Effects of a recombinant gene expression on ColE1-like plasmid segregation in Escherichia coli

<p>Abstract</p> <p>Background</p> <p>Segregation of expression plasmids leads to loss of recombinant DNA from transformed bacterial cells due to the irregular distribution of plasmids between the daughter cells during cell division. Under non-selective conditions this segregational instability results in a heterogeneous population of cells, where the non-productive plasmid-free cells overgrow the plasmid-bearing cells thus decreasing the yield of recombinant protein. Amongst the factors affecting segregational plasmid instability are: the plasmid design, plasmid copy-number, host cell genotype, fermentation conditions etc. This study aims to investigate the influence of transcription and translation on the segregation of recombinant plasmids designed for constitutive gene expression in <it>Escherichia coli </it>LE392 at glucose-limited continuous cultivation. To this end a series of pBR322-based plasmids carrying a synthetic human interferon-gamma (hIFNγ) gene placed under the control of different regulatory elements (promoter and ribosome-binding sites) were used as a model.</p> <p>Results</p> <p>Bacterial growth and product formation kinetics of transformed <it>E. coli </it>LE392 cells cultivated continuously were described by a structured kinetic model proposed by Lee et al. (1985). The obtained results demonstrated that both transcription and translation efficiency strongly affected plasmid segregation. The segregation of plasmid having a deleted promoter did not exceed 5% after 190 h of cultivation. The observed high plasmid stability was not related with an increase in the plasmid copy-number. A reverse correlation between the yield of recombinant protein (as modulated by using different ribosome binding sites) and segregational plasmid stability (determined by the above model) was also observed.</p> <p>Conclusions</p> <p>Switching-off transcription of the hIFNγ gene has a stabilising effect on ColE1-like plasmids against segregation, which is not associated with an increase in the plasmid copy-number. The increased constitutive gene expression has a negative effect on segregational plasmid stability. A kinetic model proposed by Lee et al. (1985) was appropriate for description of <it>E. coli </it>cell growth and recombinant product formation in chemostat cultivations.</p

Absorptive capacity and innovation: When is it better to cooperate?

Cooperation can benefit and hurt firms at the same time. An important question then is: when is it better to cooperate? And, once the decision to cooperate is made, how can an appropriate partner be selected? In this paper we present a model of inter-firm cooperation driven by cognitive distance, appropriability conditions and external knowledge. Absorptive capacity of firms develops as an outcome of the interaction between absorptive R&D and cognitive distance from voluntary and involuntary knowledge spillovers. Thus, we offer a revision of the original model by Cohen and Levinthal (Econ J 99(397):569-596, 1989), accounting for recent empirical findings and explicitly modeling absorptive capacity within the framework of interactive learning. We apply that to the analysis of firms' cooperation and R&D investment preferences. The results show that cognitive distance and appropriability conditions between a firm and its cooperation partner have an ambiguous effect on the profit generated by the firm. Thus, a firm chooses to cooperate and selects a partner conditional on the investments in absorptive capacity it is willing to make to solve the understandability/novelty trade-off. © 2014 Springer-Verlag Berlin Heidelberg

Determining crystal structures through crowdsourcing and coursework

We show here that computer game players can build high-quality crystal structures. Introduction of a new feature into the computer game Foldit allows players to build and real-space refine structures into electron density maps. To assess the usefulness of this feature, we held a crystallographic model-building competition between trained crystallographers, undergraduate students, Foldit players and automatic model-building algorithms. After removal of disordered residues, a team of Foldit players achieved the most accurate structure. Analysing the target protein of the competition, YPL067C, uncovered a new family of histidine triad proteins apparently involved in the prevention of amyloid toxicity. From this study, we conclude that crystallographers can utilize crowdsourcing to interpret electron density information and to produce structure solutions of the highest quality

Virtual Monoenergetic Images from Spectral Detector CT Enable Radiation Dose Reduction in Unenhanced Cranial CT

BACKGROUND AND PURPOSE: Our aim was to evaluate whether improved gray-white matter differentiation in cranial CT by means of 65-keV virtual monoenergetic images enables a radiation dose reduction compared to conventional images. MATERIALS AND METHODS: One hundred forty consecutive patients undergoing 171 spectral detector CTs of the head between February and November 2017 (56 +/- 19 years of age; male/female ratio, 56%/44%) were retrospectively included. The tube current-time product was reduced during the study period, resulting in 61, 55, and 55 patients being examined with 320, 290, and 260 mAs, respectively. All other scanning parameters were kept identical. The volume CT dose index was recorded. ROIs were placed in gray and white matter on conventional images and copied to identical positions in 65- keV virtual monoenergetic images. The contrast-to-noise ratio was calculated. Two radiologists blinded to the reconstruction technique evaluated image quality on a 5-point Likert-scale. Statistical assessment was performed using ANOVA and Wilcoxon test adjusted for multiple comparisons. RESULTS: The mean volume CT dose index was 55, 49.8, and 44.7 mGy using 320, 290, and 260 mAs, respectively. Irrespective of the volume CT dose index, noise was significantly lower in 65- keV virtual monoenergetic images compared with conventional images (65- keV virtual monoenergetic images/conventional images: extraocular muscle with 49.8 mGy, 3.7 +/- 1.3/5.6 +/- 1.6 HU, P < .001). Noise slightly increased with a reduced radiation dose (eg, extraocular muscle in conventional images: 5.3 +/- 1.4/5.6 +/- 1.6/6.1 +/- 2.1 HU). Overall, the contrast-to-noise ratio in 65- keV virtual monoenergetic images was superior to that in conventional images irrespective of the volume CT dose index (P < .001). Particularly, 65-keV virtual monoenergetic images with 44.7 mGy showed significantly lower noise and a higher contrast-to-noise ratio than conventional images with 55 mGy (P < .001). Subjective analysis confirmed better image quality in 65- keV virtual monoenergetic images, even using 44.7 mGy. CONCLUSIONS: The 65-keV virtual monoenergetic images from spectral detector CT allow radiation dose reduction in cranial CT. While this proof of concept included a radiation dose reduction of 19%, our data suggest that even greater reduction appears achievable

Interacting polariton fluids in a monolayer of tungsten disulfide

Atomically thin transition metal dichalcogenides (TMDs) possess a number of properties that make them attractive for realizing room-temperature polariton devices. An ideal platform for manipulating polariton fluids within monolayer TMDs is that of Bloch surface waves, which confine the electric field to a small volume near the surface of a dielectric mirror. Here we demonstrate that monolayer tungsten disulfide ($\text{WS}_2$) can sustain Bloch surface wave polaritons (BSWPs) with a Rabi splitting of 43 meV and propagation constants reaching 33 $\mu$m. In addition, we evidence strong polariton-polariton nonlinearities within BSWPs, which manifest themselves as a reversible blueshift of the lower polariton resonance by up to 12.9$\pm$0.5 meV. Such nonlinearities are at the heart of polariton devices and have not yet been demonstrated in TMD polaritons. As a proof of concept, we use the nonlinearity to implement a nonlinear polariton source. Our results demonstrate that BSWPs using TMDs can support long-range propagation combined with strong nonlinearities, enabling potential applications in integrated optical processing and polaritonic circuits.Comment: 7 pages, 4 figure