DoubletDecon: Deconvoluting Doublets from Single-Cell RNA-Sequencing Data.
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ABSTRACT:
SUBMITTER: DePasquale EAK
PROVIDER: S-EPMC6983270 | biostudies-literature | 2019 Nov
REPOSITORIES: biostudies-literature
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Project description:MotivationSingle-cell RNA sequencing (scRNA-seq) continues to expand our knowledge by facilitating the study of transcriptional heterogeneity at the level of single cells. Despite this technology's utility and success in biomedical research, technical artifacts are present in scRNA-seq data. Doublets/multiplets are a type of artifact that occurs when two or more cells are tagged by the same barcode, and therefore they appear as a single cell. Because this introduces non-existent transcriptional profiles, doublets can bias and mislead downstream analysis. To address this limitation, computational methods to annotate and remove doublets form scRNA-seq datasets are needed.ResultsWe introduce vaeda (Variational Auto-Encoder for Doublet Annotation), a new approach for computational annotation of doublets in scRNA-seq data. Vaeda integrates a variational auto-encoder and Positive-Unlabeled learning to produce doublet scores and binary doublet calls. We apply vaeda, along with seven existing doublet annotation methods, to 16 benchmark datasets and find that vaeda performs competitively in terms of doublet scores and doublet calls. Notably, vaeda outperforms other python-based methods for doublet annotation. Altogether, vaeda is a robust and competitive method for scRNA-seq doublet annotation and may be of particular interest in the context of python-based workflows.Availability and implementationVaeda is available at https://github.com/kostkalab/vaeda, and the version used for the results we present here is archived at zenodo (https://doi.org/10.5281/zenodo.7199783).Supplementary informationSupplementary data are available at Bioinformatics online.
| S-EPMC9805559 | biostudies-literature
Project description:MotivationSingle-cell RNA sequencing (scRNA-seq) technologies enable the study of transcriptional heterogeneity at the resolution of individual cells and have an increasing impact on biomedical research. However, it is known that these methods sometimes wrongly consider two or more cells as single cells, and that a number of so-called doublets is present in the output of such experiments. Treating doublets as single cells in downstream analyses can severely bias a study's conclusions, and therefore computational strategies for the identification of doublets are needed.ResultsWith scds, we propose two new approaches for in silico doublet identification: Co-expression based doublet scoring (cxds) and binary classification based doublet scoring (bcds). The co-expression based approach, cxds, utilizes binarized (absence/presence) gene expression data and, employing a binomial model for the co-expression of pairs of genes, yields interpretable doublet annotations. bcds, on the other hand, uses a binary classification approach to discriminate artificial doublets from original data. We apply our methods and existing computational doublet identification approaches to four datasets with experimental doublet annotations and find that our methods perform at least as well as the state of the art, at comparably little computational cost. We observe appreciable differences between methods and across datasets and that no approach dominates all others. In summary, scds presents a scalable, competitive approach that allows for doublet annotation of datasets with thousands of cells in a matter of seconds.Availability and implementationscds is implemented as a Bioconductor R package (doi: 10.18129/B9.bioc.scds).Supplementary informationSupplementary data are available at Bioinformatics online.
| S-EPMC7703774 | biostudies-literature
Project description:MotivationWhile single-cell DNA sequencing (scDNA-seq) has enabled the study of intratumor heterogeneity at an unprecedented resolution, current technologies are error-prone and often result in doublets where two or more cells are mistaken for a single cell. Not only do doublets confound downstream analyses, but the increase in doublet rate is also a major bottleneck preventing higher throughput with current single-cell technologies. Although doublet detection and removal are standard practice in scRNA-seq data analysis, options for scDNA-seq data are limited. Current methods attempt to detect doublets while also performing complex downstream analyses tasks, leading to decreased efficiency and/or performance.ResultsWe present doubletD, the first standalone method for detecting doublets in scDNA-seq data. Underlying our method is a simple maximum likelihood approach with a closed-form solution. We demonstrate the performance of doubletD on simulated data as well as real datasets, outperforming current methods for downstream analysis of scDNA-seq data that jointly infer doublets as well as standalone approaches for doublet detection in scRNA-seq data. Incorporating doubletD in scDNA-seq analysis pipelines will reduce complexity and lead to more accurate results.Availability and implementationhttps://github.com/elkebir-group/doubletD.Supplementary informationSupplementary data are available at Bioinformatics online.
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Project description:Estimating cell type compositions for complex diseases is an important step to investigate the cellular heterogeneity for understanding disease etiology and potentially facilitate early disease diagnosis and prevention. Here, we developed a computationally statistical method, referring to Multi-Omics Matrix Factorization (MOMF), to estimate the cell-type compositions of bulk RNA sequencing (RNA-seq) data by leveraging cell type-specific gene expression levels from single-cell RNA sequencing (scRNA-seq) data. MOMF not only directly models the count nature of gene expression data, but also effectively accounts for the uncertainty of cell type-specific mean gene expression levels. We demonstrate the benefits of MOMF through three real data applications, i.e., Glioblastomas (GBM), colorectal cancer (CRC) and type II diabetes (T2D) studies. MOMF is able to accurately estimate disease-related cell type proportions, i.e., oligodendrocyte progenitor cells and macrophage cells, which are strongly associated with the survival of GBM and CRC, respectively.
