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| Author | : Almut Lütge |
| Publisher | : |
| Total Pages | : 0 |
| Release | : 2023 |
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| ISBN | : |
| Author | : Nan Xi |
| Publisher | : |
| Total Pages | : 203 |
| Release | : 2021 |
| Genre | : |
| ISBN | : |
The large-scale, high-dimensional, and sparse single-cell RNA sequencing (scRNA-seq) data have raised great challenges in the pipeline of data analysis. A large number of statistical and machine learning methods have been developed to analyze scRNA-seq data and answer related scientific questions. Although different methods claim advantages in certain circumstances, it is difficult for users to select appropriate methods for their analysis tasks. Benchmark studies aim to provide recommendations for method selection based on an objective, accurate, and comprehensive comparison among cutting-edge methods. They can also offer suggestions for further methodological development through massive evaluations conducted on real data. In Chapter 2, we conduct the first, systematic benchmark study of nine cutting-edge computational doublet-detection methods. In scRNA-seq, doublets form when two cells are encapsulated into one reaction volume by chance. The existence of doublets, which appear as but are not real cells, is a key confounder in scRNA-seq data analysis. Computational methods have been developed to detect doublets in scRNA-seq data; however, the scRNA-seq field lacks a comprehensive benchmarking of these methods, making it difficult for researchers to choose an appropriate method for their specific analysis needs. Our benchmark study compares doublet-detection methods in terms of their detection accuracy under various experimental settings, impacts on downstream analyses, and computational efficiency. Our results show that existing methods exhibited diverse performance and distinct advantages in different aspects. In Chapter 3, we develop an R package DoubletCollection to integrate the installation and execution of different doublet-detection methods. Traditional benchmark studies can be quickly out-of-date due to their static design and the rapid growth of available methods. DoubletCollection addresses this issue in benchmarking doublet-detection methods for scRNA-seq data. DoubletCollection provides a unified interface to perform and visualize downstream analysis after doublet-detection. Additionally, we created a protocol using DoubletCollection to execute and benchmark doublet-detection methods. This protocol can automatically accommodate new doublet-detection methods in the fast-growing scRNA-seq field. In Chapter 4, we conduct the first comprehensive empirical study to explore the best modeling strategy for autoencoder-based imputation methods specific to scRNA-seq data. The autoencoder-based imputation method is a family of promising methods to denoise sparse scRNA-seq data; however, the design of autoencoders has not been formally discussed in the literature. Current autoencoder-based imputation methods either borrow the practice from other fields or design the model on an ad hoc basis. We find that the method performance is sensitive to the key hyperparameter of autoencoders, including architecture, activation function, and regularization. Their optimal settings on scRNA-seq are largely different from those on other data types. Our results emphasize the importance of exploring hyperparameter space in such complex and flexible methods. Our work also points out the future direction of improving current methods.
| Author | : Guo-Cheng Yuan |
| Publisher | : Humana Press |
| Total Pages | : 271 |
| Release | : 2019-02-14 |
| Genre | : Science |
| ISBN | : 9781493990566 |
This detailed book provides state-of-art computational approaches to further explore the exciting opportunities presented by single-cell technologies. Chapters each detail a computational toolbox aimed to overcome a specific challenge in single-cell analysis, such as data normalization, rare cell-type identification, and spatial transcriptomics analysis, all with a focus on hands-on implementation of computational methods for analyzing experimental data. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Computational Methods for Single-Cell Data Analysis aims to cover a wide range of tasks and serves as a vital handbook for single-cell data analysis.
