Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology (original) (raw)
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The Design and Construction of a Set of Modular Synthetic BioLogic Devices for Programming Cells
IFMBE Proceedings, 2009
Modularity is an essential property for rationally engineered standard parts and devices. This principle is now being extended to biological based parts and devices for programming cells. However, the design principles and building blocks which are currently in Synthetic Biology are somewhat limited. In addition, it is important to explore the underlying mechanisms of existing, natural biological systems in order to utilise them in designing novel genetic circuit modules. In this paper, we will describe a set of modular synthetic biological parts and devices that are based in rational design. Particularly, a modular tight-controlled and hypersensitive genetic circuit with digital logic AND function is rationally designed and engineered. They use a sigma factor 54 dependent heteroregulation module in the hrp (hypersensitive response and pathogenicity) gene regulatory system for Type III secretion in Pseudomonas syringae. Their inputs and outputs are both promoters and thus do not rely on specific inducible promoters and could drive various cellular responses. It shows that the hrp system has significant potential for building a range of biological parts and devices with good performance and flexibility.
Foundations for the design and implementation of synthetic genetic circuits
Nature Reviews Genetics, 2012
Synthetic gene circuits are designed to program new biological behaviour, dynamics and logic control. For all but the simplest synthetic phenotypes, this requires a structured approach to map the desired functionality to available molecular and cellular parts and processes. In other engineering disciplines, a formalized design process has greatly enhanced the scope and rate of success of projects. When engineering biological systems, a desired function must be achieved in a context that is incompletely known, is influenced by stochastic fluctuations and is capable of rich nonlinear interactions with the engineered circuitry. Here, we review progress in the provision and engineering of libraries of parts and devices, their composition into large systems and the emergence of a formal design process for synthetic biology.
Construction of integrated gene logic-chip
Nature nanotechnology, 2018
In synthetic biology, the control of gene expression requires a multistep processing of biological signals. The key steps are sensing the environment, computing information and outputting products. To achieve such functions, the laborious, combinational networking of enzymes and substrate-genes is required, and to resolve problems, sophisticated design automation tools have been introduced. However, the complexity of genetic circuits remains low because it is difficult to completely avoid crosstalk between the circuits. Here, we have made an orthogonal self-contained device by integrating an actuator and sensors onto a DNA origami-based nanochip that contains an enzyme, T7 RNA polymerase (RNAP) and multiple target-gene substrates. This gene nanochip orthogonally transcribes its own genes, and the nano-layout ability of DNA origami allows us to rationally design gene expression levels by controlling the intermolecular distances between the enzyme and the target genes. We further inte...
Design principles of transcriptional logic circuits
Using a set of genetic logic gates (AND, OR and XOR), we constructed a binary full-adder. The optimality analysis of the full-adder showed that, based on the position of the regulation threshold, the system displays different optimal configurations for speed and accuracy under fixed metabolic cost. In addition, the analysis identified an optimal trade-off curve bounded by these two optimal configurations. Any configuration outside this optimal trade-off curve is sub-optimal in both speed and accuracy. This type of analysis represents a useful tool for synthetic biologists to engineer faster, more accurate and cheaper genes.
Modular, Multi-Input Transcriptional Logic Gating with Orthogonal LacI/GalR Family Chimeras
ACS Synthetic Biology, 2014
In prokaryotes, the construction of synthetic, multi-input promoters is constrained by the number of transcription factors that can simultaneously regulate a single promoter. This fundamental engineering constraint is an obstacle to synthetic biologists because it limits the computational capacity of engineered gene circuits. Here, we demonstrate that complex multi-input transcriptional logic gating can be achieved through the use of ligand-inducible chimeric transcription factors assembled from the LacI/GalR family. These modular chimeras each contain a ligand-binding domain and a DNA-binding domain, both of which are chosen from a library of possibilities. When two or more chimeras have the same DNA-binding domain, they independently and simultaneously regulate any promoter containing the appropriate operator site. In this manner, simple transcriptional AND gating is possible through the combination of two chimeras, and multiple-input AND gating is possible with the simultaneous use of three or even four chimeras. Furthermore, we demonstrate that orthogonal DNA-binding domains and their cognate operators allow the coexpression of multiple, orthogonal AND gates. Altogether, this work provides synthetic biologists with novel, ligand-inducible logic gates and greatly expands the possibilities for engineering complex synthetic gene circuits.
