Cell-Free Gene Expression from DNA Brushes

Lévy, Michael; Vonshak, Ohad; Divon, Yiftach; Greiss, Ferdinand; Avidan, Noa; Daube, Shirley S.; Bar‐Ziv, Roy

doi:10.1007/978-1-0716-1998-8_8

Cell-Free Gene Expression

2022

DOI: 10.1007/978-1-0716-1998-8_8

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Cell-Free Gene Expression from DNA Brushes

Michael Lévy

Ohad Vonshak

Yiftach Divon

et al.

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2024

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Cited by 1 publication

References 22 publications

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An Automated Cell-Free Workflow for Transcription Factor Engineering

Ekas,

Wang,

Silverman

et al. 2024

ACS Synth. Biol.

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The design and optimization of metabolic pathways, genetic systems, and engineered proteins rely on highthroughput assays to streamline design-build-test-learn cycles. However, assay development is a time-consuming and laborious process. Here, we create a generalizable approach for the tailored optimization of automated cell-free gene expression (CFE)-based workflows, which offers distinct advantages over in vivo assays in reaction flexibility, control, and time to data. Centered around designing highly accurate and precise transfers on the Echo Acoustic Liquid Handler, we introduce pilot assays and validation strategies for each stage of protocol development. We then demonstrate the efficacy of our platform by engineering transcription factor-based biosensors. As a model, we rapidly generate and assay libraries of 127 MerR and 134 CadR transcription factor variants in 3682 unique CFE reactions in less than 48 h to improve limit of detection, selectivity, and dynamic range for mercury and cadmium detection. This was achieved by assessing a panel of ligand conditions for sensitivity (to 0.1, 1, 10 μM Hg and 0, 1, 10, 100 μM Cd for MerR and CadR, respectively) and selectivity (against Ag, As, Cd, Co, Cu, Hg, Ni, Pb, and Zn). We anticipate that our Echo-based, cell-free approach can be used to accelerate multiple design workflows in synthetic biology.

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An Automated Cell-Free Workflow for Transcription Factor Engineering

Ekas,

Wang,

Silverman

et al. 2024

ACS Synth. Biol.

View full text Add to dashboard Cite

show abstract

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Cell-Free Gene Expression from DNA Brushes

Cited by 1 publication

References 22 publications

An Automated Cell-Free Workflow for Transcription Factor Engineering

An Automated Cell-Free Workflow for Transcription Factor Engineering

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