Abstract
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.
Original language | English |
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Pages (from-to) | 5892-5901 |
Number of pages | 13 |
Journal | Proceedings of the National Academy of Sciences (PNAS) |
Volume | 116 |
Issue number | 13 |
Early online date | 8 Mar 2019 |
DOIs | |
Publication status | Published - 26 Mar 2019 |
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Nadanai Laohakunakorn
- School of Biological Sciences - Chancellor's Fellow - Biotechnology
- Centre for Engineering Biology
Person: Academic: Research Active (Teaching)