The effects of mismatches on hybridization in DNA microarrays: determination of nearest neighbor parameters

Quantifying interactions in DNA microarrays is of central importance for a better understanding of their functioning. Hybridization thermodynamics for nucleic acid strands in aqueous solution can be described by the so-called nearest-neighbor model, which estimates the hybridization free energy of a given sequence as a sum of dinucleotide terms. Compared with its solution counterparts, hybridization in DNA microarrays may be hindered due to the presence of a solid surface and of a high density of DNA strands. We present here a study aimed at the determination of hybridization free energies in DNA microarrays. Experiments are performed on custom Agilent slides. The solution contains a single oligonucleotide. The microarray contains spots with a perfect matching complementary sequence and other spots with one or two mismatches: in total 1006 different probe spots, each replicated 15 times per microarray. The free energy parameters are directly fitted from microarray data. The experiments demonstrate a clear correlation between hybridization free energies in the microarray and in solution. The experiments are fully consistent with the Langmuir model at low intensities, but show a clear deviation at intermediate (non-saturating) intensities. These results provide new interesting insights for the quantification of molecular interactions in DNA microarrays.Comment: 31 pages, 5 figure

Level I fieldwork today: A study of contexts and perceptions.

The last comprehensive examination of the Level I fieldwork experience was performed 15 years ago (Shalik, 1990) and addressed the different types of settings in which fieldwork occurred; amounts and types of supervision; structure and scheduling of the Level I experiences; and the effects of supervising Level I students on productivity. Although every occupational therapy and occupational therapy assistant student encounters a number of Level I fieldwork opportunities, little is available describing the process and contexts of the Level I fieldwork experience today. This study, which examines 1,002 student reports on Level I fieldwork experiences, finds that Level I fieldwork today occurs in a wide variety of physical disability, pediatric, mental health, and emerging practice settings. Findings also indicate that, whereas most fieldwork educators are occupational therapy practitioners, more fieldwork educators are non-occupational therapists than in the past. Furthermore, although students reported opportunities to practice observation and communication across all settings, practice of other clinical skills was specific to type of settings, and opportunities to practice were limited. Student perceptions about opportunities for experiencing occupation-based practice, observation of theory in practice, and how students value different types of fieldwork experiences are addressed. In addition, this study explores the expansion of Level I fieldwork into emerging practice arenas and how students perceive those experiences

In Situ Assembly of Well-Defined MoS2 Slabs on Shape-Tailored Anatase TiO2 Nanostructures: Heterojunctions Role in Phenol Photodegradation

MoS2/TiO2-based nanostructures have attracted extensive attention due to their high performance in many fields, including photocatalysis. In this contribution, MoS2 nanostructures were prepared via an in situ bottom-up approach at the surface of shape-controlled TiO2 nanoparticles (TiO2 nanosheets and bipyramids). Furthermore, a multi-technique approach by combining electron microscopy and spectroscopic methods was employed. More in detail, the morphology/structure and vibrational/optical properties of MoS2 slabs on TiO2 anatase bipyramidal nanoparticles, mainly exposing {101} facets, and on TiO2 anatase nanosheets exposing both {001} and {101} facets, still covered by MoS2, were compared. It was shown that unlike other widely used methods, the bottom-up approach enabled the atomic-level growth of well-defined MoS2 slabs on TiO2 nanostructures, thus aiming to achieve the most effective chemical interactions. In this regard, two kinds of synergistic heterojunctions, namely, crystal face heterojunctions between anatase TiO2 coexposed {101} and {001} facets and semiconductor heterojunctions between MoS2 and anatase TiO2 nanostructures, were considered to play a role in enhancing the photocatalytic activity, together with a proper ratio of (101), (001) coexposed surfaces

