardial strain increased linearly relative to HR, VO2 and O2 pulse (P < 0.05) for circumferential deformation, but showed attenuation for longitudinal deformation. Conclusion: This study describes LV myocardial deformation dynamics by 2-D strain and provides reference values for LV myocardial strain and strain rate during exercise in adolescent footballers. It found important differences between LV longitudinal and circumferential myocardial mechanics during exercise and introduces a methodology that can be used to quantify LV function and cardiac reserve during exercise in adolescent athletes.National Institute for Health Research (NIHR

Walk well:a randomised controlled trial of a walking intervention for adults with intellectual disabilities: study protocol

Background - Walking interventions have been shown to have a positive impact on physical activity (PA) levels, health and wellbeing for adult and older adult populations. There has been very little work carried out to explore the effectiveness of walking interventions for adults with intellectual disabilities. This paper will provide details of the Walk Well intervention, designed for adults with intellectual disabilities, and a randomised controlled trial (RCT) to test its effectiveness. Methods/design - This study will adopt a RCT design, with participants allocated to the walking intervention group or a waiting list control group. The intervention consists of three PA consultations (baseline, six weeks and 12 weeks) and an individualised 12 week walking programme. A range of measures will be completed by participants at baseline, post intervention (three months from baseline) and at follow up (three months post intervention and six months from baseline). All outcome measures will be collected by a researcher who will be blinded to the study groups. The primary outcome will be steps walked per day, measured using accelerometers. Secondary outcome measures will include time spent in PA per day (across various intensity levels), time spent in sedentary behaviour per day, quality of life, self-efficacy and anthropometric measures to monitor weight change. Discussion - Since there are currently no published RCTs of walking interventions for adults with intellectual disabilities, this RCT will examine if a walking intervention can successfully increase PA, health and wellbeing of adults with intellectual disabilities

Fine sediment reduces vertical migrations of Gammarus pulex (Crustacea: Amphipoda) in response to surface water loss

Surface and subsurface sediments in river ecosystems are recognized as refuges that may promote invertebrate survival during disturbances such as floods and streambed drying. Refuge use is spatiotemporally variable, with environmental factors including substrate composition, in particular the proportion of fine sediment (FS), affecting the ability of organisms to move through interstitial spaces. We conducted a laboratory experiment to examine the effects of FS on the movement of Gammarus pulex Linnaeus (Crustacea: Amphipoda) into subsurface sediments in response to surface water loss. We hypothesized that increasing volumes of FS would impede and ultimately prevent individuals from migrating into the sediments. To test this hypothesis, the proportion of FS (1–2 mm diameter) present within an open gravel matrix (4–16 mm diameter) was varied from 10 to 20% by volume in 2.5% increments. Under control conditions (0% FS), 93% of individuals moved into subsurface sediments as the water level was reduced. The proportion of individuals moving into the subsurface decreased to 74% at 10% FS, and at 20% FS no individuals entered the sediments, supporting our hypothesis. These results demonstrate the importance of reducing FS inputs into river ecosystems and restoring FS-clogged riverbeds, to promote refuge use during increasingly common instream disturbances