Seminario de Título entregado a la Universidad de Chile para optar al Título de Ingeniero en Biotecnología Molecular. / The transcription process is initiated by the Core Promoter Elements (CPE), leading the formation of the Pre-Initiation Complex (PIC) which reclutes the transcription proteins (General Transcription Factors, GTFs) and RNA Polymerase II (RNA Pol II) that bind to the CPE. These GTFs are: TBP, TFIIA, TFIIB, TFIIE, TFIIF and TFIIH in the case of promoter recognized by the RNA Pol II that synthesizes the mRNA molecule using DNA sequence as template. Among the CPE are the TATA box, the TFIIB-recognition Element (BRE), the Initiator (Inr), the Motif Ten Element (MTE), Downstream Promoter Element (DPE) and the Downstream Core Element (DCE) (Thomas y Chiang, 2006) Data collected during the laboratory experimental work suggest that the DNA sequence named Homol D is a universal CPE in S. pombe and that the genes containing this element are transcribed by the RNA Pol II, which is also the target of a new factor or protein complex that determined the initiation site of transcription. We aim to determine which GTFs are required for Homol D box dependent gene transcription, through in vitro transcription assays, inmunodepletion and Electrophoresis Mobility Shift Assay (EMSA). Here we show the analysis of the following specific antibodies for GTFs of interest (TBP, TAF1, TFIIB, TFIIE, TFIIH, TFIIF and the subunit of Srb4 mediator). xii Protein extract of S. pombe were used as GTFs source, while ribosomal K5 gene (that possess the Homol D box) were used as DNA template. To carry out transcription was used a purified RNA Pol II. The in vitro transcription and inmunodepletion experiments allowed us to observe that transcription does not occur when each transcription factor studied is depleted by using a specific antibody for each GTF. When using a promoter with an inverted Homol D as DNA template the same results were observed. When a rescue of the transcription reaction is performed by supplementing the depleted GTF after the inmunodepletion, the reaction occurred for both the normal and the inverted Homol D box. Therefore the studied GTFs, TBP, TFIIB, TFIIE, TFIIH and TFIIF are necessary for initiation of the transcription process in the K5 promoter which contains Homol D box. For the EMSA assays it was initially obtained a purified protein fraction with Homol D union and antibodies against each GTF to make an inmunodepletion of each GTFs and afterwards the reaction was incubated with P32γATP labeled probe. This assays allowed us to observe that the studied transcription factors are not the ones that recognize directly the Homol D box sequence. According with the work performed in this work, the GTFs TBP, TAF1, TFIIB, TFIIE, TFIIH, TFIIF and Srb4 are necessary for the formation of PIC but not for the recognition of the Homol D box in the K5 promoter, suggesting the existence of another factor/s o protein complex that recognizes the sequence of Homol D box. / El proceso de la transcripción es iniciado por los Elementos del Núcleo del Promotor (Core Promoter Element, CPE), que dirigen la formación del Complejo de Preiniciación (Pre-Initiation Complex, PIC), el cual está compuesto por los Factores de Transcripción Generales (General Transcription Factors, GTFs) y la RNA Polimerasa II (RNA Pol II). TBP, TFIIA, TFIIB, TFIIE, TFIIF y TFIIH corresponden a los GTFs que participan en la transcripción dirigida por promotores reconocidos por la RNA Pol II que sintetiza la molécula de mRNA a partir de una secuencia de DNA. Entre los CPE se encuentran la caja TATA, el elemento de reconocimiento de TFIIB (BRE), el Iniciador (Inr), el Motif Ten Element (MTE), el Downstream Promoter Element (DPE) y el Downstream Core Element (DCE) (Thomas y Chiang, 2006). Antecedentes recopilados durante el trabajo experimental de laboratorio sugieren que la secuencia de DNA denominada Homol D es un CPE universal en S. pombe y que los genes que poseen este elemento son transcritos por la RNA Pol II, la que además es blanco de un nuevo factor o complejo proteico que determina el sitio de inicio de la transcripción. Como objetivo de esta tesis se determinaron los requerimientos de GTFs necesarios para la transcripción de genes con promotores que poseen la caja Homol D, a través de ensayos de transcripción in vitro, inmunodepleción y geles de retardo (EMSA), en los cuales se utilizaron anticuerpos específicos para los distintos GTFs estudiados (TBP, TAF1, TFIIB, TFIIE, TFIIH, TFIIF y una subunidad del mediador Srb4). xiv Como fuente de GTFs se utilizaron extractos de proteínas del organismo S. pombe y como DNA molde fue utilizado el promotor del gen ribosomal K5 que posee la caja Homol D. Para llevar a cabo la transcripción se utilizó una RNA Pol II purificada. Los experimentos de transcripción in vitro e inmunodepleción, permitieron observar que la transcripción no ocurre al retirar cualquiera de los factores de transcripción por acción del anticuerpo específico para dicho GTFs. Además los ensayos usando como DNA molde un promotor con la caja Homol D invertida, permitieron observar los mismos resultados, es decir que la transcripción no ocurre al faltar alguno de los GTFs analizados en esta tesis. Al realizar un rescate de la reacción de transcripción, es decir, luego de hacer la inmunodepleción se suplementa el GTFs eliminado, se observó que la reacción ocurría, tanto para la caja Homol D normal como para la invertida, por lo tanto la presencia de todos los GTFs estudiados, TBP, TFIIB, TFIIE, TFIIH y TFIIF son necesarios para que se inicie el proceso de transcripción en el promotor K5 que posee la caja Homol D. Para los ensayos EMSA se utilizó extracto de proteínas totales que contenía la proteína con unión a Homol D y anticuerpos contra cada GTF, para realizar una inmunodepleción de ellos y luego se incubó con una sonda de DNA que contenía la caja Homol D marcada con P32γATP. Estos ensayos permitieron observar que los factores de transcripción estudiados no son los que reconocen de manera directa la secuencia de la caja Homol D, ya que a pesar de que ellos son retirados de la reacción se observa la aparición del complejo proteína/Homol D, reflejado como una banda en el film fotográfico. xv De acuerdo al trabajo realizado en este seminario de título, los GTFs TBP, TAF1, TFIIB, TFIIE, TFIIH, TFIIF y Srb4 son necesarios para la formación del PIC pero no para el reconocimiento de la caja Homol D en el promotor K5, sugiriendo que existiría otro factor o complejo proteico que reconoce la secuencia de la caja Homol D.