| Author | : Ruochen Jiang |
| Publisher | : |
| Total Pages | : 175 |
| Release | : 2021 |
| Genre | : |
| ISBN | : |
Next generation sequencing (NGS) has revolutionized biomedical research and has a broad impact and applications. Since its advent around 15 years ago, this high scalable DNA sequencing technology has generated numerous biological data with new features and brought new challenges to data analysis. For example, researchers utilize RNA sequencing (RNA-seq) technology to more accurately quantify the gene expression levels. However, the NGS technology involves many processing steps and technical variations when measuring the expression values in the biological samples. In other words, the NGS data researchers observed could be biased due to the randomness and constraints in the NGS technology. This dissertation will mainly focus on microbiome sequencing data and single-cell RNA-seq (scRNA-seq) data. Both of them are highly sparse matrix-form count data. The zeros could either be biological or non-biological, and the high sparsity in the data have brought challenges to data analysis. Missing data imputation problem has been studied in statistics and social science as the survey data often experience non-response to some of the survey questions and those unresponded questions will be marked as "NA" or missing values in the data. Imputation methods are used to provide a sophisticated guess for the missing values, and the purpose is to avoid discarding the collected samples and for the ease of using the state-of-the-art statistical methods. In machine learning, the famous Netflix data challenge regarding film recommendation system also falls into the missing data imputation problem category. Netflix wants to find a way to predict users' fondness of the movies they have not watched. The potential scores these users would give to the unwatched films are regarded as missing values in the data. NGS data imputation problem is different from the previous two cases in that the missing values in the NGS data are not so well-defined. The zeros in the NGS data could either come from the biological origin (should not be regarded as missing values) or non-biological origin (due to the limitation of the sequencing technology and should be regarded as missing values). The size (number of samples and features) of the NGS matrix data is usually larger than the size of survey data but smaller than the size of the recommendation system data. In addition, in most cases, the percentage of missing values in the survey data is less than the percentage of zeros in the NGS data, and the missing values in the film recommendation system data have the highest percentage (> 99.9%). As a result, the commonly used missing data imputation methods in statistics and machine learning are not directly applicable to NGS data. In recent years, numerous imputation methods have been proposed to deal with the highly sparse scRNA-seq data. In light of this, this dissertation aims to address two questions. First, the microbiome sequencing data, having additional information comparing to the scRNA-seq data, lacks an imputation method. Secondly, whether to use imputation or not in scRNA-seq data analysis is still a controversial problem. The first part of this dissertation focuses on the first imputation method developed for the microbiome sequencing data: mbImpute. Microbiome studies have gained increased attention since many discoveries revealed connections between human microbiome compositions and diseases. A critical challenge in microbiome data analysis is the existence of many non-biological zeros, which distort taxon abundance distributions, complicate data analysis, and jeopardize the reliability of scientific discoveries. To address this issue, we propose the first imputation method for microbiome data---mbImpute---to identify and recover likely non-biological zeros by borrowing information jointly from similar samples, similar taxa, and optional metadata including sample covariates and taxon phylogeny. Comprehensive simulations verify that mbImpute achieves better imputation accuracy under multiple metrics, compared with five state-of-the-art imputation methods designed for non-microbiome data. In real data applications, we demonstrate that mbImpute improves the power of identifying disease-related taxa from microbiome data of type 2 diabetes and colorectal cancer, and mbImpute preserves non-zero distributions of taxa abundances. The second part of this dissertation focuses on how to deal with high sparsity in the scRNA-seq data. ScRNA-seq technologies have revolutionized biomedical sciences by enabling genome-wide profiling of gene expression levels at an unprecedented single-cell resolution. A distinct characteristic of scRNA-seq data is the vast proportion of zeros unseen in bulk RNA-seq data. Researchers view these zeros differently: some regard zeros as biological signals representing no or low gene expression, while others regard zeros as false signals or missing data to be corrected. As a result, the scRNA-seq field faces much controversy regarding how to handle zeros in data analysis. We first discuss the sources of biological and non-biological zeros in scRNA-seq data. Second, we evaluate the impacts of non-biological zeros on cell clustering and differential gene expression analysis. Third, we summarize the advantages, disadvantages, and suitable users of three input data types: observed counts, imputed counts, and binarized counts and evaluate the performance of downstream analysis on these three input data types. Finally, we discuss the open questions regarding non-biological zeros, the need for benchmarking, and the importance of transparent analysis.
