Journal
NUCLEIC ACIDS RESEARCH
Volume 41, Issue 5, Pages 3471-3481Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkt052
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Funding
- Medical Research Council (MRC)
- Engineering and Physical Sciences Research Council (EPSRC)
- Engineering and Physical Sciences Research Council [EP/G036004/1]
- Center for Synthetic Biology and Innovation at Imperial College London
- EPSRC [EP/G036004/1] Funding Source: UKRI
- Engineering and Physical Sciences Research Council [EP/G036004/1] Funding Source: researchfish
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A bottleneck in our capacity to rationally and predictably engineer biological systems is the limited number of well-characterized genetic elements from which to build. Current characterization methods are tied to measurements in living systems, the transformation and culturing of which are inherently time-consuming. To address this, we have validated a completely in vitro approach for the characterization of DNA regulatory elements using Escherichia coli extract cell-free systems. Importantly, we demonstrate that characterization in cell-free systems correlates and is reflective of performance in vivo for the most frequently used DNA regulatory elements. Moreover, we devise a rapid and completely in vitro method to generate DNA templates for cell-free systems, bypassing the need for DNA template generation and amplification from living cells. This in vitro approach is significantly quicker than current characterization methods and is amenable to high-throughput techniques, providing a valuable tool for rapidly prototyping libraries of DNA regulatory elements for synthetic biology.
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