Benchmarking algorithms for pathway activity transformation of single-cell RNA-seq data
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ABSTRACT:
SUBMITTER: Zhang Y
PROVIDER: S-EPMC7642725 | biostudies-literature | 2020 Jan
REPOSITORIES: biostudies-literature
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Project description:BackgroundSingle-cell RNA-sequencing (scRNA-seq) measures gene expression in single cells, while single-nucleus ATAC-sequencing (snATAC-seq) quantifies chromatin accessibility in single nuclei. These two data types provide complementary information for deciphering cell types and states. However, when analyzed individually, they sometimes produce conflicting results regarding cell type/state assignment. The power is compromised since the two modalities reflect the same underlying biology. Recently, it has become possible to measure both gene expression and chromatin accessibility from the same nucleus. Such paired data enable the direct modeling of the relationships between the two modalities. Given the availability of the vast amount of single-modality data, it is desirable to integrate the paired and unpaired single-modality datasets to gain a comprehensive view of the cellular complexity.ResultsWe benchmark nine existing single-cell multi-omic data integration methods. Specifically, we evaluate to what extent the multiome data provide additional guidance for analyzing the existing single-modality data, and whether these methods uncover peak-gene associations from single-modality data. Our results indicate that multiome data are helpful for annotating single-modality data. However, we emphasize that the availability of an adequate number of nuclei in the multiome dataset is crucial for achieving accurate cell type annotation. Insufficient representation of nuclei may compromise the reliability of the annotations. Additionally, when generating a multiome dataset, the number of cells is more important than sequencing depth for cell type annotation.ConclusionsSeurat v4 is the best currently available platform for integrating scRNA-seq, snATAC-seq, and multiome data even in the presence of complex batch effects.
| S-EPMC10594700 | biostudies-literature
Project description:Recent advancements in single-cell sequencing technologies have generated extensive omics data in various modalities and revolutionized cell research, especially in the single-cell RNA and ATAC data. The joint analysis across scRNA-seq data and scATAC-seq data has paved the way to comprehending the cellular heterogeneity and complex cellular regulatory networks. Multi-omics integration is gaining attention as an important step in joint analysis, and the number of computational tools in this field is growing rapidly. In this paper, we benchmarked 12 multi-omics integration methods on three integration tasks via qualitative visualization and quantitative metrics, considering six main aspects that matter in multi-omics data analysis. Overall, we found that different methods have their own advantages on different aspects, while some methods outperformed other methods in most aspects. We therefore provided guidelines for selecting appropriate methods for specific scenarios and tasks to help obtain meaningful insights from multi-omics data integration.
| S-EPMC10944570 | biostudies-literature
Project description:Accurate identification of human leukocyte antigen (HLA) alleles is essential for various clinical and research applications, such as transplant matching and drug sensitivities. Recent advances in RNA-seq technology have made it possible to impute HLA types from sequencing data, spurring the development of a large number of computational HLA typing tools. However, the relative performance of these tools is unknown, limiting the ability for clinical and biomedical research to make informed choices regarding which tools to use. Here we report the study design of a comprehensive benchmarking of the performance of 12 HLA callers across 682 RNA-seq samples from 8 datasets with molecularly defined gold standard at 5 loci, HLA-A, -B, -C, -DRB1, and -DQB1. For each HLA typing tool, we will comprehensively assess their accuracy, compare default with optimized parameters, and examine for discrepancies in accuracy at the allele and loci levels. We will also evaluate the computational expense of each HLA caller measured in terms of CPU time and RAM. We also plan to evaluate the influence of read length over the HLA region on accuracy for each tool. Most notably, we will examine the performance of HLA callers across European and African groups, to determine discrepancies in accuracy associated with ancestry. We hypothesize that RNA-Seq HLA callers are capable of returning high-quality results, but the tools that offer a good balance between accuracy and computational expensiveness for all ancestry groups are yet to be developed. We believe that our study will provide clinicians and researchers with clear guidance to inform their selection of an appropriate HLA caller.
| S-EPMC10827116 | biostudies-literature
Project description:BACKGROUND:Single-cell RNA-sequencing (scRNA-seq) technologies and associated analysis methods have rapidly developed in recent years. This includes preprocessing methods, which assign sequencing reads to genes to create count matrices for downstream analysis. While several packaged preprocessing workflows have been developed to provide users with convenient tools for handling this process, how they compare to one another and how they influence downstream analysis have not been well studied. RESULTS:Here, we systematically benchmark the performance of 10 end-to-end preprocessing workflows (Cell Ranger, Optimus, salmon alevin, alevin-fry, kallisto bustools, dropSeqPipe, scPipe, zUMIs, celseq2, and scruff) using datasets yielding different biological complexity levels generated by CEL-Seq2 and 10x Chromium platforms. We compare these workflows in terms of their quantification properties directly and their impact on normalization and clustering by evaluating the performance of different method combinations. While the scRNA-seq preprocessing workflows compared vary in their detection and quantification of genes across datasets, after downstream analysis with performant normalization and clustering methods, almost all combinations produce clustering results that agree well with the known cell type labels that provided the ground truth in our analysis. CONCLUSIONS:In summary, the choice of preprocessing method was found to be less important than other steps in the scRNA-seq analysis process. Our study comprehensively compares common scRNA-seq preprocessing workflows and summarizes their characteristics to guide workflow users.