Tillgänglighet är något som går att mäta på olika sätt där ett tillvägagångssätt är med hjälp av mobilitet. Kollektivtrafiken spelar en stor roll för invånarna i en större tätort som Uppsalas mobilitet och för att knyta ihop stadens olika delar. Mobiliteten blir här ett sätt att mäta tillgänglighet, för att nå de olika destinationer måste en individ resa. I denna studie undersöks invånarna i Uppsalas tillgång till kollektivtrafiken för att kunna resa inom tätorten men även om General Transit Feed Specification är en lämplig metod. Detta görs genom en kvantitativ metod där befolkningsdata kombineras med data om kollektivtrafiken. Befolkningsdata hämtas från Statistiska Centralbyrån och data om kollektivtrafiken har hämtats i formatet General Transit Feed Specification. Med hjälp av dessa data ges möjlighet att undersöka vilka delar av staden och vilken del av befolkningen som har eller saknar tillgänglighet till kollektivtrafik. Resultatet av denna undersökning visar att tillgängligheten för detta sätt att resa skiljer sig mellan olika inkomstgrupper och delar av Uppsala. Denna undersöknings resultat visar även att General Transit Feed Specification är ett bra steg på vägen men kompletteringar behövs.

Taking accessibility into consideration is crucial when planning for public transport in a city. The bus network of the Swedish city Uppsala has undergone major changes since 2017, featuring a new circular bus line and several exchange points where commuters can change to regional and city buses easily. Measuring the accessibility and mobility of the residents of Uppsala will be the focus of this study, to detect underserviced areas and to measure the availability of buses throughout the week. Measuring accessibility and mobility were visualised primarily with the help of Geographic Information System [GIS] and data from General Transit Feed Specification [GTFS]. This is conducted through a quantitative method by comparing population data, ridership count and public transport data. The data is collected from GTFS, Region Uppsala and Uppsala University. In the results, the supply of buses in central Uppsala meets the demand of the commuters, and most importantly during peak hours. The exchange points are valuable for commuters to execute their journeys by having broader route alternatives to choose from. Nevertheless, the usage of bus stations is sufficient in most parts of Uppsala. However, there are certain areas in central Uppsala that are lacking accessibility to bus stations.

