SUMMARY: HTSeq 2.0 provides a more extensive application programming interface including a new representation for sparse genomic data, enhancements for htseq-count to suit single-cell omics, a new script for data using cell and molecular barcodes, improved documentation, testing and deployment, bug fixes and Python 3 support. AVAILABILITY AND IMPLEMENTATION: HTSeq 2.0 is released as an open-source software under the GNU General Public License and is available from the Python Package Index at https://pypi.python.org/pypi/HTSeq. The source code is available on Github at https://github.com/htseq/htseq. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

BACKGROUND: Visualization is an important tool for generating meaning from scientific data, but the visualization of structures in high-dimensional data (such as from high-throughput assays) presents unique challenges. Dimension reduction methods are key in solving this challenge, but these methods can be misleading- especially when apparent clustering in the dimension-reducing representation is used as the basis for reasoning about relationships within the data. RESULTS: We present two interactive visualization tools, distnet and focusedMDS, that help in assessing the validity of a dimension-reducing plot and in interactively exploring relationships between objects in the data. The distnet tool is used to examine discrepancies between the placement of points in a two dimensional visualization and the points' actual similarities in feature space. The focusedMDS tool is an intuitive, interactive multidimensional scaling tool that is useful for exploring the relationships of one particular data point to the others, that might be useful in a personalized medicine framework. CONCLUSIONS: We introduce here two freely available tools for visually exploring and verifying the validity of dimension-reducing visualizations and biological information gained from these. The use of such tools can confirm that conclusions drawn from dimension-reducing visualizations are not simply artifacts of the visualization method, but are real biological insights.

MOTIVATION: A large choice of tools exists for many standard tasks in the analysis of high-throughput sequencing (HTS) data. However, once a project deviates from standard workflows, custom scripts are needed. RESULTS: We present HTSeq, a Python library to facilitate the rapid development of such scripts. HTSeq offers parsers for many common data formats in HTS projects, as well as classes to represent data, such as genomic coordinates, sequences, sequencing reads, alignments, gene model information and variant calls, and provides data structures that allow for querying via genomic coordinates. We also present htseq-count, a tool developed with HTSeq that preprocesses RNA-Seq data for differential expression analysis by counting the overlap of reads with genes. AVAILABILITY AND IMPLEMENTATION: HTSeq is released as an open-source software under the GNU General Public Licence and available from http://www-huber.embl.de/HTSeq or from the Python Package Index at https://pypi.python.org/pypi/HTSeq.

MOTIVATION: Circularized Chromosome Conformation Capture (4C) is a powerful technique for studying the spatial interactions of a specific genomic region called the 'viewpoint' with the rest of the genome, both in a single condition or comparing different experimental conditions or cell types. Observed ligation frequencies typically show a strong, regular dependence on genomic distance from the viewpoint, on top of which specific interaction peaks are superimposed. Here, we address the computational task to find these specific peaks and to detect changes between different biological conditions. RESULTS: We model the overall trend of decreasing interaction frequency with genomic distance by fitting a smooth monotonically decreasing function to suitably transformed count data. Based on the fit, z-scores are calculated from the residuals, and high z-scores are interpreted as peaks providing evidence for specific interactions. To compare different conditions, we normalize fragment counts between samples, and call for differential contact frequencies using the statistical method DESEQ2: adapted from RNA-Seq analysis. AVAILABILITY AND IMPLEMENTATION: A full end-to-end analysis pipeline is implemented in the R package FourCSeq available at www.bioconductor.org. CONTACT: [email protected] or [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

UNLABELLED: In many genomic studies, one works with genome-position-dependent data, e.g. ChIP-chip or ChIP-Seq scores. Using conventional tools, it can be difficult to get a good feel for the data, especially the distribution of features. This article argues that the so-called Hilbert curve visualization can complement genome browsers and help to get further insights into the structure of one's data. This is demonstrated with examples from different use cases. An open-source application, called HilbertVis, is presented that allows the user to produce and interactively explore such plots. AVAILABILITY: http://www.ebi.ac.uk/huber-srv/hilbert/.

UNLABELLED: ShortRead is a package for input, quality assessment, manipulation and output of high-throughput sequencing data. ShortRead is provided in the R and Bioconductor environments, allowing ready access to additional facilities for advanced statistical analysis, data transformation, visualization and integration with diverse genomic resources. AVAILABILITY AND IMPLEMENTATION: This package is implemented in R and available at the Bioconductor web site; the package contains a 'vignette' outlining typical work flows.