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Title: Layering genetic circuits to build a single cell, bacterial half adder
Authors: Wong, A
Wang, H
Poh, C.L 
Kitney, R.I
Keywords: Bacteria (microorganisms)
5' untranslated region
5' untranslated region
Escherichia coli
gene regulatory network
molecular computer
promoter region
synthetic biology
5' Untranslated Regions
Computers, Molecular
Escherichia coli
Gene Regulatory Networks
Promoter Regions, Genetic
Synthetic Biology
Issue Date: 2015
Citation: Wong, A, Wang, H, Poh, C.L, Kitney, R.I (2015). Layering genetic circuits to build a single cell, bacterial half adder. BMC Biology 13 (1) : 40. ScholarBank@NUS Repository.
Abstract: Background: Gene regulation in biological systems is impacted by the cellular and genetic context-dependent effects of the biological parts which comprise the circuit. Here, we have sought to elucidate the limitations of engineering biology from an architectural point of view, with the aim of compiling a set of engineering solutions for overcoming failure modes during the development of complex, synthetic genetic circuits. Results: Using a synthetic biology approach that is supported by computational modelling and rigorous characterisation, AND, OR and NOT biological logic gates were layered in both parallel and serial arrangements to generate a repertoire of Boolean operations that include NIMPLY, XOR, half adder and half subtractor logics in a single cell. Subsequent evaluation of these near-digital biological systems revealed critical design pitfalls that triggered genetic context-dependent effects, including 5' UTR interferences and uncontrolled switch-on behaviour of the supercoiled ?54 promoter. In particular, the presence of seven consecutive hairpins immediately downstream of the promoter transcription start site severely impeded gene expression. Conclusions: As synthetic biology moves forward with greater focus on scaling the complexity of engineered genetic circuits, studies which thoroughly evaluate failure modes and engineering solutions will serve as important references for future design and development of synthetic biological systems. This work describes a representative case study for the debugging of genetic context-dependent effects through principles elucidated herein, thereby providing a rational design framework to integrate multiple genetic circuits in a single prokaryotic cell. © 2015 Wong et al.
Source Title: BMC Biology
ISSN: 17417007
DOI: 10.1186/s12915-015-0146-0
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