Single-Cell RNA Sequencing Analysis: A Step-by-Step Overview

Thanks to innovative sample-preparation and sequencing technologies, gene expression in individual cells can now be measured for thousands of cells in a single experiment. Since its introduction, single-cell RNA sequencing (scRNA-seq) approaches have revolutionized the genomics field as they created unprecedented opportunities for resolving cell heterogeneity by exploring gene expression profiles at a single-cell resolution. However, the rapidly evolving field of scRNA-seq invoked the emergence of various analytics approaches aimed to maximize the full potential of this novel strategy. Unlike population-based RNA sequencing approaches, scRNA seq necessitates comprehensive computational tools to address high data complexity and keep up with the emerging single-cell associated challenges. Despite the vast number of analytical methods, a universal standardization is lacking. While this reflects the fields’ immaturity, it may also encumber a newcomer to blend in. In this review, we aim to bridge over the abovementioned hurdle and propose four ready-to-use pipelines for scRNA-seq analysis easily accessible by a newcomer, that could fit various biological data types. Here we provide an overview of the currently available single-cell technologies for cell isolation and library preparation and a step by step guide that covers the entire canonical analytic workflow to analyse scRNA-seq data including read mapping, quality controls, gene expression quantification, normalization, feature selection, dimensionality reduction, and cell clustering useful for trajectory inference and differential expression. Such workflow guidelines will escort novices as well as expert users in the analysis of complex scRNA-seq datasets, thus further expanding the research potential of single-cell approaches in basic science, and envisaging its future implementation as best practice in the field

The impact of microRNAs on transcriptional heterogeneity and gene co-expression across single embryonic stem cells

MicroRNAs act posttranscriptionally to suppress multiple target genes within a cell population. To what extent this multi-target suppression occurs in individual cells and how it impacts transcriptional heterogeneity and gene co-expression remains unknown. Here we used single-cell sequencing combined with introduction of individual microRNAs. miR-294 and let-7c were introduced into otherwise microRNA-deficient Dgcr8 knockout mouse embryonic stem cells. Both microRNAs induce suppression and correlated expression of their respective gene targets. The two microRNAs had opposing effects on transcriptional heterogeneity within the cell population, with let-7c increasing and miR-294 decreasing the heterogeneity between cells. Furthermore, let-7c promotes, whereas miR-294 suppresses, the phasing of cell cycle genes. These results show at the individual cell level how a microRNA simultaneously has impacts on its many targets and how that in turn can influence a population of cells. The findings have important implications in the understanding of how microRNAs influence the co-expression of genes and pathways, and thus ultimately cell fate

Mantra 2.0: An online collaborative resource for drug mode of action and repurposing by network analysis

Elucidation of molecular targets of a compound (mode of action, MoA) and of its off-targets is a crucial step in drug development. We developed an online collaborative resource (MANTRA 2.0) that supports this process by exploiting similarities between drug-induced transcriptional profiles. Drugs are organised in a network of nodes (drugs) and edges (similarities) highlighting “communities” of drugs sharing a similar MoA. A user can upload gene expression profiles (GEPs) before and after drug treatment in one or multiple cell types. An automated processing pipeline transforms the GEPs into a unique drug ”node” embedded in the drug-network. Visual inspection of the neighbouring drugs and communities helps in revealing its MoA, and to suggest new applications of known drugs (drug repurposing). MANTRA 2.0 allows storing and sharing user-generated network nodes, thus making MANTRA 2.0 a collaborative ever-growing resource

05/18/1993 - Eastern\u27s 1993 All Student Show Winner Laura Brahos.pdf

This paper reports on the stages forming a model evaluation protocol for urban flow and dispersion models proposed within the COST Action 732 on "Quality Assurance and Improvement of Micro-Scale Meteorological Models". It discusses the different components forming model evaluation with emphasis on validation and implementation of the protocol for the test case Mock Urban Setting Test (MUST). The protocol was proposed with building-resolving models in mind, but integral models have also been included. The suggested approach can be used for further micro-scale model evaluation and for the standardisation of their applications

Democracy by Design: Perspectives for Digitally Assisted, Participatory Upgrades of Society

The technological revolution, particularly the availability of more data and more powerful computational tools, has led to the emergence of a new scientific field called “Computational Diplomacy”. Our work tries to define its scope and focuses on a popular subarea of it, namely “Digital Democracy”. In recent years, there has been a surge of interest in using digital technologies to promote more participatory forms of democracy. While there are numerous potential benefits to using digital tools to enhance democracy, significant challenges must be addressed. It is essential to ensure that digital technologies are used in an accessible, equitable, and fair manner rather than reinforcing existing power imbalances. This paper investigates how digital tools can be used to help design more democratic societies by investigating three key research areas: (1) the role of digital technologies for facilitating civic engagement in collective decision-making; (2) the use of digital tools to improve transparency and accountability in governance; and (3) the potential for digital technologies to enable the formation of more inclusive and representative democracies. We argue that more research on how digital technologies can be used to support democracy upgrade is needed. Along these lines, we lay out a research agenda for the future

THE MUST MODEL EVALUATION EXERCISE: PATTERNS IN MODEL PERFORMANCE

As part of the COST 732 action more than a dozen different research groups have modelled the MUST experiment, as simulated in a wind tunnel. The model evaluation guidance developed within COST 732 recommends \u27exploratory data analysis\u27 as one of the elements in model validation. Experience has shown that such exploratory analysis is crucial to reveal shortcomings of models that might otherwise pass unnoticed. Conditions are best for detecting common patterns and anomalies if you have a situation where several models are put into a common framework – like the case at hand. The available material provides a unique opportunity to identify and explore patterns within model performance

Linear B-cell epitopes in the spike and nucleocapsid proteins as markers of SARS-CoV-2 exposure and disease severity

BACKGROUND Given the unceasing worldwide surge in COVID-19 cases, there is an imperative need to develop highly specific and sensitive serology assays to define exposure to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). METHODS Pooled plasma samples from PCR positive COVID-19 patients were used to identify linear B-cell epitopes from a SARS-CoV-2 peptide library of spike (S), envelope (E), membrane (M), and nucleocapsid (N) structural proteins by peptide-based ELISA. Hit epitopes were further validated with 79 COVID-19 patients with different disease severity status, 13 seasonal human CoV, 20 recovered SARS patients and 22 healthy donors. FINDINGS Four immunodominant epitopes, S14P5, S20P2, S21P2 and N4P5, were identified on the S and N viral proteins. IgG responses to all identified epitopes displayed a strong detection profile, with N4P5 achieving the highest level of specificity (100%) and sensitivity (>96%) against SARS-CoV-2. Furthermore, the magnitude of IgG responses to S14P5, S21P2 and N4P5 were strongly associated with disease severity. INTERPRETATION IgG responses to the peptide epitopes can serve as useful indicators for the degree of immunopathology in COVID-19 patients, and function as higly specific and sensitive sero-immunosurveillance tools for recent or past SARS-CoV-2 infections. The flexibility of these epitopes to be used alone or in combination will allow for the development of improved point-of-care-tests (POCTs)

THE MUST MODEL EVALUATION EXERCISE: STATISTICAL ANALYSIS OF MODELLING RESULTS

The first validation exercise of the COST action 732 lead to a substantial number of simulation results for comparison with the MUST wind tunnel experiments. Validation metrics for selected simulation results of the flow field and the concentrations are presented and compared to the state of the art. In addition mean metrics and corresponding scatter limits are computed from the individual results