The tidyomics ecosystem: enhancing omic data analyses.

The growth of omic data presents evolving challenges in data manipulation, analysis and integration. Addressing these challenges, Bioconductor provides an extensive community-driven biological data analysis platform. Meanwhile, tidy R programming offers a revolutionary data organization and manipulation standard. Here we present the tidyomics software ecosystem, bridging Bioconductor to the tidy R paradigm. This ecosystem aims to streamline omic analysis, ease learning and encourage cross-disciplinary collaborations. We demonstrate the effectiveness of tidyomics by analyzing 7.5 million peripheral blood mononuclear cells from the Human Cell Atlas, spanning six data frameworks and ten analysis tools.

Museum of spatial transcriptomics.

The function of many biological systems, such as embryos, liver lobules, intestinal villi, and tumors, depends on the spatial organization of their cells. In the past decade, high-throughput technologies have been developed to quantify gene expression in space, and computational methods have been developed that leverage spatial gene expression data to identify genes with spatial patterns and to delineate neighborhoods within tissues. To comprehensively document spatial gene expression technologies and data-analysis methods, we present a curated review of literature on spatial transcriptomics dating back to 1987, along with a thorough analysis of trends in the field, such as usage of experimental techniques, species, tissues studied, and computational approaches used. Our Review places current methods in a historical context, and we derive insights about the field that can guide current research strategies. A companion supplement offers a more detailed look at the technologies and methods analyzed: https://pachterlab.github.io/LP_2021/ .

Identification of spatial homogeneous regions in tissues with concordex.

Spatial homogeneous regions (SHRs) in tissues are domains that are homogeneous with respect to cell type composition. We present a method for identifying SHRs using spatial transcriptomics data, and demonstrate that it is efficient and effective at finding SHRs for a wide variety of tissue types. The method is implemented in a tool called concordex, which relies on analysis of k-nearest-neighbor (kNN) graphs. The concordex tool is also useful for analysis of non-spatial transcriptomics data, and can elucidate the extent of concordance between partitions of cells derived from clustering algorithms, and transcriptomic similarity as represented in kNN graphs.

kallisto, bustools, and kb-python for quantifying bulk, single-cell, and single-nucleus RNA-seq.

The term "RNA-seq" refers to a collection of assays based on sequencing experiments that involve quantifying RNA species from bulk tissue, from single cells, or from single nuclei. The kallisto, bustools, and kb-python programs are free, open-source software tools for performing this analysis that together can produce gene expression quantification from raw sequencing reads. The quantifications can be individualized for multiple cells, multiple samples, or both. Additionally, these tools allow gene expression values to be classified as originating from nascent RNA species or mature RNA species, making this workflow amenable to both cell-based and nucleus-based assays. This protocol describes in detail how to use kallisto and bustools in conjunction with a wrapper, kb-python, to preprocess RNA-seq data.

The impact of package selection and versioning on single-cell RNA-seq analysis.

Standard single-cell RNA-sequencing analysis (scRNA-seq) workflows consist of converting raw read data into cell-gene count matrices through sequence alignment, followed by analyses including filtering, highly variable gene selection, dimensionality reduction, clustering, and differential expression analysis. Seurat and Scanpy are the most widely-used packages implementing such workflows, and are generally thought to implement individual steps similarly. We investigate in detail the algorithms and methods underlying Seurat and Scanpy and find that there are, in fact, considerable differences in the outputs of Seurat and Scanpy. The extent of differences between the programs is approximately equivalent to the variability that would be introduced in benchmarking scRNA-seq datasets by sequencing less than 5% of the reads or analyzing less than 20% of the cell population. Additionally, distinct versions of Seurat and Scanpy can produce very different results, especially during parts of differential expression analysis. Our analysis highlights the need for users of scRNA-seq to carefully assess the tools on which they rely, and the importance of developers of scientific software to prioritize transparency, consistency, and reproducibility for their tools.

kallisto, bustools and kb-python for quantifying bulk, single-cell and single-nucleus RNA-seq.

The term 'RNA-seq' refers to a collection of assays based on sequencing experiments that involve quantifying RNA species from bulk tissue, single cells or single nuclei. The kallisto, bustools and kb-python programs are free, open-source software tools for performing this analysis that together can produce gene expression quantification from raw sequencing reads. The quantifications can be individualized for multiple cells, multiple samples or both. Additionally, these tools allow gene expression values to be classified as originating from nascent RNA species or mature RNA species, making this workflow amenable to both cell-based and nucleus-based assays. This protocol describes in detail how to use kallisto and bustools in conjunction with a wrapper, kb-python, to preprocess RNA-seq data. Execution of this protocol requires basic familiarity with a command line environment. With this protocol, quantification of a moderately sized RNA-seq dataset can be completed within minutes.

The tidyomics ecosystem: Enhancing omic data analyses.

The growth of omic data presents evolving challenges in data manipulation, analysis, and integration. Addressing these challenges, Bioconductor1 provides an extensive community-driven biological data analysis platform. Meanwhile, tidy R programming2 offers a revolutionary standard for data organisation and manipulation. Here, we present the tidyomics software ecosystem, bridging Bioconductor to the tidy R paradigm. This ecosystem aims to streamline omic analysis, ease learning, and encourage cross-disciplinary collaborations. We demonstrate the effectiveness of tidyomics by analysing 7.5 million peripheral blood mononuclear cells from the Human Cell Atlas3, spanning six data frameworks and ten analysis tools.

Voyager: exploratory single-cell genomics data analysis with geospatial statistics.

Exploratory spatial data analysis (ESDA) can be a powerful approach to understanding single-cell genomics datasets, but it is not yet part of standard data analysis workflows. In particular, geospatial analyses, which have been developed and refined for decades, have yet to be fully adapted and applied to spatial single-cell analysis. We introduce the Voyager platform, which systematically brings the geospatial ESDA tradition to (spatial) -omics, with local, bivariate, and multivariate spatial methods not yet commonly applied to spatial -omics, united by a uniform user interface. Using Voyager, we showcase biological insights that can be derived with its methods, such as biologically relevant negative spatial autocorrelation. Underlying Voyager is the SpatialFeatureExperiment data structure, which combines Simple Feature with SingleCellExperiment and AnnData to represent and operate on geometries bundled with gene expression data. Voyager has comprehensive tutorials demonstrating ESDA built on GitHub Actions to ensure reproducibility and scalability, using data from popular commercial technologies. Voyager is implemented in both R/Bioconductor and Python/PyPI, and features compatibility tests to ensure that both implementations return consistent results.