Single-cell screening of photosynthetic growth and lactate production by cyanobacteria

Background Photosynthetic cyanobacteria are attractive for a range of biotechnological applications including biofuel production. However, due to slow growth, screening of mutant libraries using microtiter plates is not feasible. Results We present a method for high-throughput, single-cell analysis and sorting of genetically engineered l-lactate-producing strains of Synechocystis sp. PCC6803. A microfluidic device is used to encapsulate single cells in picoliter droplets, assay the droplets for l-lactate production, and sort strains with high productivity. We demonstrate the separation of low- and high-producing reference strains, as well as enrichment of a more productive l-lactate-synthesizing population after UV-induced mutagenesis. The droplet platform also revealed population heterogeneity in photosynthetic growth and lactate production, as well as the presence of metabolically stalled cells. Conclusions The workflow will facilitate metabolic engineering and directed evolution studies and will be useful in studies of cyanobacteria biochemistry and physiology

Towards Adaptive Technology in Routine Mental Healthcare

This paper summarizes the information technology-related research findings after 5 years with the INTROducing Mental health through Adaptive Technology project. The aim was to improve mental healthcare by introducing new technologies for adaptive interventions in mental healthcare through interdisciplinary research and development. We focus on the challenges related to internet-delivered psychological treatments, emphasising artificial intelligence, human-computer interaction, and software engineering. We present the main research findings, the developed artefacts, and lessons learned from the project before outlining directions for future research. The main findings from this project are encapsulated in a reference architecture that is used for establishing an infrastructure for adaptive internet-delivered psychological treatment systems in clinical contexts. The infrastructure is developed by introducing an interdisciplinary design and development process inspired by domain-driven design, user-centred design, and the person based approach for intervention design. The process aligns the software development with the intervention design and illustrates their mutual dependencies. Finally, we present software artefacts produced within the project and discuss how they are related to the proposed reference architecture. Our results indicate that the proposed development process, the reference architecture and the produced software can be practical means of designing adaptive mental health care treatments in correspondence with the patients’ needs and preferences. In summary, we have created the initial version of an information technology infrastructure to support the development and deployment of Internet-delivered mental health interventions with inherent support for data sharing, data analysis, reusability of treatment content, and adaptation of intervention based on user needs and preferences.publishedVersio

lac of Time : Transcription Factor Kinetics in Living Cells

Gene regulation mediated by transcription factors (TFs) is essential for all organisms. The functionality of TFs can largely be described by the fraction of time they occupy their regulatory binding sites on the chromosome. DNA-binding proteins have been shown to find their targets through facilitated diffusion in vitro. In its simplest form this means that the protein combines a random 3D search in the cytoplasm with 1D sliding along DNA. This has been proposed to speed up target location. It is difficult to mimic the in vivo conditions for gene regulation in biochemistry experiments; i.e. the ionic strength, chromosomal structure, and the presence of other DNA-binding macromolecules.    In this thesis single molecule imaging assays for live cell measurements were developed to study the kinetics of the Escherichia coli transcription factor LacI. The low copy number LacI, in fusion with a fluorescent protein (Venus) is detected as a localized near-diffraction limited spot when being DNA-bound for longer than the exposure time. An allosteric inducer is used to control binding and release. Using this method we can measure the time it takes for LacI to bind to different operator sequences. We then extend the assay and show that LacI slides in to and out from the operator site, and that it is obstructed by another DNA-binding protein positioned next to its target. We present a new model where LacI redundantly passes over the operator many times before binding.    By combining experiments with molecular dynamics simulations we can characterize the details of non-specific DNA-binding. In particular, we validate long-standing assumptions that the non-specific association is diffusion-controlled. In addition it is seen that the non-specifically bound protein diffuses along DNA in a helical path.    Using microfluidics we design a chase assay to measure in vivo dissociation rates for the LacI-Venus dimer. Based on the comparison of these rates with association rates and equilibrium binding data we suggest that there might be a short time following TF dissociation when transcription initiation is silenced. This implies that the fraction of time the operator is occupied is not enough to describe the regulatory range of the promoter

Noise, distortion and dynamic range of single mode 1.3 \ub5m InGaAs vertical cavity surface emitting lasers for radio-over-fibre links

