In vitro selection and characterization of cellulose-binding DNA aptamers

Many nucleic acid enzymes and aptamers have modular architectures that allow them to retain their functions when combined with other nucleotide sequences. This modular function facilitates the engineering of RNAs and DNAs that have more complex functions. We sought to create new DNA aptamers that bind cellulose to provide a module for immobilizing DNAs. Cellulose has been used in a variety of applications ranging from coatings and films to pharmaceutical preparations, and therefore DNA aptamers that bind cellulose might enable new applications. We used in vitro selection to isolate aptamers from a pool of random-sequence DNAs and subjected two distinct clones to additional rounds of mutagenesis and selection. One aptamer (CELAPT 14) was chosen for sequence minimization and more detailed biochemical analysis. CELAPT 14 aptamer variants exhibit robust binding both to cellulose powder and paper. Also, an allosteric aptamer construct was engineered that exhibits ATP-mediated cellulose binding during paper chromatography

In situ deformation observation via EBSD and EDS during high temperature tensile testing

Tensile testing is the backbone of mechanical characterization for materials science. The possibility to combine mechanical testing with advanced imaging and characterization methods and the option to operate at high temperatures up to 800°C opens a large variety of possibilities for materials research. In this work in situ annealing experiments are shown, where the grain growth is observed via EBSD over the course of the experiment. Different annealing states are achieved and tested after cooling to room temperature. Using the EBSD information, high Schmid factor grains can easily be identified and monitored during the in situ tensile experiment and therefore even the first yielding grains are captured. Further in situ high temperature tensile tests on steel samples up to a temperature of 800 °C are presented. An example of a tested steel specimen is shown in Figure 1. Here, slip band formation is easily observable in BSD contrast. By enabling feature tracking, the chosen region of interest remains in the field of view and is imaged correctly. Please click Download on the upper right corner to see the full abstract

Riboswitches Control Fundamental Biochemical Pathways in Bacillus subtilis and Other Bacteria

AbstractRiboswitches are metabolite binding domains within certain messenger RNAs that serve as precision sensors for their corresponding targets. Allosteric rearrangement of mRNA structure is mediated by ligand binding, and this results in modulation of gene expression. We have identified a class of riboswitches that selectively recognizes guanine and becomes saturated at concentrations as low as 5 nM. In Bacillus subtilis, this mRNA motif is located on at least five separate transcriptional units that together encode 17 genes that are mostly involved in purine transport and purine nucleotide biosynthesis. Our findings provide further examples of mRNAs that sense metabolites and that control gene expression without the need for protein factors. Furthermore, it is now apparent that riboswitches contribute to the regulation of numerous fundamental metabolic pathways in certain bacteria

Exome Sequencing of a Multigenerational Human Pedigree

Over the next few years, the efficient use of next-generation sequencing (NGS) in human genetics research will depend heavily upon the effective mechanisms for the selective enrichment of genomic regions of interest. Recently, comprehensive exome capture arrays have become available for targeting approximately 33 Mb or ∼180,000 coding exons across the human genome. Selective genomic enrichment of the human exome offers an attractive option for new experimental designs aiming to quickly identify potential disease-associated genetic variants, especially in family-based studies. We have evaluated a 2.1 M feature human exome capture array on eight individuals from a three-generation family pedigree. We were able to cover up to 98% of the targeted bases at a long-read sequence read depth of ≥3, 86% at a read depth of ≥10, and over 50% of all targets were covered with ≥20 reads. We identified up to 14,284 SNPs and small indels per individual exome, with up to 1,679 of these representing putative novel polymorphisms. Applying the conservative genotype calling approach HCDiff, the average rate of detection of a variant allele based on Illumina 1 M BeadChips genotypes was 95.2% at ≥10x sequence. Further, we propose an advantageous genotype calling strategy for low covered targets that empirically determines cut-off thresholds at a given coverage depth based on existing genotype data. Application of this method was able to detect >99% of SNPs covered ≥8x. Our results offer guidance for “real-world” applications in human genetics and provide further evidence that microarray-based exome capture is an efficient and reliable method to enrich for chromosomal regions of interest in next-generation sequencing experiments

Insights into the evolution of Darwin’s finches from comparative analysis of the Geospiza magnirostris genome sequence

