Modeling and simulation of a synthetic genetic circuit that implements a multicelled behavior in a growing microcolony of E. coli

Synthetic Biology is a relatively new discipline, born at the beginning of the New Millennium, that brings the typical engineering approach (abstraction, modularity and standardization) to biotechnology. These principles aim to tame the extreme complexity of the various components and aid the construction of artificial biological systems with specific functions, usually by means of synthetic genetic circuits implemented in bacteria or simple eukaryotes like yeast. The cell becomes a programmable machine and its low-level programming language is made of strings of DNA. This work was performed in collaboration with researchers of the Department of Electrical Engineering of the University of Washington in Seattle and also with a student of the Corso di Laurea Magistrale in Ingegneria Biomedica at the University of Bologna: Marilisa Cortesi. During the collaboration I contributed to a Synthetic Biology project already started in the Klavins Laboratory. In particular, I modeled and subsequently simulated a synthetic genetic circuit that was ideated for the implementation of a multicelled behavior in a growing bacterial microcolony. In the first chapter the foundations of molecular biology are introduced: structure of the nucleic acids, transcription, translation and methods to regulate gene expression. An introduction to Synthetic Biology completes the section. In the second chapter is described the synthetic genetic circuit that was conceived to make spontaneously emerge, from an isogenic microcolony of bacteria, two different groups of cells, termed leaders and followers. The circuit exploits the intrinsic stochasticity of gene expression and intercellular communication via small molecules to break the symmetry in the phenotype of the microcolony. The four modules of the circuit (coin flipper, sender, receiver and follower) and their interactions are then illustrated. In the third chapter is derived the mathematical representation of the various components of the circuit and the several simplifying assumptions are made explicit. Transcription and translation are modeled as a single step and gene expression is function of the intracellular concentration of the various transcription factors that act on the different promoters of the circuit. A list of the various parameters and a justification for their value closes the chapter. In the fourth chapter are described the main characteristics of the gro simulation environment, developed by the Self Organizing Systems Laboratory of the University of Washington. Then, a sensitivity analysis performed to pinpoint the desirable characteristics of the various genetic components is detailed. The sensitivity analysis makes use of a cost function that is based on the fraction of cells in each one of the different possible states at the end of the simulation and the wanted outcome. Thanks to a particular kind of scatter plot, the parameters are ranked. Starting from an initial condition in which all the parameters assume their nominal value, the ranking suggest which parameter to tune in order to reach the goal. Obtaining a microcolony in which almost all the cells are in the follower state and only a few in the leader state seems to be the most difficult task. A small number of leader cells struggle to produce enough signal to turn the rest of the microcolony in the follower state. It is possible to obtain a microcolony in which the majority of cells are followers by increasing as much as possible the production of signal. Reaching the goal of a microcolony that is split in half between leaders and followers is comparatively easy. The best strategy seems to be increasing slightly the production of the enzyme. To end up with a majority of leaders, instead, it is advisable to increase the basal expression of the coin flipper module. At the end of the chapter, a possible future application of the leader election circuit, the spontaneous formation of spatial patterns in a microcolony, is modeled with the finite state machine formalism. The gro simulations provide insights into the genetic components that are needed to implement the behavior. In particular, since both the examples of pattern formation rely on a local version of Leader Election, a short-range communication system is essential. Moreover, new synthetic components that allow to reliably downregulate the growth rate in specific cells without side effects need to be developed. In the appendix are listed the gro code utilized to simulate the model of the circuit, a script in the Python programming language that was used to split the simulations on a Linux cluster and the Matlab code developed to analyze the data.

Design of a GNSS receiver for the ESEO mission

This thesis was carried out inside the ESA's ESEO mission and focus in the design of one of the secondary payloads carried on board the spacecraft: a GNSS receiver for orbit determination. The purpose of this project is to test the technology of the orbit determination in real time applications by using commercial components. The architecture of the receiver includes a custom part, the navigation computer, and a commercial part, the front-end, from Novatel, with COCOM limitation removed, and a GNSS antenna. This choice is motivated by the goal of demonstrating the correct operations in orbit, enabling a widespread use of this technology while lowering the cost and time of the device’s assembly. The commercial front-end performs GNSS signal acquisition, tracking and data demodulation and provides raw GNSS data to the custom computer. This computer processes this raw observables, that will be both transferred to the On-Board Computer and then transmitted to Earth and provided as input to the recursive estimation filter on-board, in order to obtain an accurate positioning of the spacecraft, using the dynamic model. The main purpose of this thesis, is the detailed design and development of the mentioned GNSS receiver up to the ESEO project Critical Design Review, including requirements definition, hardware design and breadboard preliminary test phase design.

