How We Make DNA Origami

DNA origami has attracted substantial attention since its invention ten years ago, due to the seemingly infinite possibilities that it affords for creating customized nanoscale objects. Although the basic concept of DNA origami is easy to understand, using custom DNA origami in practical applications requires detailed know‐how for designing and producing the particles with sufficient quality and for preparing them at appropriate concentrations with the necessary degree of purity in custom environments. Such know‐how is not readily available for newcomers to the field, thus slowing down the rate at which new applications outside the field of DNA nanotechnology may emerge. To foster faster progress, we share in this article the experience in making and preparing DNA origami that we have accumulated over recent years. We discuss design solutions for creating advanced structural motifs including corners and various types of hinges that expand the design space for the more rigid multilayer DNA origami and provide guidelines for preventing undesired aggregation and on how to induce specific oligomerization of multiple DNA origami building blocks. In addition, we provide detailed protocols and discuss the expected results for five key methods that allow efficient and damage‐free preparation of DNA origami. These methods are agarose‐gel purification, filtration through molecular cut‐off membranes, PEG precipitation, size‐exclusion chromatography, and ultracentrifugation‐based sedimentation. The guide for creating advanced design motifs and the detailed protocols with their experimental characterization that we describe here should lower the barrier for researchers to accomplish the full DNA origami production workflow.     

Effects of polymer modification on the fuel resistance of asphalt binders

Fuel resistance is an important characteristic of asphalt binders, especially when used in airport fields and filling stations. However, the relevant standards are presently lacking and the scientific literature on the development of suited formulations is sparse. In this paper, a simple procedure to assess the fuel resistance of asphalts (considered in a dynamic sense) is proposed. A number of polymer modified asphalts (PMAs), prepared from two bases and 4 or 6 wt% of several polymers, have been analyzed with this procedure and the relationships between fuel resistance and morphology of the PMAs, the chemical structure of the polymers and the composition of the asphalt bases have been investigated.     

Role of the Initiation Factors in mRNA Start Site Selection and fMet-tRNA Recruitment by Bacterial Ribosomes

Translation initiation is a complex, multi-step process of fundamental importance in all kingdoms of life, during which the start site of the genetic message transmitted in the form of an RNA molecule (mRNA) is selected, and the level of translation determined. Being the slowest step of protein synthesis, initiation is the phase most often subject to regulation. Here we review, in a historical perspective and focusing mainly on results from our laboratory, the development of our perception of the mechanisms by which the most relevant steps of this pathway occur in bacteria. In particular, we describe: (a) the mechanistics and kinetics of translation initiation; (b) properties of mRNAs with and without Shine–Dalgarno sequence relevant for initiation site selection and translational efficiency; c) ribosomal binding and dissociation of the initiation factors, formation and properties of translation initiation intermediates; (d) the mechanisms by which translation initiation fidelity is ensured. Finally, we provide a short survey of the known translation initiation inhibitors.     

ANP and BNP Exert Anti-Inflammatory Action via NPR-1/cGMP Axis by Interfering with Canonical,Non-Canonical,and Alternative Routes of Inflammasome Activation in Human THP1 Cells

Dysregulated inflammasome activation and interleukin (IL)-1β production are associated with several inflammatory disorders. Three different routes can lead to inflammasome activation: a canonical two-step, a non-canonical Caspase-4/5- and Gasdermin D-dependent, and an alternative Caspase-8-mediated pathway. Natriuretic Peptides (NPs), Atrial Natriuretic Peptide (ANP) and B-type Natriuretic Peptide (BNP), binding to Natriuretic Peptide Receptor-1 (NPR-1), signal by increasing cGMP (cyclic guanosine monophosphate) levels that, in turn, stimulate cGMP-dependent protein kinase-I (PKG-I). We previously demonstrated that, by counteracting inflammasome activation, NPs inhibit IL-1β secretion. Here we aimed to decipher the molecular mechanism underlying NPs effects on THP-1 cells stimulated with lipopolysaccharide (LPS) + ATP. Involvement of cGMP and PKG-I were assessed pre-treating THP-1 cells with the membrane-permeable analogue, 8-Br-cGMP, and the specific inhibitor KT-5823, respectively. We found that NPs, by activating NPR-1/cGMP/PKG-I axis, lead to phosphorylation of NLRP3 at Ser295 and to inflammasome platform disassembly. Moreover, by increasing intracellular cGMP levels and activating phosphodiesterases, NPs interfere with both Gasdermin D and Caspase-8 cleavage, indicating that they disturb non-canonical and alternative routes of inflammasome activation. These results showed that ANP and BNP anti-inflammatory and immunomodulatory actions may involve the inhibition of all the known routes of inflammasome activation. Thus, NPs might be proposed for the treatment of the plethora of diseases caused by a dysregulated inflammasome activation.     