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Project description:Single-cell RNA-sequencing has become a widely used, powerful approach for studying cell populations. However, these methods often generate multiplet artifacts, where two or more cells receive the same barcode, resulting in a hybrid transcriptome. In most experiments, multiplets account for several percent of transcriptomes and can confound downstream data analysis. Here, we present Single-Cell Remover of Doublets (Scrublet), a framework for predicting the impact of multiplets in a given analysis and identifying problematic multiplets. Scrublet avoids the need for expert knowledge or cell clustering by simulating multiplets from the data and building a nearest neighbor classifier. To demonstrate the utility of this approach, we test Scrublet on several datasets that include independent knowledge of cell multiplets. Scrublet is freely available for download at github.com/AllonKleinLab/scrublet.
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Project description:Single-cell RNA-sequencing (scRNA-seq) techniques provide unprecedented opportunities to investigate phenotypic and molecular heterogeneity in complex biological systems. However, profiling massive amounts of cells brings great computational challenges to accurately and efficiently characterize diverse cell populations. Single cell discriminant analysis (scDA) solves this problem by simultaneously identifying cell groups and discriminant metagenes based on the construction of cell-by-cell representation graph, and then using them to annotate unlabeled cells in data. We demonstrate scDA is effective to determine cell types, revealing the overall variabilities between cells from eleven data sets. scDA also outperforms several state-of-the-art methods when inferring the labels of new samples. In particular, we found scDA less sensitive to drop-out events and capable to label a mass of cells within or across datasets after learning even from a small set of data. The scDA approach offers a new way to efficiently analyze scRNA-seq profiles of large size or from different batches. scDA was implemented and freely available at https://github.com/ZCCQQWork/scDA.
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Project description:Integrating single-cell RNA sequencing (scRNA-seq) data with genotypes obtained from DNA sequencing studies facilitates the detection of functional genetic variants underlying cell type-specific gene expression variation. Unfortunately, most existing scRNA-seq studies do not come with DNA sequencing data; thus, being able to call single nucleotide variants (SNVs) from scRNA-seq data alone can provide crucial and complementary information, detection of functional SNVs, maximizing the potential of existing scRNA-seq studies. Here, we perform extensive analyses to evaluate the utility of two SNV calling pipelines (GATK and Monovar), originally designed for SNV calling in either bulk or single-cell DNA sequencing data. In both pipelines, we examined various parameter settings to determine the accuracy of the final SNV call set and provide practical recommendations for applied analysts. We found that combining all reads from the single cells and following GATK Best Practices resulted in the highest number of SNVs identified with a high concordance. In individual single cells, Monovar resulted in better quality SNVs even though none of the pipelines analyzed is capable of calling a reasonable number of SNVs with high accuracy. In addition, we found that SNV calling quality varies across different functional genomic regions. Our results open doors for novel ways to leverage the use of scRNA-seq for the future investigation of SNV function.
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Project description:As single-cell RNA sequencing (scRNA-seq) technologies have rapidly developed, so have analysis methods. Many methods have been tested, developed, and validated using simulated datasets. Unfortunately, current simulations are often poorly documented, their similarity to real data is not demonstrated, or reproducible code is not available. Here, we present the Splatter Bioconductor package for simple, reproducible, and well-documented simulation of scRNA-seq data. Splatter provides an interface to multiple simulation methods including Splat, our own simulation, based on a gamma-Poisson distribution. Splat can simulate single populations of cells, populations with multiple cell types, or differentiation paths.
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Project description:Single-cell RNA sequencing (scRNA-seq) is able to give an insight into the gene-gene associations or transcriptional networks among cell populations based on the sequencing of a large number of cells. However, traditional network methods are limited to the grouped cells instead of each single cell, and thus the heterogeneity of single cells will be erased. We present a new method to construct a cell-specific network (CSN) for each single cell from scRNA-seq data (i.e. one network for one cell), which transforms the data from 'unstable' gene expression form to 'stable' gene association form on a single-cell basis. In particular, it is for the first time that we can identify the gene associations/network at a single-cell resolution level. By CSN method, scRNA-seq data can be analyzed for clustering and pseudo-trajectory from network perspective by any existing method, which opens a new way to scRNA-seq data analyses. In addition, CSN is able to find differential gene associations for each single cell, and even 'dark' genes that play important roles at the network level but are generally ignored by traditional differential gene expression analyses. In addition, CSN can be applied to construct individual network of each sample bulk RNA-seq data. Experiments on various scRNA-seq datasets validated the effectiveness of CSN in terms of accuracy and robustness.
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