Darlington, Alexander P. S. (2018) Design of genetic controllers to manage translational resource allocation in synthetic gene networks. PhD thesis, University of Warwick.

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Abstract

To control cellular processes synthetic biologists and biotechnologists often use regulation of gene expression; by regulating transcription it is assumed protein levels will follow. However the use of a common pool of gene expression resources results in the emergence of hidden interactions, couplings, between genes which are not immediately apparent from circuit topologies. This can result in a breakdown in the relationship between transcriptional regulation (input) and protein levels (output). Current evidence suggests that it is the number of free ribosomes which limits protein synthesis capacity and therefore creates these non-regulatory linkages.

In this thesis, the feasibility of dividing the cell's translational resources to reverse the breakdown in input-output responses and decouple co-expressed genes is demonstrated. A model of microbial growth which captures key aspects of host-circuit interactions is developed and demonstrates the feasibility of using orthogonal (circuit specific) ribosomes for circuit gene expression; showing that by allocating circuit genes to both host and orthogonal translational pools these genes can be decoupled and the flux through a model biotechnological pathway can be improved.

The design of negative feedback controllers to allocate translational resources is investigated. These act to increase supply of orthogonal ribosomes as the demand for translational resources by the circuit increases. The stability of a number of controller architectures is investigated. An experimental prototype of the best performing controller architecture is able to reduce gene coupling by 50%.

The best controller architecture is carried forward for further analysis, and a detailed mechanistic model which can be used as a design tool is developed. A hard trade of between gene expression and decoupling activity is identified and designs which manage this trade-o suggested. The controller is shown to ameliorate resource mediated failures of modularity in a range of synthetic biology circuits.

Finally, a discussion on ways to produce a second generation of translational resource allocation controllers is provided.

Item Type:	Thesis (PhD)
Subjects:	Q Science > QH Natural history > QH426 Genetics
Library of Congress Subject Headings (LCSH):	Cellular control mechanisms, Gene regulatory networks, Ribosomes, Gene expression
Official Date:	February 2018
Dates:	Date Event February 2018 UNSPECIFIED
Institution:	University of Warwick
Theses Department:	School of Engineering
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Bates, Declan
Sponsors:	Engineering and Physical Sciences Research Council ; Biotechnology and Biological Sciences Research Council (Great Britain)
Extent:	x, 149 leaves : illustrations, charts
Language:	eng

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