104 research outputs found
PIMS sequencing extension:a laboratory information management system for DNA sequencing facilities
No full textBackground: Facilities that provide a service for DNA sequencing typically support large numbers of users and experiment types. The cost of services is often reduced by the use of liquid handling robots but the efficiency of such facilities is hampered because the software for such robots does not usually integrate well with the systems that run the sequencing machines. Accordingly, there is a need for software systems capable of integrating different robotic systems and managing sample information for DNA sequencing services. In this paper, we describe an extension to the Protein Information Management System (PIMS) that is designed for DNA sequencing facilities. The new version of PIMS has a user-friendly web interface and integrates all aspects of the sequencing process, including sample submission, handling and tracking, together with capture and management of the data. Results: The PIMS sequencing extension has been in production since July 2009 at the University of Leeds DNA Sequencing Facility. It has completely replaced manual data handling and simplified the tasks of data management and user communication. Samples from 45 groups have been processed with an average throughput of 10000 samples per month. The current version of the PIMS sequencing extension works with Applied Biosystems 3130XL 96-well plate sequencer and MWG 4204 or Aviso Theonyx liquid handling robots, but is readily adaptable for use with other combinations of robots. Conclusions: PIMS has been extended to provide a user-friendly and integrated data management solution for DNA sequencing facilities that is accessed through a normal web browser and allows simultaneous access by multiple users as well as facility managers. The system integrates sequencing and liquid handling robots, manages the data flow, and provides remote access to the sequencing results. The software is freely available, for academic users, from http://www.pims-lims. org/
The diversity of Hippocampus abdominalis in New Zealand
No full textThis study investigates the diversity and population differentiation of the New Zealand Pot-belly seahorse Hippocampus abdominalis through the utilization of morphological and genetic data. Four microsatellite loci - Habd3, Habd6, Habd7 and Habd9 - and three mitochondrial DNA markers - cytochrome b (814 bp), cytochrome oxidase 1 (624 bp) and control region (404 bp) - in conjunction with quantified morphological features revealed a very high diversity but low population differentiation within New Zealand, suggesting very high levels of gene flow. Some sexual dimorphism was detected, in the terms of shorter snout length and trunk length, and a higher incidence of fronds and spotting in males. A sample size of 166 yielded 31-46 microsatellite alleles and no common multilocus genotypes, and 36-40 new sequences were generated for each mitochondrial DNA marker exposing 14-16 haplotypes, with a maximum of 0.7-2.2% sequence divergence. H. abdominalis were found to be widely dispersed mainly in low density populations. As this species is likely to be facing increased threats from exploitation and habitat degradation in the future it is hoped that this information contributes to the knowledge about H. abdominalis so that future conservation management would be easier to implement
Functional characterization of the γ-aminobutyric acid transporter from Mycobacterium smegmatis MC2 155 reveals sodium-driven GABA transport.