| Author | : Jessica Lu Zhou |
| Publisher | : |
| Total Pages | : 0 |
| Release | : 2023 |
| Genre | : |
| ISBN | : |
Single-cell sequencing has emerged as a powerful tool for dissecting cellular heterogeneity and providing cell type-specific biological insights. Single-cell sequencing technologies have rapidly proliferated over the last decade, leading to an explosion of data generated from such experiments. However, several challenges exist in the computational analysis of single-cell sequencing data due to its large and complex nature, including the need for sophisticated statistical methods to distinguish biologically meaningful signals from noise, the integration of single-cell sequencing data with other types of biological information, and the development of scalable and reproducible computational pipelines that can handle the large and complex nature of the data. In this dissertation, I present two distinct projects analyzing single-cell sequencing data. The first is of an analytical nature and tackles a translational question. In this project, I built computational pipelines for processing and analyzing single-nucleus RNA- and ATAC-sequencing datasets generated from the amygdalae of genetically diverse heterogenous stock rats, which were subjected to a behavioral protocol for studying addiction-like behaviors following cocaine self-administration. In doing so, I provide a standard reference for analyzing such data as well as reveal cell type-specific insights into the molecular underpinnings of cocaine addiction. The second project is oriented towards methods development and seeks to understand the fundamental biological question of transcriptional regulation. Here, I developed a statistical framework for simulating and modeling data from single-cell CRISPR regulatory screens and used it to perform a genome-wide interrogation of epistatic-like interactions between enhancer pairs. I found that multiple enhancers act together in a multiplicative fashion with little evidence for interactive effects between them. This work revealed novel insights into the collective behavior of multiple regulatory elements and provides a tool that can be applied to future datasets generated from such experiments. This dissertation exemplifies how computational methods can be applied in different contexts to extract meaning from a variety of single-cell sequencing modalities. By tackling both a translational and fundamental biological question, I have showcased the breadth of what can be revealed by studying single-cell sequencing data and the computational methods necessary to extract this information.
| Author | : Xinzhou Ge |
| Publisher | : |
| Total Pages | : 186 |
| Release | : 2021 |
| Genre | : |
| ISBN | : |
The development of high-throughput biological technologies have enabled researchers to simultaneously perform analysis on thousands of features (e.g., genes, genomic regions, and proteins). The most common goal of analyzing high-throughput data is to contrast two conditions, to identify ``interesting'' features, whose values differ between two conditions. How to contrast the features from two conditions to extract useful information from high-throughput data, and how to ensure the reliability of identified features are two increasingly pressing challenge to statistical and computational science. This dissertation aim to address these two problems regarding analysing high-throughput data from two conditions. My first project focuses on false discovery rate (FDR) control in high-throughput data analysis from two conditions. FDR is defined as the expected proportion of uninteresting features among the identified ones. It is the most widely-used criterion to ensure the reliability of the interesting features identified. Existing bioinformatics tools primarily control the FDR based on p-values. However, obtaining valid p-values relies on either reasonable assumptions of data distribution or large numbers of replicates under both conditions, two requirements that are often unmet in biological studies. In Chapter \ref{chap:clipper}, we propose Clipper, a general statistical framework for FDR control without relying on p-values or specific data distributions. Clipper is applicable to identifying both enriched and differential features from high-throughput biological data of diverse types. In comprehensive simulation and real-data benchmarking, Clipper outperforms existing generic FDR control methods and specific bioinformatics tools designed for various tasks, including peak calling from ChIP-seq data, and differentially expressed gene identification from bulk or single-cell RNA-seq data. Our results demonstrate Clipper's flexibility and reliability for FDR control, as well as its broad applications in high-throughput data analysis. My second project focuses on alignment of multi-track epigenomic signals from different samples or conditions. The availability of genome-wide epigenomic datasets enables in-depth studies of epigenetic modifications and their relationships with chromatin structures and gene expression. Various alignment tools have been developed to align nucleotide or protein sequences in order to identify structurally similar regions. However, there are currently no alignment methods specifically designed for comparing multi-track epigenomic signals and detecting common patterns that may explain functional or evolutionary similarities. We propose a new local alignment algorithm, EpiAlign, designed to compare chromatin state sequences learned from multi-track epigenomic signals and to identify locally aligned chromatin regions. EpiAlign is a dynamic programming algorithm that novelly incorporates varying lengths and frequencies of chromatin states. We demonstrate the efficacy of EpiAlign through extensive simulations and studies on the real data from the NIH Roadmap Epigenomics project. EpiAlign can also detect common chromatin state patterns across multiple epigenomes from conditions, and it will serve as a useful tool to group and distinguish epigenomic samples based on genome-wide or local chromatin state patterns.
