Hybridisation based DNA screening on peptide nucleic acid (PNA) oligomer arrays.

Arrays of up to some 1000 PNA oligomers of individual sequence were synthesised on polymer membranes using a robotic device originally designed for peptide synthesis. At approximately 96%, the stepwise synthesis efficiency was comparable to standard PNA synthesis procedures. Optionally, the individual, fully deprotected PNA oligomers could be removed from the support for further use, because an enzymatically cleavable but otherwise stable linker was used. Since PNA arrays could form powerful tools for hybridisation based DNA screening assays due to some favourable features of the PNA molecules, the hybridisation behaviour of DNA probes to PNA arrays was investigated for a precise understanding of PNA-DNA interactions on solid support. Hybridisation followed the Watson-Crick base pairing rules with higher duplex stabilities than on corresponding DNA oligonucleotide sensors. Both the affinity and specificity of DNA hybridisation to the PNA oligomers depended on the hybridisation conditions more than expected. Successful discrimination between hybridisation to full complementary PNA sequences and truncated or mismatched versions was possible at salt concentrations down to 10 mM Na+and below, although an increasing tendency to unspecific DNA binding and few strong mismatch hybridisation events were observed.     

ETHYL ALCOHOL REDUCTION OF SALMONELLA TYPHIMURIUM IN POULTRY FEED

This study was designed to evaluate the effect of ethyl alcohol as a potential treatment for reduction of Salmonella populations in poultry feed. Growth rate of S. typhimurium in tryptic soy broth was significantly reduced by addition of greater than 0.3% volume/volume of ethyl alcohol and growth was completely inhibited by addition of 5% ethyl alcohol. Ethyl alcohol concentrations of 20% volume/weight and greater significantly reduced initial S.typhimurium populations in poultry feed (for 20% treated, 2.31 ± 0.31 vs 3.39 ± 0.29 for untreated; P < 0.05). When feed treatment was administered either before or after inoculation of S. typhimurium with 60% ethyl alcohol or 0.04% buffered propionic acid, populations in feeds treated after inoculation were decreased to a nondetection level (< 1.0 log₁₀ CFU/g) by ethyl alcohol treatment but not by other treatments. Ethyl alcohol treatment may have the potential for reducing Salmonella spp. contamination in poultry feed.     

Research Design and the Politics of Abstraction: Unpacking the Environmentality of Scientific Practice in Socioecological Assessments

Scientific assessments of socio-ecological systems are becoming mainstays in guiding policymaking and other interventions in response to global environmental change. The environmentality literature emphasizes the institutional architecture of emergent science-policy regimes and how scientific research is used in political settings, creating new modes of governance and subjectivities. However, there has been relatively little attention to domain-level socio-ecological assessments as socially produced technologies where specific scientific choices are mechanisms connecting governance architecture and popular subjectivities.Combining empirical case study and literature review, assessment technologies are analyzed in three domains: vulnerability assessment, ecosystem services assessment, life-cycle assessment. Using conceptualization, operationalization, and institutionalization as analytical lenses, the cases illustrate ways that scientific choices simplify complex socio-ecological relationships with implications for both governance practices and subjectivities. Furthermore, findings explore the possibility for assessments to be more inclusive of diverse social values and practices, enabling more empowering subjectivities.     

A new device to quantify regenerate torsional stiffness in distraction osteogenesis.

We present a newly developed torsional stiffness measurement device with the potential to quantitatively assess the in vivo torsional stiffness of bone regenerate during distraction osteogenesis. We describe the form and function of this device and its application in a model of regenerate consolidation. The device was able to produce data to assess stiffness of the regenerate with an accuracy between +/- 3 and +/- 9% for material stiffness ranging between 0.1 and 2.4 Nm/o and with a precision of +/- 3.6%. This method provides advantages over similar methods of bone fracture healing assessment with guaranteed maintenance of bone axis, minimized risk of bone misalignment during the bone healing process and a close relation to the functional loading pattern in torsion of bones such as tibia and femora.     

The Agrobacterium vitis T-6b oncoprotein induces auxin-independent cell expansion in tobacco

Among the Agrobacterium T-DNA genes, rolB, rolC, orf13, orf8, lso, 6b and several other genes encode weakly homologous proteins with remarkable effects on plant growth. The 6b oncogene induces tumors and enations. In order to study its properties we have used transgenic tobacco plants that carry a dexamethasone-inducible 6b gene, dex-T-6b. Upon induction, dex-T-6b plants develop a large array of morphological modifications, some of which involve abnormal cell expansion. In the present investigation, dex-T-6b-induced expansion was studied in intact leaves and an in vitro leaf disc system. Although T-6b and indole-3-acetic acid (IAA) both induced expansion and were non-additive, T-6b expression did not increase IAA levels, nor did it induce an IAA-responsive gene. Fusicoccin (FC) is known to stimulate expansion by increasing cell wall plasticity. T-6b- and FC-induced expansion were additive at saturating FC concentrations, indicating that T-6b does not act by a similar mechanism to FC. T-6b expression led to higher leaf osmolality values, in contrast to FC, suggesting that the T-6b gene induces expansion by increasing osmolyte concentrations. Metabolite profiling showed that glucose and fructose played a major role in this increase. We infer that T-6b disrupts the osmoregulatory controls that govern cell expansion during development and wound healing.     

Protein kinase A-mediated phosphorylation of connexin36 in mouse retina results in decreased gap junctional communication between AII amacrine cells

Gap junctions in AII amacrine cells of mammalian retina participate in the coordination of the rod and cone signaling pathway involved in visual adaptation. Upon stimulation by light, released dopamine binds to D(1) receptors on AII amacrine cells leading to increased intracellular cAMP (cyclic adenosine monophosphate) levels. AII amacrine cells express the gap junctional protein connexin36 (Cx36). Phosphorylation of Cx36 has been hypothesized to regulate gap junctional activity of AII amacrine cells. However, until now in vivo phosphorylation of Cx36 has not been reported. Indeed, it had been concluded that Cx36 in bovine retina is not phosphorylated, but in vitro phosphorylation for Cx35, the bass ortholog of Cx36, had been shown. To clarify this experimental discrepancy, we examined protein kinase A (PKA)-induced phosphorylation of Cx36 in mouse retina as a possible mechanism to modulate the extent of gap junctional coupling. The cytoplasmic domains of Cx36 and the total Cx36 protein were phosphorylated in vitro by PKA. Mass spectroscopy revealed that all four possible PKA consensus motifs were phosphorylated; however, domains point mutated at the sites in question showed a prevalent usage of Ser-110 and Ser-293. Additionally, we demonstrated that Cx36 was phosphorylated in cultured mouse retina. Furthermore, activation of PKA increased the level of phosphorylation of Cx36. cAMP-stimulated, PKA-mediated phosphorylation of Cx36 protein was accompanied by a decrease of tracer coupling between AII amacrine cells. Our results link increased phosphorylation of Cx36 to down-regulation of permeability through gap junction channels mediating light adaptation in the retina.