No full textCharacterizing the mycobacterial transporters involved in the uptake and/or catabolism of host-derived nutrients required by mycobacteria may identify novel drug targets against tuberculosis. Here, we identify and characterize a member of the amino acid-polyamine- organocation superfamily, a potential γ-aminobutyric acid transport protein, GabP, from Mycobacterium smegmatis The protein was expressed to a level allowing its purification to homogeneity and Size Exclusion Chromatography-Multi Angle Laser Light Scattering analysis of the purified protein showed that it was dimeric. We showed that GabP transported γ-aminobutyric acid in vitro and when over-expressed in E. coli Additionally, transport was greatly reduced in the presence of β-alanine, suggesting that it could be either substrate or inhibitor of GabP. Using GabP reconstituted into proteoliposomes, we demonstrated that γ-aminobutyric acid uptake is driven by the sodium gradient and is stimulated by membrane potential. Molecular docking showed that γ-aminobutyric acid binds MsGabP, another Mycobacterium smegmatis putative GabP and the Mycobacterium tuberculosis homologue in the same manner. This study represents the first expression, purification and characterization of an active γ-aminobutyric acid transport protein from mycobacteria.IMPORTANCE The spread of multidrug resistant tuberculosis increases its global health impact in humans. As there is transmission both to and from animals, the spread of the disease also increases its effects in a broad range of animal species. Identifying new mycobacterial transporters will enhance our understanding of mycobacterial physiology and furthermore provides new drug targets. Our target protein is the gene product of msmeg_6196, annotated as GABA permease, from Mycobacterium smegmatis strain MC2 155. Our current study demonstrates that it is a sodium-dependent GABA transporter that may also transport β-alanine. As GABA may well be an essential nutrient for mycobacterial metabolism inside the host, this could be an attractive target for the development of new drugs against tuberculosis
Free serum cortisol during the postoperative acute phase response determined by equilibrium dialysis liquid chromatography-tandem mass spectrometry
No full textIn severely ill patients low concentrations of the corticosteroid binding globulin are typically found; the aim of this study was to quantify directly free bioactive cortisol concentrations in the sera of postoperative cardiosurgical patients. Serum samples of 12 consecutive patients undergoing aortocoronary bypass surgery taken preoperatively and on the postoperative days 1 to 4 were analyzed. Total serum cortisol was quantified using liquid chromatographytandem mass spectrometry with an online sample extraction system and trideuterated cortisol as the internal standard, and free serum cortisol was measured after overnight equilibrium dialysis. Whereas on the first postoperative day, the median total serum cortisol concentration was approximately twofold increased compared to preoperative samples (preoperatively, 245 nmol/l (interquartile range (IQR) 203293 nmol/l); first postoperative day, 512 nmol/l (IQR 410611 nmol/l)), median dialyzable free cortisol concentration was almost sevenfold increased (preoperatively, 14.2 nmol/l (IQR 10.920.7 nmol/l); first postoperative day, 98.3 nmol/l (IQR 81.3134 nmol/l)). On the fourth postoperative day, median free cortisol was still significantly increased compared to baseline sampling (p < 0.05), whereas median total cortisol was not. A median of 5.7% (IQR 5.47.0%) of total cortisol was found as free cortisol on the preoperative day, 21.2% (IQR 18.9 23.5%) on the first postoperative day and 10.5% (IQR 9.814.0%) on the fourth postoperative day. It is concluded that during the postoperative period the freeto bound ratio of cortisol is highly variable and that during the acute phase response direct quantification of free bioactive cortisol concentrations seems to be biologically more appropriate than the measurement of total cortisol concentrations
The synthesis and properties of some well-defined fluorinated polymers
No full textThis thesis describes studies directed to the ring opening metathesis polymerisation (ROMP) of some fluorinated compounds using a range of well-defined initiators. Chapter 1 reviews some general aspects of olefin metathesis and ring opening metathesis polymerisation of relevance to the work described in this thesis. Topics such as piezo- and pyro-electricity and optical and electrical properties of conjugated polymers are introduced and these receive more detailed attention later in the thesis. Chapter 2 describes the synthesis, characterisation and properties of . poly(bis(trifluororaethyl)norbomadiene) in detail. The use of various initiating systems that have been used previously and the effect they have on the tacticity of the resulting polymer raicrostructure are discussed. The latter part of this chapter reviews some of the current thinking