Model-based probe set optimization for high-performance microarrays

A major challenge in microarray design is the selection of highly specific oligonucleotide probes for all targeted genes of interest, while maintaining thermodynamic uniformity at the hybridization temperature. We introduce a novel microarray design framework (Thermodynamic Model-based Oligo Design Optimizer, TherMODO) that for the first time incorporates a number of advanced modelling features: (i) A model of position-dependent labelling effects that is quantitatively derived from experiment. (ii) Multi-state thermodynamic hybridization models of probe binding behaviour, including potential cross-hybridization reactions. (iii) A fast calibrated sequence-similarity-based heuristic for cross-hybridization prediction supporting large-scale designs. (iv) A novel compound score formulation for the integrated assessment of multiple probe design objectives. In contrast to a greedy search for probes meeting parameter thresholds, this approach permits an optimization at the probe set level and facilitates the selection of highly specific probe candidates while maintaining probe set uniformity. (v) Lastly, a flexible target grouping structure allows easy adaptation of the pipeline to a variety of microarray application scenarios. The algorithm and features are discussed and demonstrated on actual design runs. Source code is available on request

Sex differences in clinical presentation, severity and outcome of stroke: Results from a hospital-based registry

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Articles Nearest-Neighbor Thermodynamics and NMR of DNA Sequences with Internal A‚A, C‚C, G‚G, and T‚T Mismatches †

ABSTRACT: Thermodynamic measurements are reported for 51 DNA duplexes with A‚A, C‚C, G‚G, and T‚T single mismatches in all possible Watson-Crick contexts. These measurements were used to test the applicability of the nearest-neighbor model and to calculate the 16 unique nearest-neighbor parameters for the 4 single like with like base mismatches next to a Watson-Crick pair. The observed trend in stabilities of mismatches at 37°C is G‚G > T‚T ≈ A‚A > C‚C. The observed stability trend for the closing Watson-Crick pair on the 5′ side of the mismatch is G‚C g C‚G g A‚T g T‚A. The mismatch contribution to duplex stability ranges from -2.22 kcal/mol for GGC‚GGC to +2.66 kcal/mol for ACT‚ ACT. The mismatch nearest-neighbor parameters predict the measured thermodynamics with average deviations of ∆G°3 7 ) 3.3%, ∆H°) 7.4%, ∆S°) 8.1%, and T M ) 1.1°C. The imino proton region of 1-D NMR spectra shows that G‚G and T‚T mismatches form hydrogen-bonded structures that vary depending on the Watson-Crick context. The data reported here combined with our previous work provide for the first time a complete set of thermodynamic parameters for molecular recognition of DNA by DNA with or without single internal mismatches. The results are useful for primer design and understanding the mechanism of triplet repeat diseases. DNA mismatches occur in vivo due to misincorporation of bases during replication (1), heteroduplex formation during homologous recombination (2), mutagenic chemicals (3, 4), ionizing radiation (5), and spontaneous deamination (6). Knowledge of the thermodynamics of DNA mismatches will be useful for elucidating the mechanisms of polymerase fidelity and mismatch repair efficiency. Moreover, thermodynamic parameters for mismatch formation are important for DNA secondary structure prediction (see http://sun2.science.wayne.edu/∼jslsun2 and http://mfold1.wustl.edu/∼mfold/dna/form1.cgi). Recent work has shown that triplet repeat sequences form transiently stable hairpins that contain like with like base mismatche

iCODEHOP: a new interactive program for designing COnsensus-DEgenerate Hybrid Oligonucleotide Primers from multiply aligned protein sequences

PCR amplification using COnsensus DEgenerate Hybrid Oligonucleotide Primers (CODEHOPs) has proven to be highly effective for identifying unknown pathogens and characterizing novel genes. We describe iCODEHOP; a new interactive web application that simplifies the process of designing and selecting CODEHOPs from multiply-aligned protein sequences. iCODEHOP intelligently guides the user through the degenerate primer design process including uploading sequences, creating a multiple alignment, deriving CODEHOPs and calculating their annealing temperatures. The user can quickly scan over an entire set of degenerate primers designed by the program to assess their relative quality and select individual primers for further analysis. The program displays phylogenetic information for input sequences and allows the user to easily design new primers from selected sequence sub-clades. It also allows the user to bias primer design to favor specific clades or sequences using sequence weights. iCODEHOP is freely available to all interested researchers at https://icodehop.cphi.washington.edu/i-codehop-context/Welcome