2011
Building biological devices to perform computational and signal processing tasks is one of the main research issues in synthetic biology. Herein, two modular biological systems that could mimic multiplexing and demultiplexing logic functions are proposed and discussed. These devices, called multiplexer (mux) and demultiplexer (demux), respectively, have a remarkable importance in electronic, telecommunication, and signal processing systems and, similarly, they could play a crucial role if implemented in a living organism, such as Escherichia coli. BioBrick standard parts were used to design mux and demux and to construct two genetic circuits that could carry out the desired tasks. A modular approach, mimicking basic logic gates (AND, OR, and NOT) with protein/autoinducer or protein/DNA interactions and interconnecting them to create the final circuits, was adopted. A mathematical model of the designed gene networks was defined and simulations performed to validate the expected behavior of the systems. In addition, circuit subparts were tested in vivo and the results used to determine some of the parameters of the mathematical model. According to both the experimental and simulated results, guidelines for future finalization of mux and demux are provided.
Engineering Gene Circuits: Foundations and Applications
Nanotechnology in Biology and Medicine, 2007
Synthetic biology has emerged as a useful approach to decoding fundamental laws underlying biological control. Recent efforts have produced many exciting systems and generated substantial insights. These progresses highlight the potential of synthetic biology to impact diverse areas including biology, computation, engineering, and medicine. 20.1 Introduction Biological systems often function reliably in diverse environments despite internal or external perturbations. This behavior is often characterized as ''robustness.'' Based on extensive studies over the last several decades, much of this robustness can be attributed to the control of gene expression through complex cellular networks [1-4]. These networks are known to consist of various regulatory modules, including feedback [5] and feed-forward [6] regulation and cell-cell communication [7]. With these basic regulatory modules and motifs, researchers are now constructing artificial networks that mimic nature to gain fundamental biological insight and understanding [8]. In addition, other artificial networks that are engineered with novel functions will serve as building blocks for future practical applications. These efforts form the foundation of the recent emergence of synthetic biology [3,9,10]. These artificial networks are interchangeably called ''synthetic gene circuits'' or ''engineered gene circuits.'' Recent accomplishments in synthetic biology include engineered switches [11-14], oscillators [15,16], logic gates [17-19], metabolic control [20], reengineered translational machinery [21], population control [22] and pattern formation [23] using natural or synthetic [24] cell-cell communication, reengineered viral genome [25], and hierarchically complex circuits built upon smaller, well-characterized
Engineering in the biological substrate: information processing in genetic circuits
Proceedings of the IEEE, 2000
We review the rapidly evolving efforts to analyze, model, simulate, and engineer genetic and biochemical information processing systems within living cells. We begin by showing that the fundamental elements of information processing in electronic and genetic systems are strikingly similar, and follow this theme through a review of efforts to create synthetic genetic circuits. In particular, we describe and review the "silicon mimetic" approach, where genetic circuits are engineered to mimic the functionality of semiconductor devices such as logic gates, latched circuits, and oscillators. This is followed with a review of the analysis, modeling, and simulation of natural and synthetic genetic circuits, which often proceed in a manner similar to that used for electronic systems. We conclude by presenting examples of naturally occurring genetic and biochemical systems that recently have been conceptualized in terms familiar to systems engineers. Our review of these newly forming fields of research demonstrates that the expertise and skills contained within electrical and computer engineering disciplines apply not only to design within biological systems, but also to the development of a deeper understanding of biological functionality. This review of these efforts points to the emergence of both engineering and basic science disciplines following parallel paths.
Methods in …, 2007
Synthetic biology is an emerging field in which the procedures and methods of engineering are extended to living organisms, with the long-term goal of producing novel cell types that aid human society. For example, engineered cell types may sense a particular environment and express gene products that serve as an indicator of that environment, or effect a change in that environment. While we are still some way from producing cells with significant practical applications, the immediate goals of synthetic biology are to develop a quantitative understanding of genetic circuitry and its interactions with the environment and to develop modular genetic circuitry derived from standard, interoperable, parts, that can be introduced into cells and results in some desired input/output function. Using an engineering approach, the input/output function of each modular element is characterized independently, providing a toolkit of elements that can be linked in different ways to provide various circuit topologies. The principle of modularity, yet largely unproven for biological systems, suggests that modules will function appropriately based on their design characteristics when combined into larger synthetic genetic devices. This modularity concept is similar to that used to develop large computer programs, where inpendent software modules can be independently developed and later combined into the final program.