An expansion of the railway, East Coast Line is essential in order to ensure transportation of passenger and goods back and forth to Northern Sweden. The preliminary studies of the planned expansion to a double track have identified vulnerabilities linked to how our climate changes. Because of these risks and vulnerabilities, the railway station in Hudiksvall needs to be relocated or the Current station needs to be adapted to potential future climate scenarios. Furthermore, social sustainability and the aspect of accessibility is also a vital perspective to consider during the development of railway infrastructure. This study compares the three different station locations from an accessibility perspective and from different climate scenarios through Network Analyst in ArcGIS Pro. To visualize future climate scenarios, two RCP-scenarios (Representative Concentration Pathways) are considered, which is RCP 4.5 and RCP8.5. Additionally, the GTFS specification in ArcGIS Pro is used to model public transit to these railway stations in an accessibility perspective. Because one strategy when developing the East Coast Line is to increase the active transportation in comparison to car transportation.  Results from this study indicates that the Current station, which is located in a coastal area will be worst affected of potential future climate scenarios from an accessibility perspective. Other findings are that vulnerable groups in the society, such as low-income earners and elderly will be most affected, if the railway station remains in the current location. The results from the performed Service area analysis and Location-allocation analysis advocates the Eastern station as a location for the new railway station. / En expansion av Ostkustbanan är viktigt för att kunna säkerställa transport av passagerare och gods till och från norra Sverige. Förstudierna av den planerade expansionen till ett dubbelspår har identifierat sårbarheter kopplat till hur vårt klimat förändras. Som en följd av dessa risker och sårbarheter behöver järnvägsstationen i Hudiksvall flyttas. Alternativt behöver den nuvarande klimatanpassas. Detta ställer krav på att ta hänsyn till den sociala hållbarheten och tillgängligheten för befolkningen i Hudiksvall. Denna studie ämnar att jämför de tre olika stationslägena ur ett tillgänglighetsperspektiv samt utifrån olika klimatscenarier i ArcGIS Pro. För att visualisera framtida klimatscenarier beaktas två RCP-scenarier, det vill säga representativa koncentrationsvägar, vilket är RCP4.5 och RCP8.5. Dessutom används GTFS-specifikationen i ArcGIS Pro för att modellera kollektivtrafik till dessa järnvägsstationer ur ett tillgänglighetsperspektiv, då ett mål med utvecklingen av Ostkustbanan är att utöka andelen av personer som väljer aktiv transport i jämförelse med biltransporter.  Resultatet av denna studie visar att den nuvarande stationen, som är placerad i närheten av Hudiksvalls kust, kommer att vara hårdast drabbad av potentiella framtida klimatscenarier ur ett tillgänglighetsperspektiv. Vidare visar studien på att låginkomsttagare och äldre kommer att drabbas hårdast om järnvägsstationen ligger kvar på den nuvarande platsen. Resultatet från de utförda Service area analyserna och Location-allocation analyserna, visar att det östra alternativet är det alternativ som kommer att vara minst påverkad av ett framtida klimat från ett tillgänglighetsperspektiv.

