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• Rapid cell-free forward engineering of novel genetic ring oscillators  + (While complex dynamic biological networks … While complex dynamic biological networks control gene expression in all living organisms, the forward engineering of comparable synthetic networks remains challenging. The current paradigm of characterizing synthetic networks in cells results in lengthy design-build-test cycles, minimal data collection, and poor quantitative characterization. Cell-free systems are appealing alternative environments, but it remains questionable whether biological networks behave similarly in cell-free systems and in cells. We characterized in a cell-free system the 'repressilator,' a three-node synthetic oscillator. We then engineered novel three, four, and five-gene ring architectures, from characterization of circuit components to rapid analysis of complete networks. When implemented in cells, our novel 3-node networks produced population-wide synchronized oscillations and 95% of 5-node oscillator cells oscillated for up to 72 hours. Oscillation periods in cells matched the cell-free system results for all networks tested. An alternate forward engineering paradigm using cell-free systems can thus accurately capture cellular behavior.thus accurately capture cellular behavior.)
• Moving bounding boxes and incremental synthesis for dynamic obstacles  + (While the use of formal synthesis for robo … While the use of formal synthesis for robotics problems in which the environment may act adversarially provides for exactârather than probabilisticâcorrectness of controllers, such methods are impractical when the adversary can move freely in a large portion of the workspace. As is well-known, this is due to exponential growth in the state space with the addition of each new problem variable. Furthermore, such an approach is overly conservative because most configurations will not be reached in typical runs. Rather than entirely abandon the discrete game view, we propose a combined method that ensures exact satisfaction of a given specification, expressed in linear temporal logic, while providing a lower bound on robot-obstacle distance throughout execution. Our method avoids explicit encoding of the moving obstacle and thus substantially reduces the reactive synthesis problem size, while allowing other nondeterministic variables to still be included in the specification. Our approaches centers on modeling obstacle motion as changes in the presence of a virtual static obstacle, and performing incremental synthesis in response. The algorithm is tested in application to a planar surveillance task.application to a planar surveillance task.)
• Metabolic engineering of Pseudomonas putida for production of vanillylamine from lignin-derived substrates  + (Whole-cell bioconversion of technical lign … Whole-cell bioconversion of technical lignins using Pseudomonas putida strains overexpressing amine transaminases (ATAs) has the potential to become an eco-efficient route to produce phenolic amines. Here, a novel cell growth-based screening method to evaluate the in vivo activity of recombinant ATAs towards vanillylamine in P. putida KT2440 was developed. It allowed the identification of the native enzyme Pp-SpuC-II and ATA from Chromobacterium violaceum (Cv-ATA) as highly active towards vanillylamine in vivo. Overexpression of Pp-SpuC-II and Cv-ATA in the strain GN442ΔPP_2426, previously engineered for reduced vanillin assimilation, resulted in 94- and 92-fold increased specific transaminase activity, respectively. Whole-cell bioconversion of vanillin yielded 0.70 ± 0.20 mM and 0.92 ± 0.30 mM vanillylamine, for Pp-SpuC-II and Cv-ATA, respectively. Still, amine production was limited by a substantial re-assimilation of the product and formation of the by-products vanillic acid and vanillyl alcohol. Concomitant overexpression of Cv-ATA and alanine dehydrogenase from Bacillus subtilis increased the production of vanillylamine with ammonium as the only nitrogen source and a reduction in the amount of amine product re-assimilation. Identification and deletion of additional native genes encoding oxidoreductases acting on vanillin are crucial engineering targets for further improvement.gineering targets for further improvement.)
• Real-Time Optimal Trajectory Generation for Constrained Dynamical Systems  + (With the advent of powerful computing and … With the advent of powerful computing and efficient computational algorithms,</br>real-time solutions to constrained optimal control problems are nearing a reality. In</br>this thesis, we develop a computationally efficient Nonlinear Trajectory Generation</br>(NTG) algorithm and describe its software implementation to solve, in real-time,</br>nonlinear optimal trajectory generation problems for constrained systems. NTG is</br>a nonlinear trajectory generation software package that combines nonlinear control</br>theory, B-spline basis functions, and nonlinear programming. We compare NTG</br>with other numerical optimal control problem solution techniques, such as direct</br>collocation, shooting, adjoints, and differential inclusions.</br>We demonstrate the performance of NTG on the Caltech Ducted Fan testbed.</br>Aggressive, constrained optimal control problems are solved in real-time for hover-</br>to-hover, forward flight, and terrain avoidance test cases. Real-time trajectory</br>generation results are shown for both the two-degree of freedom and receding</br>horizon control designs. Further experimental demonstration is provided with the</br>station-keeping, reconfiguration, and deconfiguration of micro-satellite formation</br>with complex nonlinear constraints. Successful application of NTG in these cases</br>demonstrates reliable real-time trajectory generation, even for highly nonlinear</br>and non-convex systems. The results are among the first to apply receding horizon</br>control techniques for agile flight in an experimental setting, using representative dynamics and computation.g representative dynamics and computation.)
