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GS: DNA assembly for synthetic biology: from parts to pathways and beyond
- Integr Biol (Camb
"... The assembly of DNA from small fragments into large constructs has seen significant recent development, becoming a pivotal technology in the ability to implement the vision of synthetic biology. As the cost of whole gene synthesis is decreasing, whole genome synthesis at the other end of the spectru ..."
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The assembly of DNA from small fragments into large constructs has seen significant recent development, becoming a pivotal technology in the ability to implement the vision of synthetic biology. As the cost of whole gene synthesis is decreasing, whole genome synthesis at the other end of the spectrum has expanded our horizons to the prospect of fully engineered synthetic cells. However, the recently proven ability to synthesise genome-scale DNA is at odds with our ability to rationally engineer biological devices, which lags significantly behind. Most work in synthetic biology takes place on an intermediate scale with the combinatorial construction of networks and metabolic pathways from registries of modular biopart components. Implementation for rapid prototyping of engineered biological circuits requires quick and reliable DNA assembly according to specific architectures. It is apparent that DNA assembly is now a limiting technology in advancing synthetic biology. Current techniques employ standardised restriction enzyme assembly protocols such as BioBrickst, BglBricks and Golden Gate methods. Alternatively, sequence-independent overlap techniques, such as In-Fusiont, SLIC and Gibson isothermal assembly are becoming popular for larger assemblies, and in vivo DNA assembly in yeast and bacillus appears adept for chromosome fabrication. It is important to consider how the use of different technologies may impact the outcome of a construction, since the assembly technique can direct the architecture and diversity of systems that can be made. This review provides a critical examination of recent DNA assembly strategies and considers how this important facilitating aspect of synthetic biology may proceed in the future.
A synthetic low-frequency mammalian oscillator
- Nucleic Acids Res
, 2010
"... Circadian clocks have long been known to be essential for the maintenance of physiological and behavioral processes in a variety of organisms ranging from plants to humans. Dysfunctions that subvert gene expression of oscillatory circadian-clock components may result in severe pathologies, including ..."
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Circadian clocks have long been known to be essential for the maintenance of physiological and behavioral processes in a variety of organisms ranging from plants to humans. Dysfunctions that subvert gene expression of oscillatory circadian-clock components may result in severe pathologies, including tumors and metabolic disorders. While the underlying molecular mechanisms and dynamics of complex gene behavior are not fully understood, synthetic approaches have provided substantial insight into the operation of complex control circuits, including that of oscillatory networks. Using iterative cycles of mathematical model-guided design and experimental analyses, we have developed a novel low-frequency mammalian oscillator. It incorporates intronically encoded siRNA-based silencing of the tetracycline-dependent transactivator to enable the autonomous and robust expression of a fluorescent transgene with periods of 26h, a circadian clock-like oscilla-tory behavior. Using fluorescence-based time-lapse microscopy of engineered CHO-K1 cells, we profiled expression dynamics of a destabilized yellow fluo-rescent protein variant in single cells and real time. The novel oscillator design may enable further insights into the system dynamics of natural periodic processes as well as into siRNA-mediated transcription silencing. It may foster advances in design, analysis and application of complex syn-thetic systems in future gene therapy initiatives.
Genetic programs constructed from layered logic gates in single cells
- Nature
, 2013
"... Genetic programs function to integrate environmental sensors, implement signal processing algorithms and control expression dynamics We designed the architecture of an AND gate according to two constraints The transcription factors and chaperones were gleaned from gene clusters encoding type III ..."
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Cited by 9 (0 self)
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Genetic programs function to integrate environmental sensors, implement signal processing algorithms and control expression dynamics We designed the architecture of an AND gate according to two constraints The transcription factors and chaperones were gleaned from gene clusters encoding type III secretion systems, which are found in many pathogenic bacteria 9
Collective sensing-capacity of bacteria populations
- in Proc. International Symposium on Information Theory
, 2012
"... Abstract—The design of biological networks using bacteria as the basic elements of the network is initially motivated by a phenomenon called quorum sensing. Through quorum sensing, each bacterium performs sensing the medium and communi-cating it to others via molecular communication. As a result, ba ..."
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Abstract—The design of biological networks using bacteria as the basic elements of the network is initially motivated by a phenomenon called quorum sensing. Through quorum sensing, each bacterium performs sensing the medium and communi-cating it to others via molecular communication. As a result, bacteria can orchestrate and act collectively and perform tasks impossible otherwise. In this paper, we consider a population of bacteria as a single node in a network. In our version of biological communication networks, such a node would communicate with one another via molecular signals. As a first step toward such networks, this paper focuses on the study of the transfer of information to the population (i.e., the node) by stimulating it with a concentration of special type of a molecules signal. These molecules trigger a chain of processes inside each bacteria that results in a final output in the form of light or fluorescence.