concerning the detailed mechanistic aspects of this polymerisation. Chapter 3 reviews attempts directed to an improvement in tacticity control in the synthesis of poly(bis(trifluoromethyl)norbomadiene). The synthesis and activity of the new well-defined initiators used in these studies are reported. It is shown that varying the nature of the ancillary Ligands surrounding the metal centre can have a dramatic influence on the tacticity of the resulting polymer. Chapter 4 reports studies directed to an examination of the limits of the well controlled synthesis of poly(bis(trifluoromethyl)norbornadiene). The syntheses of high trans and high cis, highly tactic poly(bis(trifluoromethyl)norbornadiene samples using well-defined initiating systems are described. It is shown that by varying the monomerinitiator ratio, samples with a wide range of molecular weights can be achieved and these are reported. Chapter 5 describes experiments concerning the ROMP of fluorinated monomers containing six membered rings. In particular ROMP studies of the monomers, 2,3-bis(trifluoromethyl)bicyclo[2.2.2]octa-2,5-diene . and 2,3- (tetrafluorobenzo)bicyclo[2.2.2]octatriene are described finally, Chapter 6 provides a summary of the work reported and outlines some ideas for future studies
CREDIT RATING AGENCIES AND THEIR POTENTIAL IMPACT ON DEVELOPING COUNTRIES
No full textCredit rating agencies (CRAs) play a key role in financial markets by helping to reduce the informative asymmetry between lenders and investors, on one side, and issuers on the other side, about the creditworthiness of companies or countries. CRAs´ role has expanded with financial globalization and has received an additional boost from Basel II which incorporates the ratings of CRAs into the rules for setting weights for credit risk. Ratings tend to be sticky, lagging markets, and overreact when they do change. This overreaction may have aggravated financial crises in the recent past, contributing to financial instability and cross-country contagion. The recent bankruptcies of Enron, WorldCom, and Parmalat have prompted legislative scrutiny of the agencies. Criticism has been especially directed towards the high degree of concentration of the industry. Promotion of competition may require policy action at national and international level to encourage the establishment of new agencies and to channel business generated by new regulatory requirements in their direction.
Crystal structure of a prokaryotic homologue of the mammalian oligopeptide–proton symporters, PepT1 and PepT2
No full textPepT1 and PepT2 are major facilitator superfamily (MFS) transporters that utilize a proton gradient to drive the uptake of di- and tri-peptides in the small intestine and kidney, respectively. They are the major routes by which we absorb dietary nitrogen and many orally administered drugs. Here, we present the crystal structure of PepTSo, a functionally similar prokaryotic homologue of the mammalian peptide transporters from Shewanella oneidensis. This structure, refined using data up to 3.6 Å resolution, reveals a ligand-bound occluded state for the MFS and provides new insights into a general transport mechanism. We have located the peptide-binding site in a central hydrophilic cavity, which occludes a bound ligand from both sides of the membrane. Residues thought to be involved in proton coupling have also been identified near the extracellular gate of the cavity. Based on these findings and associated kinetic data, we propose that PepTSo represents a sound model system for understanding mammalian peptide transport as catalysed by PepT1 and PepT2
Exploring high-end scenarios for local sea level rise to develop flood protection strategies for a low-lying delta-the Netherlands as an example
No full textSea level rise, especially combined with possible changes in storm surges and increased river discharge resulting from climate change, poses a major threat in low-lying river deltas. In this study we focus on a specific example of such a delta: the Netherlands. To evaluate whether the country’s flood protection strategy is capable of coping with future climate conditions, an assessment of low-probability/high-impact scenarios is conducted, focusing mainly on sea level rise. We develop a plausible high-end scenario of 0.55 to 1.15 m global mean sea level rise, and 0.40 to 1.05 m rise on the coast of the Netherlands by 2100 (excluding land subsidence), and more than three times these local values by 2200. Together with projections for changes in storm surge height and peak river discharge, these scenarios depict a complex, enhanced flood risk for the Dutch delta
Energy-efficient cooperative single-carrier frequency-division multiple-access
