Layering genetic circuits to build a single cell, bacterial half adder
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 compil...
| Main Authors: | , , , |
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| Other Authors: | |
| Format: | Journal Article |
| Language: | English |
| Published: |
2015
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| Online Access: | https://hdl.handle.net/10356/80944 http://hdl.handle.net/10220/38986 |
| _version_ | 1811685061104238592 |
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| author | Wong, Adison Wang, Huijuan Poh, Chueh Loo Kitney, Richard I. |
| author2 | School of Chemical and Biomedical Engineering |
| author_facet | School of Chemical and Biomedical Engineering Wong, Adison Wang, Huijuan Poh, Chueh Loo Kitney, Richard I. |
| author_sort | Wong, Adison |
| collection | NTU |
| description | 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 σ54 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. |
| first_indexed | 2024-10-01T04:38:32Z |
| format | Journal Article |
| id | ntu-10356/80944 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T04:38:32Z |
| publishDate | 2015 |
| record_format | dspace |
| spelling | ntu-10356/809442023-12-29T06:48:06Z Layering genetic circuits to build a single cell, bacterial half adder Wong, Adison Wang, Huijuan Poh, Chueh Loo Kitney, Richard I. School of Chemical and Biomedical Engineering 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 σ54 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. Published version 2015-12-07T09:16:46Z 2019-12-06T14:17:58Z 2015-12-07T09:16:46Z 2019-12-06T14:17:58Z 2015 Journal Article 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(40). 1741-7007 https://hdl.handle.net/10356/80944 http://hdl.handle.net/10220/38986 10.1186/s12915-015-0146-0 26078033 en BMC Biology © 2015 Wong et al. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. 16 p. application/pdf |
| spellingShingle | Wong, Adison Wang, Huijuan Poh, Chueh Loo Kitney, Richard I. Layering genetic circuits to build a single cell, bacterial half adder |
| title | Layering genetic circuits to build a single cell, bacterial half adder |
| title_full | Layering genetic circuits to build a single cell, bacterial half adder |
| title_fullStr | Layering genetic circuits to build a single cell, bacterial half adder |
| title_full_unstemmed | Layering genetic circuits to build a single cell, bacterial half adder |
| title_short | Layering genetic circuits to build a single cell, bacterial half adder |
| title_sort | layering genetic circuits to build a single cell bacterial half adder |
| url | https://hdl.handle.net/10356/80944 http://hdl.handle.net/10220/38986 |
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