Electrical detection of the temperature induced melting transition of a DNA hairpin covalently attached to gold interdigitated microelectrodes

The temperature induced melting transition of a self-complementary DNA strand covalently attached at the 5′ end to the surface of a gold interdigitated microelectrode (GIME) was monitored in a novel, label-free, manner. The structural state of the hairpin was assessed by measuring four different electronic properties of the GIME (capacitance, impedance, dissipation factor and phase angle) as a function of temperature from 25°C to 80°C. Consistent changes in all four electronic properties of the GIME were observed over this temperature range, and attributed to the transition of the attached single-stranded DNA (ssDNA) from an intramolecular, folded hairpin structure to a melted ssDNA. The melting curve of the self-complementary single strand was also measured in solution using differential scanning calorimetry (DSC) and UV absorbance spectroscopy. Temperature dependent electronic measurements on the surface and absorbance versus temperature values measured in solution experiments were analyzed assuming a two-state process. The model analysis provided estimates of the thermodynamic transition parameters of the hairpin on the surface. Two-state analyses of optical melting data and DSC measurements provided evaluations of the thermodynamic transition parameters of the hairpin in solution. Comparison of surface and solution measurements provided quantitative evaluation of the effect of the surface on the thermodynamics of the melting transition of the DNA hairpin

Transformative learning as pedagogy for the health professions : a scoping review

Context Transformative learning (TL) has been described as learning that challenges established perspectives, leading to new ways of being in the world. As a learning theory it has resonated with educators globally, including those in the health professions. Described as a complex metatheory, TL has evolved over time, eliciting divergent interpretations of the construct. This scoping review provides a comprehensive synthesis of how TL is currently represented in the health professions education literature, including how it influences curricular activities, to inform its future application in the field. Methods Arksey and O'Malley's six‐step framework was adopted to review the period from 2006 to May 2018. A total of 10 bibliographic databases were searched, generating 1532 potential studies. After several rounds of review, first of abstracts and then of full texts, 99 studies were mapped by two independent reviewers onto the internally developed data extraction sheet. Descriptive information about included studies was aggregated. Discursive data were subjected to content analysis. Results A mix of conceptual and empirical research papers, which used a range of qualitative methodologies, were included. Studies from the USA, the UK and Australia were most prevalent. Insights relating to how opportunities for TL were created, how it manifests and influences behaviour, as well as how it is experienced, demonstrated much congruency. Conceptions of TL were seen to be clustered around the work of key theorists. Conclusions The training of health professionals often takes place in unfamiliar settings where students are encouraged to be active participants in providing care. This increases the opportunity for exposure to learning experiences that are potentially transformative, allowing for a pedagogy of uncertainty that acknowledges the complexity of the world we live in and questions what we believe we know about it. TL provides educators in the health professions with a theoretical lens through which they can view such student learning

Modeling Bacterial DNA: Simulation of Self-avoiding Supercoiled Worm-Like Chains Including Structural Transitions of the Helix

Under supercoiling constraints, naked DNA, such as a large part of bacterial DNA, folds into braided structures called plectonemes. The double-helix can also undergo local structural transitions, leading to the formation of denaturation bubbles and other alternative structures. Various polymer models have been developed to capture these properties, with Monte-Carlo (MC) approaches dedicated to the inference of thermodynamic properties. In this chapter, we explain how to perform such Monte-Carlo simulations, following two objectives. On one hand, we present the self-avoiding supercoiled Worm-Like Chain (ssWLC) model, which is known to capture the folding properties of supercoiled DNA, and provide a detailed explanation of a standard MC simulation method. On the other hand, we explain how to extend this ssWLC model to include structural transitions of the helix.Comment: Book chapter to appear in The Bacterial Nucleoid, Methods and Protocols, Springer serie