Data gathering in networks of bacteria colonies: Collective sensing and relaying using molecular communication
- in NetSciCom Workshop at 31th Annual IEEE Conference on Computer Communications (INFOCOM
, 2012
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Efficient Modeling and Simulation of Bacteria-based Nanonetworks with BNSim
"... Abstract—Bacteria-based networks are formed using native or engineered bacteria that communicate at nano-scale. This definition includes the micro-scale molecular transportation system which uses chemotactic bacteria for targeted cargo delivery, as well as genetic circuits for intercellular interact ..."
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Abstract—Bacteria-based networks are formed using native or engineered bacteria that communicate at nano-scale. This definition includes the micro-scale molecular transportation system which uses chemotactic bacteria for targeted cargo delivery, as well as genetic circuits for intercellular interactions like quorum sensing or light communication. To characterize the dynamics of bacterial networks accurately, we introduce BNSim, an opensource, parallel, stochastic, and multiscale modeling platform which integrates various simulation algorithms, together with genetic circuits and chemotactic pathway models in a complex 3D environment. Moreover, we show how this platform can be used to model synthetic bacterial consortia which implement a XOR function and aggregate nearby bacteria using light communication. Consequently, the results demonstrate how BNSim can predict various properties of realistic bacterial networks and provide guidance for their actual wet-lab implementations. Index Terms—Nanotechnology, bacteria-based nanonetworks, bacteria consortia, multiscale modeling, stochastic simulation, heterogeneous multicellular system, cyber-physical systems I.
Biology by design: From top to bottom and back
- Journal of Biomedicine and Biotechnology, Volume 2010, Article ID
, 2010
"... Synthetic biology is a nascent technical discipline that seeks to enable the design and construction of novel biological systems to meet pressing societal needs. However, engineering biology still requires much trial and error because we lack effective approaches for connecting basic "parts&qu ..."
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Synthetic biology is a nascent technical discipline that seeks to enable the design and construction of novel biological systems to meet pressing societal needs. However, engineering biology still requires much trial and error because we lack effective approaches for connecting basic "parts" into higher-order networks that behave as predicted. Developing strategies for improving the performance and sophistication of our designs is informed by two overarching perspectives: "bottom-up" and "top-down" considerations. Using this framework, we describe a conceptual model for developing novel biological systems that function and interact with existing biological components in a predictable fashion. We discuss this model in the context of three topical areas: biochemical transformations, cellular devices and therapeutics, and approaches that expand the chemistry of life. Ten years after the construction of synthetic biology's first devices, the drive to look beyond what does exist to what can exist is ushering in an era of biology by design.
System-Theoretic Analysis and Least-Squares Design of Microfluidic Channels for Flow-Induced Molecular Communication
"... where molecular transport is performed via flow, is utilized in microfluidic channels to enhance diffusion-based molecular communication. The incorporation of the microfluidic channel and the transport of molecules by flow, i.e., convection, require a rigorous analysis to develop an end-to-end conce ..."
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where molecular transport is performed via flow, is utilized in microfluidic channels to enhance diffusion-based molecular communication. The incorporation of the microfluidic channel and the transport of molecules by flow, i.e., convection, require a rigorous analysis to develop an end-to-end concentration propagation model and a design for microfluidic channels. To the best of our knowledge, this is the firstattempttoanalyzeconcentration propagation in microfluidic channels from FMC perspective and devise them specifically to enhance the FMC. In this paper, a system-theoretic analysis of molecular transport is presented first. The system-theoretic model incorporates the solution of flow velocity in microfluidic channels and yields an end-to-end transfer function for concentration propagation based on building blocks of microfluidic channels. Then, the design of microfluidic channels is performed based on the least-squares Finite Impulse Response (FIR) filtering to achieve the desired end-to-end transfer function in FMC. According to the desired pass and stop bands, the required length and aspect-ratio parameters of the microfluidic channels are obtained for FIR filtering. The transfer functions for FMC is elaborated via numerical results. Furthermore, two example designs of microfluidic channels are presented for least-squares FIR band-pass and band-stop filtering in FMC. Index Terms—Least-squares linear filtering, microfluidics, molecular communication, nano communications, systems theory. I.