Connexin 43 and Connexin 26 Involvement in the Ponatinib-Induced Cardiomyopathy: Sex-Related Differences in a Murine Model

Cardiac connexins (Cxs) are proteins responsible for proper heart function. They form gap junctions that mediate electrical and chemical signalling throughout the cardiac system, and thus enable a synchronized contraction. Connexins can also individually participate in many signal transduction pathways, interacting with intracellular proteins at various cellular compartments. Altered connexin expression and localization have been described in diseased myocardium and the aim of this study is to assess the involvement of Cx43, Cx26, and some related molecules in ponatinib-induced cardiac toxicity. Ponatinib is a new multi-tyrosine kinase inhibitor that has been successfully used against human malignancies, but its cardiotoxicity remains worrisome. Therefore, understanding its signaling mechanism is important to adopt potential anti cardiac damage strategies. Our experiments were performed on hearts from male and female mice treated with ponatinib and with ponatinib plus siRNA-Notch1 by using immunofluorescence, Western blotting, and proteomic analyses. The altered cardiac function and the change in Cxs expression observed in mice after ponatinib treatment, were results dependent on the Notch1 pathway and sex. Females showed a lower susceptibility to ponatinib than males. The downmodulation of cardiac Cx43, Cx26 and miR-122, high pS368-Cx43 phosphorylation, cell viability and survival activation could represent some of the female adaptative/compensatory reactions to ponatinib cardiotoxicity.     

A control strategy for variable-speed variable-pitch wind turbines within the regions of partial- and full-load operation without wind speed feedback

In this paper, the control of a variable-speed variable-pitch wind turbine in the whole wind speed range is addressed, without any feedback measurement of wind speed. In addition to an aerodynamic torque observer able to ensure the tracking of the maximum delivered power in the partial-load region, a novel wind speed observer is proposed for power regulation in the full-load region, along with a sliding surface ensuring finite-time set-point stabilization of the speed tracking error. The proposed control solution has been validated on the National Renewable Energy Laboratory 5-MW three-blade wind turbine model using the recognized high-fidelity simulation tool FAST.     

Efficient multiresolution time-domain analysis of arbitrarily shaped photonic devices

A multiresolution time-domain (MRTD) technique is proposed and applied to the analysis of arbitrarily shaped photonic devices. The suggested method implements the multiresolution analysis in the context of method-of-moments for the solution of Maxwell?s equations. To improve the capabilities of the proposed method, the uniaxial perfectly matched layers absorber for the MRTD is rigorously incorporated and better performance is reported over the conventional finite-difference time-domain. Various numerical examples demonstrate the stability and numerical precision of the suggested MRTD method for both linear and nonlinear applications. Moreover, the application of the suggested MRTD scheme for the design of a photonic crystal-based optical wavelength filter and for the analysis of a frequency converter is presented.     

Development and testing of the Clinical Post-Conference Learning Environment Survey

Clinical postconference is an integral component of the required laboratory hours in undergraduate baccalaureate nursing education. Empirical evidence supporting the educational benefits of this activity are nearly nonexistent. This article describes the development and psychometric testing of a self-report instrument designed to measure clinical postconference learning environments as perceived by undergraduate nursing students and faculty. The Clinical Post-Conference Learning Environment Survey (CPCLES) consists of 54 items and has been tested with more than 500 participants in three schools of nursing in the Midwest. Based on theoretical support and comprehensive review of the literature, six components of the learning environment are measured in two forms of the instrument; an actual scale and an importance scale. Content validity, internal consistency, and test-retest reliability are discussed. Using the CPCLES, perceptions of undergraduate nursing students and faculty were measured. Significant differences between the actual components of the learning environment and the correlated ratings of the importance of these components were found. Faculty perceived a significantly greater amount of teacher support, task orientation, and innovation in the postconference learning environment than did undergraduate students. No differences were noted between faculty and students regarding the importance of the learning environment components. This study presents findings with the use of the first instrument developed to measure clinical postconference learning environments. The congruence and discrepancy among and between learning groups' perceptions carry implications for educational practices in this setting. Subseqent investigations using this tool may be able to link the perceived learning environment to valued cognitive, affective, and behavioral learning outcomes.     

Response of obese binge eaters to treatment by behavior therapy combined with very low calorie diet.

Examined attrition and weight loss in 235 female obese binge eaters, episodic overeaters, and nonbingers treated by a 26-wk program of behavior modification and very low calorie diet. No significant differences were observed among conditions in the number of Ss who completed treatment. Episodic overeaters, however, were more likely than Ss in the other 2 conditions to drop out during the last 7 wks of treatment, when Ss resumed consumption of a conventional diet. End-of-treatment weight losses for the 3 conditions, which did not differ significantly, averaged 21.5, 19.4, and 21.7 kg, respectively. No significant differences were observed among conditions in weight regain (which averaged 8.8 kg) in the year following treatment, although small sample sizes prevented an adequate evaluation. (PsycINFO Database Record (c) 2010 APA, all rights reserved)