| S-EPMC8672463 | biostudies-literature
Project description:Single-cell RNA sequencing (scRNA-seq) technologies allow numerous opportunities for revealing novel and potentially unexpected biological discoveries. scRNA-seq clustering helps elucidate cell-to-cell heterogeneity and uncover cell subgroups and cell dynamics at the group level. Two important aspects of scRNA-seq data analysis were introduced and discussed in the present review: relevant datasets and analytical tools. In particular, we reviewed popular scRNA-seq datasets and discussed scRNA-seq clustering models including K-means clustering, hierarchical clustering, consensus clustering, and so on. Seven state-of-the-art scRNA clustering methods were compared on five public available datasets. Two primary evaluation metrics, the Adjusted Rand Index (ARI) and the Normalized Mutual Information (NMI), were used to evaluate these methods. Although unsupervised models can effectively cluster scRNA-seq data, these methods also have challenges. Some suggestions were provided for future research directions.
| S-EPMC7549635 | biostudies-literature
Project description:Single-cell RNA-sequencing (scRNA-seq) enables researchers to quantify transcriptomes of thousands of cells simultaneously and study transcriptomic changes between cells. scRNA-seq datasets increasingly include multisubject, multicondition experiments to investigate cell-type-specific differential states (DS) between conditions. This can be performed by first identifying the cell types in all the subjects and then by performing a DS analysis between the conditions within each cell type. Naïve single-cell DS analysis methods that treat cells statistically independent are subject to false positives in the presence of variation between biological replicates, an issue known as the pseudoreplicate bias. While several methods have already been introduced to carry out the statistical testing in multisubject scRNA-seq analysis, comparisons that include all these methods are currently lacking. Here, we performed a comprehensive comparison of 18 methods for the identification of DS changes between conditions from multisubject scRNA-seq data. Our results suggest that the pseudobulk methods performed generally best. Both pseudobulks and mixed models that model the subjects as a random effect were superior compared with the naïve single-cell methods that do not model the subjects in any way. While the naïve models achieved higher sensitivity than the pseudobulk methods and the mixed models, they were subject to a high number of false positives. In addition, accounting for subjects through latent variable modeling did not improve the performance of the naïve methods.
| S-EPMC9487674 | biostudies-literature
Project description:We present a systematic evaluation of state-of-the-art algorithms for inferring gene regulatory networks from single-cell transcriptional data. As the ground truth for assessing accuracy, we use synthetic networks with predictable trajectories, literature-curated Boolean models and diverse transcriptional regulatory networks. We develop a strategy to simulate single-cell transcriptional data from synthetic and Boolean networks that avoids pitfalls of previously used methods. Furthermore, we collect networks from multiple experimental single-cell RNA-seq datasets. We develop an evaluation framework called BEELINE. We find that the area under the precision-recall curve and early precision of the algorithms are moderate. The methods are better in recovering interactions in synthetic networks than Boolean models. The algorithms with the best early precision values for Boolean models also perform well on experimental datasets. Techniques that do not require pseudotime-ordered cells are generally more accurate. Based on these results, we present recommendations to end users. BEELINE will aid the development of gene regulatory network inference algorithms.
| S-EPMC7098173 | biostudies-literature
Project description:Single-cell RNA-seq has the potential to facilitate isoform quantification as the confounding factor of a mixed population of cells is eliminated. However, best practice for using existing quantification methods has not been established. We carry out a benchmark for five popular isoform quantification tools. Performance is generally good for simulated data based on SMARTer and SMART-seq2 data. The reduction in performance compared with bulk RNA-seq is small. An important biological insight comes from our analysis of real data which shows that genes that express two isoforms in bulk RNA-seq predominantly express one or neither isoform in individual cells.
| S-EPMC6223048 | biostudies-literature
Project description:With the development of single-cell RNA sequencing (scRNA-seq) technology, analysts need to integrate hundreds of thousands of cells with multiple experimental batches. It is becoming increasingly difficult for users to select the best integration methods to remove batch effects. Here, we compared the advantages and limitations of four commonly used Scanpy-based batch-correction methods using two representative and large-scale scRNA-seq datasets. We quantitatively evaluated batch-correction performance and efficiency. Furthermore, we discussed the performance differences among the evaluated methods at the algorithm level.
| S-EPMC7338326 | biostudies-literature
Project description:Single-cell RNA-seq data contain a large proportion of zeros for expressed genes. Such dropout events present a fundamental challenge for various types of data analyses. Here, we describe the SCRABBLE algorithm to address this problem. SCRABBLE leverages bulk data as a constraint and reduces unwanted bias towards expressed genes during imputation. Using both simulation and several types of experimental data, we demonstrate that SCRABBLE outperforms the existing methods in recovering dropout events, capturing true distribution of gene expression across cells, and preserving gene-gene relationship and cell-cell relationship in the data.
| S-EPMC6501316 | biostudies-literature