Biology subjects

Swank, Z.

Publications and source records attributed to Swank, Z..

4 recordsLinked to original sources

bioRxiv2023

Heterotypic responses against nsp12/nsp13 from prior SARS-CoV-2 infection associates with lower subsequent endemic coronavirus incidence

Immune responses from prior SARS-CoV-2 infection and COVID-19 vaccination do not prevent re-infections and may not protect against future novel coronaviruses (CoVs). We examined the incidence of and immune differences against human endemic CoVs (eCoV) as a proxy for response against future emerging CoVs. Assessment was among those with known SARS-CoV-2 infection, COVID-19 vaccination but no documented SARS-CoV-2 infection, or neither exposure. Retrospective cohort analyses suggest that prior SARS-CoV-2 infection, but not COVID-19 vaccination alone, protects against subsequent symptomatic eCoV infection. CD8+ T cell responses to the non-structural eCoV proteins, nsp12 and nsp13, were significantly higher in individuals with previous SARS-CoV-2 infection as compared to the other groups. The three groups had similar cellular responses against the eCoV spike and nucleocapsid, and those with prior spike exposure had lower eCoV-directed neutralizing antibodies. Incorporation of non-structural viral antigens in a future pan-CoV vaccine may improve protection against future heterologous CoV infections.

immunology↗

bioRxiv2020

CFPU: a cell-free processing unit for high-throughput, automated in vitro circuit characterization

Forward engineering synthetic circuits is at the core of synthetic biology. Automated solutions will be required to facilitate circuit design and implementation. Circuit design is increasingly being automated with design software, but innovations in experimental automation are lagging behind. Microfluidic technologies made it possible to perform in vitro transcription-translation (tx-tl) reactions with increasing throughput and sophistication, enabling screening and characterization of individual circuit elements and complete circuit designs. Here we developed an automated microfluidic cell-free processing unit (CFPU) that extends high-throughput screening capabilities to a continuous reaction environment, which is essential for the implementation and analysis of more complex and dynamic circuits. The CFPU contains 280 chemostats that can be individually programmed with DNA circuits. Each chemostat is periodically supplied with tx-tl reagents, giving rise to sustained, long-term steady state conditions. Using microfluidic pulse width modulation (PWM) the device is able to generate tx-tl reagent compositions in real-time. The device has higher throughput, lower reagent consumption, and overall higher functionality than current chemostat devices. We applied this technology to map transcription factor based repression under equilibrium conditions and implemented dynamic gene circuits switchable by small molecules. We expect the CFPU to help bridge the gap between circuit design and experimental automation for in vitro development of synthetic gene circuits.

synthetic biology↗

bioRxiv2018

Cell-free gene regulatory network engineering with synthetic transcription factors

Gene regulatory networks are ubiquitous in nature and critical for bottom-up engineering of synthetic networks. Transcriptional repression is a fundamental function that can be tuned at the level of DNA, protein, and cooperative protein - protein interactions, necessitating high-throughput experimental approaches for in-depth characterization. Here we used a cell-free system in combination with a high-throughput microfluidic device to comprehensively study the different tuning mechanisms of a synthetic zinc-finger repressor library, whose affinity and cooperativity can be rationally engineered. The device is integrated into a comprehensive workflow that includes determination of transcription factor binding energy landscapes and mechanistic modeling, enabling us to generate a library of well-characterized synthetic transcription factors and corresponding promoters, which we then used to build gene regulatory networks de novo. The well-characterized synthetic parts and insights gained should be useful for rationally engineering gene regulatory networks and for studying the biophysics of transcriptional regulation.

synthetic biology↗

bioRxiv2018

Microfluidic device for on-chip mixing and encapsulation of lysates

Emulsion drops are often employed as picoliter-sized containers to perform screening assays. These assays usually entail the formation of drops encompassing discrete objects such as cells or microparticles and reagents to study interactions between the different encapsulants. Drops are also used to screen influences of reagent concentrations on the final product. However, these latter assays are less frequently performed because it is difficult to change the reagent concentration over a wide range with high precision within a single experiment. In this paper, we present a microfluidic double emulsion drop maker containing pneumatic valves that enable injection of different reagents using pulsed width modulation and subsequent mixing. This device can produce drops from reagent volumes as low as 10 l with minimal sample loss, thereby enabling experiments that would be prohibitively expensive using droplet generators that do not contain valves. We employ this device to monitor the kinetics of cell free synthesis of green fluorescent proteins inside double emulsions. To demonstrate the potential of this device, we perform DNA titration experiments in double emulsion drops to test the influence of the DNA concentration on the amount of green fluorescence proteins produced.

synthetic biology↗