No full textA variety of cooperative relaying schemes are designed for the single-carrier frequency-division multiple-access (SC-FDMA) uplink, when communicating over broadband wireless channels. Our goal is to reduce the battery power dissipated both by transmission and signal processing, so that the overall energy-efficiency may be increased. We assume that there are a number of inactive mobile terminals acting as potential relays, which have either fixed or time-variant positions in a cell. Our investigations are focused on the optimum exploitation of all the resources, when considering relay selection, power allocation and channel-quality-aided adaptive subband allocation. We exploit the benefits of combining the path-loss reduction and diversity gains arising from both fixed and opportunistic relaying, user cooperation and from all the propagation paths, as well as from multiple antennas. Novel frequency-domain equalisation and diversity combining approaches are also conceived.Specifically, we firstly conceive two single-relay assisted topologies for the sake of exploiting the achievable cooperative diversity, namely the single-dedicated-relaying (SDR), where each relay is dedicated to a single user, and the single-shared-relaying (SSR), when a single relay assists multiple users. In order to eliminate both the multi-user interference and for the sake of mitigating the noise-amplification imposed by amplify-and-forward (AF) relaying, we propose an efficient subband-based AF scheme, which is benchmarked against the conventional AF regime in the context of both the SDR and SSR topologies. Furthermore, by assuming that the channel state information (CSI) is available at the base station (BS)’s receiver, a joint frequency-domain equalisation and diversity-combining scheme is proposed for the sake of increasing the achievable cooperative diversity gain. In this case, when considering the different number of available relays that are geographically dispersed across a large-scale environment subject to both path-loss and shadowing, we propose three different dynamic relay selection schemes, namely single-user relay selection (SU-RS), multi-user relay selection (MU-RS), and multiple-access relay selection (MA-RS), combined with source/relay vi power allocation in the context of opportunistic cooperation (OC) for the sake of increasing the multi-user system’s throughput. By contrast, when the source-to-destination (S-D) direct links are of low quality and hence are deemed to be unavailable, we exploit the relays which are roaming within each other’s vicinity in geographically localised manner in a cluster. Therefore, by assuming that these cooperating relays are capable of exchanging their channel quality information (CQI), we propose two first-hop-quality-aware (FHQA) joint dynamic resource allocation (DRA) schemes for opportunistic relaying (OR) based SCFDMA uplink, which beneficially combines channel-quality-aware subband allocation with efficient relay selection. The FHQA joint DRA schemes optimise the multi-user multi-relay networks relying on whether it is the source-to-relay (S-R) or the relay-to-destination (R-D) link, which dominates the attainable performance, when the BS’s receiver employs either single or multiple antennas. Additionally, the benefits of OR are quantified in the context of interleaver-aided decode-and-forward (DF) relaying for transmission over correlated fading channels. Therefore, the length of the interleavers combined with channel coding may be shortened. As a result, we benefit from a reduced interleaving delay and/or from a total transmit power reduction. In comparison to the benchmark schemes considered in the literature, the reliability and energy-efficiency of our proposed systems are significantly improved
Cryo-EM Structure and Molecular Dynamics Analysis of the Fluoroquinolone Resistant Mutant of the AcrB Transporter from Salmonella
No full textSalmonella is an important genus of Gram-negative pathogens, treatment of which has become problematic due to increases in antimicrobial resistance. This is partly attributable to the overexpression of tripartite efflux pumps, particularly the constitutively expressed AcrAB-TolC. Despite its clinical importance, the structure of the Salmonella AcrB transporter remained unknown to-date, with much of our structural understanding coming from the Escherichia coli orthologue. Here, by taking advantage of the styrene maleic acid (SMA) technology to isolate membrane proteins with closely associated lipids, we report the very first experimental structure of Salmonella AcrB transporter. Furthermore, this novel structure provides additional insight into mechanisms of drug efflux as it bears the mutation (G288D), originating from a clinical isolate of Salmonella Typhimurium presenting an increased resistance to fluoroquinolones. Experimental data are complemented by state-of-the-art molecular dynamics (MD) simulations on both the wild type and G288D variant of Salmonella AcrB. Together, these reveal several important differences with respect to the E. coli protein, providing insights into the role of the G288D mutation in increasing drug efflux and extending our understanding of the mechanisms underlying antibiotic resistance
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