Analysis of robot dynamics through complexity measures: an application in automatic design

L'applicazione di misure, derivanti dalla teoria dell'informazione, fornisce un valido strumento per quantificare alcune delle proprietà dei sistemi complessi. Le stesse misure possono essere utilizzate in robotica per favorire l'analisi e la sintesi di sistemi di controllo per robot. In questa tesi si è analizzata la correlazione tra alcune misure di complessità e la capacità dei robot di portare a termine, con successo, tre differenti task. I risultati ottenuti suggeriscono che tali misure di complessità rappresentano uno strumento promettente anche nel campo della robotica, ma che il loro utilizzo può diventare difficoltoso quando applicate a task compositi.

Characterization of ENSO changes under idealised radiative forcing in an intermediate complexity general circulation model

Intermediate-complexity general circulation models are a fundamental tool to investigate the role of internal and external variability within the general circulation of the atmosphere and ocean. The model used in this thesis is an intermediate complexity atmospheric general circulation model (SPEEDY) coupled to a state-of-the-art modelling framework for the ocean (NEMO). We assess to which extent the model allows a realistic simulation of the most prominent natural mode of variability at interannual time scales: El-Niño Southern Oscillation (ENSO). To a good approximation, the model represents the ENSO-induced Sea Surface Temperature (SST) pattern in the equatorial Pacific, despite a cold tongue-like bias. The model underestimates (overestimates) the typical ENSO spatial variability during the winter (summer) seasons. The mid-latitude response to ENSO reveals that the typical poleward stationary Rossby wave train is reasonably well represented. The spectral decomposition of ENSO features a spectrum that lacks periodicity at high frequencies and is overly periodic at interannual timescales. We then implemented an idealised transient mean state change in the SPEEDY model. A warmer climate is simulated by an alteration of the parametrized radiative fluxes that corresponds to doubled carbon dioxide absorptivity. Results indicate that the globally averaged surface air temperature increases of 0.76 K. Regionally, the induced signal on the SST field features a significant warming over the central-western Pacific and an El-Niño-like warming in the subtropics. In general, the model features a weakening of the tropical Walker circulation and a poleward expansion of the local Hadley cell. This response is also detected in a poleward rearrangement of the tropical convective rainfall pattern. The model setting that has been here implemented provides a valid theoretical support for future studies on climate sensitivity and forced modes of variability under mean state changes.

Validation of an intermediate-complexity flood risk model for climate change adaptation

In questo lavoro di tesi viene presentato e validato un modello di rischio di alluvione a complessità intermedia per scenari climatici futuri. Questo modello appartiene a quella categoria di strumenti che mirano a soddisfare le esigenze identificate dal World Climate Research Program (WRCP) per affrontare gli effetti del cambiamento climatico. L'obiettivo perseguito è quello di sviluppare, seguendo un approccio ``bottom-up" al rischio climatico regionale, strumenti che possano aiutare i decisori a realizzare l'adattamento ai cambiamenti climatici. Il modello qui presentato è interamente basato su dati open-source forniti dai servizi Copernicus. Il contributo di questo lavoro di tesi riguarda lo sviluppo di un modello, formulato da (Ruggieri et al.), per stimare i danni di eventi alluvionali fluviali per specifici i livelli di riscaldamento globale (GWL). Il modello è stato testato su tre bacini idrografici di medie dimensioni in Emilia-Romagna, Panaro, Reno e Secchia. In questo lavoro, il modello viene sottoposto a test di sensibilità rispetto a un'ipotesi enunciata nella formulazione del modello, poi vengono effettuate analisi relative all'ensemble multi-modello utilizzato per le proiezioni. Il modello viene quindi validato, confrontando i danni stimati nel clima attuale per i tre fiumi con i danni osservati e confrontando le portate simulate con quelle osservate. Infine, vengono stimati i danni associati agli eventi alluvionali in tre scenari climatici futuri caratterizzati da GWL di 1.5° C, 2.0° C e 3.0°C.