Background: A classical example of repeated speciation coupled with ecological diversification is the evolution of 14 closely related species of Darwin’s (Galápagos) finches (Thraupidae, Passeriformes). Their adaptive radiation in the Galápagos archipelago took place in the last 2–3 million years and some of the molecular mechanisms that led to their diversification are now being elucidated. Here we report evolutionary analyses of genome of the large ground finch, Geospiza magnirostris. Results: 13,291 protein-coding genes were predicted from a 991.0 Mb G. magnirostris genome assembly. We then defined gene orthology relationships and constructed whole genome alignments between the G. magnirostris and other vertebrate genomes. We estimate that 15% of genomic sequence is functionally constrained between G. magnirostris and zebra finch. Genic evolutionary rate comparisons indicate that similar selective pressures acted along the G. magnirostris and zebra finch lineages suggesting that historical effective population size values have been similar in both lineages. 21 otherwise highly conserved genes were identified that each show evidence for positive selection on amino acid changes in the Darwin's finch lineage. Two of these genes (Igf2r and Pou1f1) have been implicated in beak morphology changes in Darwin’s finches. Five of 47 genes showing evidence of positive selection in early passerine evolution have cilia related functions, and may be examples of adaptively evolving reproductive proteins. Conclusions: These results provide insights into past evolutionary processes that have shaped G. magnirostris genes and its genome, and provide the necessary foundation upon which to build population genomics resources that will shed light on more contemporaneous adaptive and non-adaptive processes that have contributed to the evolution of the Darwin’s finches.Organismic and Evolutionary Biolog

Bedrock geology of DFDP-2B, central Alpine Fault, New Zealand

<p>During the second phase of the Alpine Fault, Deep Fault Drilling Project (DFDP) in the Whataroa River, South Westland, New Zealand, bedrock was encountered in the DFDP-2B borehole from 238.5–893.2 m Measured Depth (MD). Continuous sampling and meso- to microscale characterisation of whole rock cuttings established that, in sequence, the borehole sampled amphibolite facies, Torlesse Composite Terrane-derived schists, protomylonites and mylonites, terminating 200–400 m above an Alpine Fault Principal Slip Zone (PSZ) with a maximum dip of 62°. The most diagnostic structural features of increasing PSZ proximity were the occurrence of shear bands and reduction in mean quartz grain sizes. A change in composition to greater mica:quartz + feldspar, most markedly below c. 700 m MD, is inferred to result from either heterogeneous sampling or a change in lithology related to alteration. Major oxide variations suggest the fault-proximal Alpine Fault alteration zone, as previously defined in DFDP-1 core, was not sampled.</p

Standardised pre-operative diagnostics and treatment of peripheral arterial disease reduce wound complications in geriatric ankle fractures

Purpose The aim of this study was to evaluate a standardised algorithm to assess and treat impaired limb perfusion prior to surgical fixation of geriatric ankle fractures and determine the prevalence of peripheral arterial disease (PAD) in geriatric patients presenting with ankle fractures. Methods Eighty-four patients > 65 years pre-operatively diagnosed and treated according to an algorithm (study group) were compared with 84 patients diagnosed and treated before the algorithm was introduced (control group). Results In 14 patients of the study group, clinical noninvasive examination revealed signs of relevant PAD, which was confirmed with computed tomographic angiography (CTA) in nine patients, all of whom had successful angioplasty prior to surgical fixation of the ankle fracture. In three of these patients, PAD had previously been diagnosed. After standardised diagnostics and treatment of malperfusion, a significantly reduced overall and, particularly, wound complication rate was found. Conclusion PAD is an underdiagnosed condition in geriatric patients presenting with ankle fractures. This study underlines the relevance of limb perfusion for adequate wound healing in geriatric ankle fractures. Therefore, special attention should be paid to diagnose and-if indicated-optimise limb perfusion prior to surgical fixation of geriatric ankle fractures

Femoral offset following trochanteric femoral fractures: a prospective observational study

Background: Reconstruction of the femoral offset reportedly improves outcome following total hip arthroplasty, but little is known of its influence following hip fractures. We aimed to establish the effect of the femoral offset on the medium-term functional outcome in elderly patients who had sustained trochanteric fractures requiring proximal femoral nailing. Patients and Methods: We measured the rotation corrected femoral offset (FORC) and relative femoral offset (FORL) on plain anteroposterior radiographs of the hip in 188 patients (58 male, 130 female) with a trochanteric fracture who underwent proximal femoral nailing at our institution. The primary outcome measure was the Harris hip score (HSS) 6 and 12 months postoperatively; the Barthel index was assessed as a secondary outcome. Results: The mean FORC after surgery was 58 mm (+/- 11 mm), while the mean FORL was 1.21 (+/- 0.22). At final follow up, we found significant inverse relationships (Spearman's rank correlation coefficient,.) between FORC and FORL and the functional outcome assessed by the HSS (FORC : rho = -0.207, p = 0.036; FORL : rho = -0.247, p = 0.012), and FORL and the Barthel index (FORC : rho = -147, p = 0.129; FORL : rho = -0.192, p = 0.046). A consistent trend was observed after adjustment for confounding variables. Conclusions: Our results underline the biomechanical importance of the femoral offset for medium-term outcomes in elderly patients with trochanteric fractures. In contrast with the published findings on total hip arthroplasty, we found an inverse correlation between functional outcome and the extent of the reconstructed femoral offset. (C) 2015 Elsevier Ltd. All rights reserved