The analogue modulation characteristics, including second order harmonic and third order intermodulation distortion, relative intensity noise (RIN) and spurious free dynamic range (SFDR), of single mode, GaAs-based 1.28 μm vertical cavity surface emitting laser (VCSELs) with highly strained InGaAs quantum wells have been investigated. The VCSELs utilise an oxide aperture for current and optical confinement and an inverted surface relief (SR) for suppression of higher order transverse modes. The inverted SR structure also has the advantage of suppressing oxide modes that, otherwise, appear in VCSELs with a large detuning of the cavity resonance with respect to the gain peak, which is needed to extend the emission wavelength. RIN levels comparable with those of single mode VCSELs emitting at 850 nm are demonstrated, with values from 140 to 150 dB/Hz in the 2 5 GHz range. SFDR values of 100 and 95 dBHz2/3 are obtained at 2 and 5 GHz, respectively. These values are in the range of those required in radio-over-fibre links

Noise, distortion and dynamic range of single mode 1.3 \ub5m InGaAs vertical cavity surface emitting lasers for radio-over-fibre links

The analogue modulation characteristics, including second order harmonic and third order intermodulation distortion, relative intensity noise (RIN) and spurious free dynamic range (SFDR), of single mode, GaAs-based 1.28 μm vertical cavity surface emitting laser (VCSELs) with highly strained InGaAs quantum wells have been investigated. The VCSELs utilise an oxide aperture for current and optical confinement and an inverted surface relief (SR) for suppression of higher order transverse modes. The inverted SR structure also has the advantage of suppressing oxide modes that, otherwise, appear in VCSELs with a large detuning of the cavity resonance with respect to the gain peak, which is needed to extend the emission wavelength. RIN levels comparable with those of single mode VCSELs emitting at 850 nm are demonstrated, with values from 140 to 150 dB/Hz in the 2 5 GHz range. SFDR values of 100 and 95 dBHz2/3 are obtained at 2 and 5 GHz, respectively. These values are in the range of those required in radio-over-fibre links

N.B.: When citing this work, cite the original article. Original Publication:

Hypoxia mediates low cell-cycle activity and increases the proportion of long-term reconstituting hematopoietic stem cells during in vitro cultur

Single mode 1.3 μm InGaAs VCSELs for access network applications

GaAs-based VCSELs emitting near 1.3 μm are realized using highly strained InGaAs quantum wells and a large detuning of the cavity resonance with respect to the gain peak. The VCSELs have an oxide aperture for current and optical confinement and an inverted surface relief for suppression of higher-order transverse modes. The inverted surface relief structure also has the advantage of suppressing oxide modes that otherwise appear in VCSELs with a large detuning between the cavity resonance and the gain peak. Under large signal, digital modulation, clear and open eyes and error free transmission over 9 km of single mode fiber have been demonstrated at the OC-48 and 10 GbE bit rates up to 85\ub0C. Here we review these results and present results from a complementary study of the RF modulation characteristics, including second order harmonic and third order intermodulation distortion, relative intensity noise (RIN), and spurious free dynamic range (SFDR). RIN levels comparable to those of single mode VCSELs emitting at 850 nm are demonstrated, with values from -140 to -150 dB/Hz. SFDR values of 100 and 95 dBHz2/3 were obtained at 2 and 5 GHz, respectively, which is in the range of those required in radio-over-fiber systems

Single mode 1.3 μm InGaAs VCSELs for access network applications

GaAs-based VCSELs emitting near 1.3 μm are realized using highly strained InGaAs quantum wells and a large detuning of the cavity resonance with respect to the gain peak. The VCSELs have an oxide aperture for current and optical confinement and an inverted surface relief for suppression of higher-order transverse modes. The inverted surface relief structure also has the advantage of suppressing oxide modes that otherwise appear in VCSELs with a large detuning between the cavity resonance and the gain peak. Under large signal, digital modulation, clear and open eyes and error free transmission over 9 km of single mode fiber have been demonstrated at the OC-48 and 10 GbE bit rates up to 85\ub0C. Here we review these results and present results from a complementary study of the RF modulation characteristics, including second order harmonic and third order intermodulation distortion, relative intensity noise (RIN), and spurious free dynamic range (SFDR). RIN levels comparable to those of single mode VCSELs emitting at 850 nm are demonstrated, with values from -140 to -150 dB/Hz. SFDR values of 100 and 95 dBHz2/3 were obtained at 2 and 5 GHz, respectively, which is in the range of those required in radio-over-fiber systems