• Quantifying Resource Competition and its Effects in the TX-TL System  + (Without accounting for the limited availab … Without accounting for the limited availability of shared cellular resources, the standard model of gene expression fails to reliably predict experimental data obtained in vitro. To overcome this limitation, we develop a dynamical model of gene expression explicitly modeling competition for scarce resources. In addition to accurately describing the experimental data, this model only depends on a handful of easily identifiable parame- ters with clear physical interpretation. Based on this model, we then characterize the combinations of protein concentrations that are simultaneously realizable with shared resources. As application examples, we demonstrate how the results can be used to explain similarities/differences among different in vitro extracts, furthermore, we illustrate that accounting for resource usage is essential in circuit design considering the toggle switch.cuit design considering the toggle switch.)

• Bacterial Controller Aided Wound Healing: A Case Study in Dynamical Population Controller Design  + (Wound healing is a complicated biological … Wound healing is a complicated biological process consisting of many types of cellular dynamics and functions regulated by chemical and molecular signals. Recent advances in synthetic biology have made it possible to predictably design and build closed-loop controllers that can function appropriately alongside biological species. In this paper we develop a simple dynamical population model mimicking the sequential relay-like dynamics of cellular populations involved in the wound healing process. Our model consists of four nodes and five signals whose parameters we can tune to simulate various chronic healing conditions. We also develop a set of regulator functions based on type-1 incoherent feed forward loops (IFFL) that can sense the change from acute healing to incomplete chronic wounds, improving the system in a timely manner. Both the wound healing and type-1 IFFL controller architectures are compatible with available synthetic biology experimental tools for potential applications.rimental tools for potential applications.)
• Aspects of Geometric Mechanics and Control of Mechanical Systems  + (any interesting control systems are mechan … any interesting control systems are mechanical control systems. In spite of</br>this, there has not been much effort to develop methods which use the special</br>structure of mechanical systems to obtain analysis tools which are</br>suitable for these systems. In this dissertation we take the first steps</br>towards a methodical treatment of mechanical control systems.</br><p></br>First we develop a framework for analysis of certain classes of</br>mechanical control systems. In the Lagrangian formulation we study ``simple</br>mechanical control systems'' whose Lagrangian is ``kinetic energy minus</br>potential energy.'' We propose a new and useful definition of</br>controllability for these systems and obtain a computable set of conditions</br>for this new version of controllability. We also obtain decompositions of</br>simple mechanical systems in the case when they are not controllable. In the</br>Hamiltonian formulation we study systems whose control vector fields are</br>Hamiltonian. We obtain decompositions which describe the controllable and</br>uncontrollable dynamics. In each case, the dynamics are shown to be</br>Hamiltonian in a suitably general sense.</br><p></br>Next we develop intrinsic descriptions of Lagrangian and Hamiltonian</br>mechanics in the presence of external inputs. This development is a first</br>step towards a control theory for general Lagrangian and Hamiltonian</br>control systems. Systems with constraints are also studied. We first give a</br>thorough overview of variational methods including a comparison of the</br>``nonholonomic'' and ``vakonomic'' methods. We also give a generalised</br>definition for a constraint and, with this more general definition, we are</br>able to give some preliminary controllability results for constrained systems.inary controllability results for constrained systems.)
• Specification and Synthesis of Reactive Protocols for Aircraft Electric Power Distribution  + (he increasing complexity of electric power … he increasing complexity of electric power systems leads to challenges in integration and verification. We consider the problem of designing a control protocol for an aircraft electric power system that meets a set of requirements describing the correct behaviors of the system and reacts dynamically to changes in internal system states. We formalize the requirements by translating them into a temporal logic specification language and apply game-based, temporal logic formal methods to automatically synthesize a controller protocol that satisfies these overall properties and requirements. Through a case study, we perform a design exploration to show the benefits and tradeoffs between centralized and distributed control architectures.zed and distributed control architectures.)
• Efficient control synthesis for augmented finite transition systems with an application to switching protocols  + (ugmented finite transition systems general … ugmented finite transition systems generalize nondeterministic transition systems with additional liveness conditions. We propose efficient algorithms for synthesizing control protocols for augmented finite transition systems to satisfy high-level specifications expressed in a fragment of linear temporal logic (LTL). We then use these algorithms within a framework for switching protocol synthesis for discrete-time dynamical systems, where augmented finite transition systems are used for abstracting the underlying dynamics. We introduce a notion of minimality for abstractions of certain fidelity and show that such minimal abstractions can be exactly computed for switched affine systems. Additionally, based on this framework, we present a procedure for computing digitally implementable switching protocols for continuous-time systems. The effectiveness of the proposed framework is illustrated through two examples of temperature control for buildings.ples of temperature control for buildings.)
• Designing Robustness to Temperature in a Feedforward Loop Circuit  + (âIncoherent feedforward loopsâ represent i … âIncoherent feedforward loopsâ represent important biomolecular circuit elements capable of a rich set of dynamic behavior including adaptation and pulsed responses. Temperature can modulate some of these properties through its effect on the underlying reaction rate parameters. It is generally unclear how to design a circuit where these properties are robust to variations in temperature. Here, we address this issue using a combination of tools from control and dynamical systems theory as well as preliminary experimental measurements towards such a design. Using a structured uncertainty representation, we analyze a standard incoherent feedforward loop circuit, noting mechanisms that intrinsically confer temperature robustness to some of its properties. Further, we study design variants that can enhance this robustness to temperature, including different negative feedback configurations as well as conditions for perfect temperature compensation. Finally, we find that the response of an incoherent feedforward loop circuit in cells can change with temperature. These results present groundwork for the design of a temperature-robust incoherent feedforward loop circuit.obust incoherent feedforward loop circuit.)