Transport models are key in predicting travel behaviour and planning transport systems. Transport models can be either aggregated or disaggregated. Disaggregation means that travel behaviour is represented on an individual level, which can be beneficial because it offers a higher detail level and reduces aggregation bias. Input data for transport models can be both expensive and inaccessible, especially comprehensive data. Thus, it is advantageous to explore the utilisation of open data, which is free and accessible. The objective of the thesis was to evaluate how OpenStreetMap and other Open Data can be utilised in disaggregated transport modelling. The scope of the study was Uppsala, Sweden. In the thesis, a disaggregate transport model was designed, which only considered commuting trips made by public transport. Destinations and a synthetic population were estimated based on OpenStreetMap map features, SCB census data, and LuTRANS land use data. A travel survey was utilised in model calibration, and UL boarding data was used for model validation. The results showed that OpenStreetMap provided sufficient data for estimating a synthetic population and destinations for a disaggregate transport model when combined with other open data sources. Population and land usecensus data were essential for calibrating the model. However, the model came with limitations caused by assumptions, generalisation, technical constraints, and the partial incompleteness of open data. The thesis concludes that Open Data, such as OpenStreetMap, can be utilised sufficiently for transport modelling, with proper assumptions and processing. The openness of the data also increases the replicability of such a model. / Transportmodeller är viktiga i att förutspå resvanemönster och för att kunna planera transportsystemet. Transportmodeller kan vara antingen aggregeradeeller disaggregerade. Disaggregering betyder att resvanor är representerade påindividuell nivå, vilket kan vara fördelaktigt då det innebär en högre detalj nivå och mindre partiskhet orsakad av aggregering (aggregation bias). Indata förtransportmodeller kan vara både dyrt och svåråtkomligt, speciellt för mer omfattande data. Därav kan det vara till stor nytta att utforska möjligheten att använda öppnadata (Open Data), som är gratis och lättåtkomligt. Syftet med examensarbetetvar att utvärdera hur OpenStreetMap och annan Open Data kan användas idisaggregerad transportmodellering. Den geografiska omfattningen av studien är Uppsala tätort. En disaggregerad transportmodell togs fram i examensarbetet, sombara tog hänsyn till jobbresor med kollektivtrafik. Destinationer och en syntetiskbefolkning uppskattades utifrån OpenStreetMap objekt, befolkningsdata från SCB, samt markanvändningsdata från LuTRANS. En resvaneundersökning utnyttjadesför modellkalibrering och påstigningsdata från UL användes för modellvalidering.Resultaten visade att OpenStreetMap erbjöd tillräckligt med data för att ta framoch uppskatta en syntetisk befolkning och destinationer för en disaggregeradtransportmodell, om den kombineras med andra öppna datakällor. Befolkning- ochmarkanvändningsdata var avgörande i att kalibrera modellen. Dock så innefattar modellen vissa begränsningar som är orsakada av antaganden, generalisering, tekniskabegränsningar, samt ofullständigheten av Open Data. Slutsatsen är att Open Data, så som OpenStreetMap, kan utnyttjas för transportmodellering, om det kombineras med välformulerade antaganden och processering av datan. Datans öppenheten medför även en ökad replikerbarhet för en sådan modell.

