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Synthetic Biology underpins advances in the bioeconomy
Biological systems - including the simplest cells - exhibit a broad range of functions to thrive in their environment. Research in the Imperial College Centre for Synthetic Biology is focused on the possibility of engineering the underlying biochemical processes to solve many of the challenges facing society, from healthcare to sustainable energy. In particular, we model, analyse, design and build biological and biochemical systems in living cells and/or in cell extracts, both exploring and enhancing the engineering potential of biology.
As part of our research we develop novel methods to accelerate the celebrated Design-Build-Test-Learn synthetic biology cycle. As such research in the Centre for Synthetic Biology highly multi- and interdisciplinary covering computational modelling and machine learning approaches; automated platform development and genetic circuit engineering ; multi-cellular and multi-organismal interactions, including gene drive and genome engineering; metabolic engineering; in vitro/cell-free synthetic biology; engineered phages and directed evolution; and biomimetics, biomaterials and biological engineering.
@article{Broedel:2017:10.1038/nprot.2017.084,
author = {Broedel, AK and Jaramillo, A and Isalan, M},
doi = {10.1038/nprot.2017.084},
journal = {Nature Protocols},
pages = {1830--1843},
title = {Intracellular directed evolution of proteins from combinatorial libraries based on conditional phage replication},
url = {http://dx.doi.org/10.1038/nprot.2017.084},
volume = {12},
year = {2017}
}
TY - JOUR
AB - Directed evolution is a powerful tool to improve the characteristics of biomolecules. Here we present a protocol for the intracellular evolution of proteins with distinct differences and advantages in comparison with established techniques. These include the ability to select for a particular function from a library of protein variants inside cells, minimizing undesired coevolution and propagation of nonfunctional library members, as well as allowing positive and negative selection logics using basally active promoters. A typical evolution experiment comprises the following stages: (i) preparation of a combinatorial M13 phagemid (PM) library expressing variants of the gene of interest (GOI) and preparation of the Escherichia coli host cells; (ii) multiple rounds of an intracellular selection process toward a desired activity; and (iii) the characterization of the evolved target proteins. The system has been developed for the selection of new orthogonal transcription factors (TFs) but is capable of evolving any gene—or gene circuit function—that can be linked to conditional M13 phage replication. Here we demonstrate our approach using as an example the directed evolution of the bacteriophage λ cI TF against two synthetic bidirectional promoters. The evolved TF variants enable simultaneous activation and repression against their engineered promoters and do not cross-react with the wild-type promoter, thus ensuring orthogonality. This protocol requires no special equipment, allowing synthetic biologists and general users to evolve improved biomolecules within ~7 weeks.
AU - Broedel,AK
AU - Jaramillo,A
AU - Isalan,M
DO - 10.1038/nprot.2017.084
EP - 1843
PY - 2017///
SN - 1750-2799
SP - 1830
TI - Intracellular directed evolution of proteins from combinatorial libraries based on conditional phage replication
T2 - Nature Protocols
UR - http://dx.doi.org/10.1038/nprot.2017.084
UR - http://hdl.handle.net/10044/1/50850
VL - 12
ER -