Alors que plus de cinquante pays dans le monde ont entrepris une démarche d’ouverture des données publiques, la thèse enquête sur l’émergence et la mise en oeuvre des politiques d’open data. Elle repose sur l’analyse de sources publiques et sur une enquête ethnographique conduite dans sept collectivités locales et institutions françaises. Revenant sur six moments de définition de grands « principes » de l’open data et leur traduction en politique publique par une institution française, Etalab, ce travail montre comment la catégorisation par l’open data a porté l’attention sur les données, en particulier sous leur forme « brute », considérées comme une ressource inexploitée, le « nouveau pétrole » gisant sous les organisations. L’enquête montre que le processus de l’ouverture débute généralement par une phase d’identification marquée par des explorations progressives et incertaines. Elle permet de comprendre que l’identification constitue un geste d’instauration qui transforme progressivement les fichiers de gestion de l’administration en données. Leur mise en circulation provoque des frictions : pour sortir des réseaux sociotechniques de l’organisation, les données doivent généralement passer à travers des circuits de validation et des chaînes de traitement. Par ailleurs, les données doivent souvent subir d’importantes transformations avant leur ouverture pour devenir intelligibles à la fois par les machines et par les humains. Cette thèse montre enfin que l’instauration concerne aussi les publics dont il est attendu qu’ils visualisent, inspectent et exploitent les données ouvertes. L’instauration des publics par des instruments très divers constitue un autre pan du travail invisible des politiques d’open data. Il ressort enfin de cette thèse que l’obligation à l’ouverture des données publiques, une suite possible des politiques d’open data, pose de manière saillante une question fondamentale « qu’est-ce qu’une donnée ? » Plutôt que de réduire la donnée à une catégorie relative, qui s’appliquerait à toutes sortes de matériaux informationnels, les cas étudiés montrent qu’elle est généralement attribuée dès lors que les données sont le point de départ de réseauxsociotechniques dédiés à leur circulation, leur exploitation et leur mise en visibilité. / As more than fifty countries have launched an open data policy, this doctoral dissertation investigates on the emergence and implementation of such policies. It is based on the analysis of public sources and an ethnographic inquiry conducted in seven French local authorities and institutions. By retracing six moments of definitions of the “open data principles” and their implementation by a French institution, Etalab, this work shows how open data has brought attention to data, particularly in their raw form, considered as an untapped resource, the “new oil” lying under the organisations. The inquiry shows that the process of opening generally begins by a phase of identification marked by progressive and uncertain explorations. It allows to understand that data are progressively instantiated from management files into data. Their circulation provoke frictions: to leave the sociotechnical network of organisations, data generally go through validation circuits and chains of treatment. Besides, data must often undergo important treatments before their opening in order to become intelligible by machines as well as humans. This thesis shows eventually that data publics are also instantiated as they are expected to visualize, inspect and process the data. Data publics are instantiated through various tools, which compose another area of the invisible work of open data projects. Finally, it appears from this work that the possible legal requirement to open data asks a fundamental question, “what is data?” Instead of reducing data to a relational category, which would apply to any informational material, studied cases show that they generally are applied when data are a starting point of sociotechnical networks dedicated to their circulation, their exploitation and their visibility.

Mobile applications and real-time data are excellent tools for rapidly sharing information. Such information may concern public transportation, such as time tables and traffic delays. This project has involved the development of a trip planner, which can subscribe to real-time data in order to inform the end user about the position of transit vehicles and trip updates. A trip planner is an application which, after having been given a start and a destination by the user, generates the possible trip between these two locations. The route is displayed upon a map, along with information of how the trip is travelled. The real-time data, which is pushed to the application, will inform the user if vehicles are delayed and if the trip needs to be updated due to a missed bus or train. The trip planner for Itract developed in this project is using the graphical interface and some necessary Java classes from the open source application Open Trip Planner. The new trip planner, developed in this project, is compatible with the API of Itract, has some additional functionality and can subscribe to real-time information. To subscribe to real-time information, a database called Redis has been set up in connection to Itract. Another database, known as MongoDB, is used for persistant storage. / Itract

The thesis deals with the benefits of modern technologies to public land transport, an area which has not been notably impacted by modern technologies to this day. The work anal-yses both the way modern technologies can help transport companies, and also which tech-nologies in particular can bring something new to the conservative sector. The specific ob-jectives of the thesis are the analysis of D2D systems key components, benefits of Yield Management to the transport industry and implementation of carriers into the Bileto plat-form.

Transport simulation models play a crucial role in transportation planning, design, and operations, allowing for the replication of various scenarios through the incorporation of real-world data and parameters. Recently, agent-based transport models have gained prominence for their ability to simulate intricate metropolitan transport systems. These models take into account the distinct characteristics, decision-making processes, and interactions of individual agents. Among the array of agent-based transport models, MATSim stands out as a potent and adaptable tool for modeling transportation systems. A critical aspect of MATSim’s input preparation involves assigning activity location points using land use raster data. However, the characteristics of land use raster data present limitations in certain urban case studies such as Stockholm. In response, some researchers have turned their attention to buildings shapefile data, a commonly used geospatial data format. This study aims to improve the activity location assignment model by developing an evaluation workflow of model uncertainty for different geospatial input data in MATSim and empirically analyzing their impacts on simulation outcomes. Despite acknowledging data availability and activity representation limitations, the study’s results demonstrate that utilizingbuildings shapefiles as input data yields more consistent outcomes with reduced uncertainty. This suggests the promising potential of buildings shapefiles as a favorable data source for transportation modeling and planning within the studied scenarios.

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