| Author | : Filippo Geraci |
| Publisher | : Frontiers Media SA |
| Total Pages | : 169 |
| Release | : 2020-06-08 |
| Genre | : |
| ISBN | : 2889637050 |
| Author | : Hui Ting Grace Yeo |
| Publisher | : |
| Total Pages | : 176 |
| Release | : 2020 |
| Genre | : |
| ISBN | : |
Single-cell RNA-sequencing (scRNA-seq) enables transcriptome-wide measurements of single cells at scale. As scRNA-seq datasets grow in complexity and size, more complex computational methods are required to distill raw data into biological insight. In this thesis, we introduce computational methods that enable analysis of novel scRNA-seq perturbational assays. We also develop computational models that seek to move beyond simple observations of cell states toward more complex models of underlying biological processes. In particular, we focus on cellular differentiation, which is the process by which cells acquire some specific form or function. First, we introduce barcodelet scRNA-seq (barRNA-seq), an assay which tags individual cells with RNA ‘barcodelets’ to identify them based on the treatments they receive. We apply barRNA-seq to study the effects of the combinatorial modulation of signaling pathways during early mESC differentiation toward germ layer and mesodermal fates. Using a data-driven analysis framework, we identify combinatorial signaling perturbations that drive cells toward specific fates. Second, we describe poly-adenine CRISPR gRNA-based scRNA-seq (pAC-seq), a method that enables the direct observation of guide RNAs (gRNAs) in scRNA-seq. We apply it to assess the phenotypic consequences of CRISPR/Cas9-based alterations of gene cis-regulatory regions. We find that power to detect transcriptomic effects depend on factors such as rate of mono/biallelic loss, baseline gene expression, and the number of cells per target gRNA. Third, we propose a generative model for analyzing scRNA-seq containing unwanted sources of variation. Using only weak supervision from a control population, we show that the model enables removal of nuisance effects from the learned representation without prior knowledge of the confounding factors. Finally, we develop a generative modeling framework that learns an underlying differentiation landscape from population-level time-series data. We validate the modeling framework on an experimental lineage tracing dataset, and show that it is able to recover the expected effects of known modulators of cell fate in hematopoiesis.
| Author | : Jingyi Li |
| Publisher | : |
| Total Pages | : 226 |
| Release | : 2013 |
| Genre | : |
| ISBN | : |
In the burgeoning field of genomics, high-throughput technologies (e.g. microarrays, next-generation sequencing and label-free mass spectrometry) have enabled biologists to perform global analysis on thousands of genes, mRNAs and proteins simultaneously. Extracting useful information from enormous amounts of high-throughput genomic data is an increasingly pressing challenge to statistical and computational science. In this thesis, I will address three problems in which statistical and computational methods were used to analyze high-throughput genomic data to answer important biological questions. The first part of this thesis focuses on addressing an important question in genomics: how to identify and quantify mRNA products of gene transcription (i.e., isoforms) from next-generation mRNA sequencing (RNA-Seq) data? We developed a statistical method called Sparse Linear modeling of RNA-Seq data for Isoform Discovery and abundance Estimation (SLIDE) that employs probabilistic modeling and L1 sparse estimation to answer this ques- tion. SLIDE takes exon boundaries and RNA-Seq data as input to discern the set of mRNA isoforms that are most likely to present in an RNA-Seq sample. It is based on a linear model with a design matrix that models the sampling probability of RNA-Seq reads from different mRNA isoforms. To tackle the model unidentifiability issue, SLIDE uses a modified Lasso procedure for parameter estimation. Compared with existing deterministic isoform assembly algorithms, SLIDE considers the stochastic aspects of RNA-Seq reads in exons from different isoforms and thus has increased power in detecting more novel isoforms. Another advantage of SLIDE is its flexibility of incorporating other transcriptomic data into its model to further increase isoform discovery accuracy. SLIDE can also work downstream of other RNA-Seq assembly algorithms to integrate newly discovered genes and exons. Besides isoform discovery, SLIDE sequentially uses the same linear model to estimate the abundance of discovered isoforms. Simulation and real data studies show that SLIDE performs as well as or better than major competitors in both isoform discovery and abundance estimation. The second part of this thesis demonstrates the power of simple statistical analysis in correcting biases of system-wide protein abundance estimates and in understanding the rela- tionship between gene transcription and protein abundances. We found that proteome-wide surveys have significantly underestimated protein abundances, which differ greatly from previously published individual measurements. We corrected proteome-wide protein abundance estimates by using individual measurements of 61 housekeeping proteins, and then found that our corrected protein abundance estimates show a higher correlation and a stronger linear relationship with mRNA abundances than do the uncorrected protein data. To estimate the degree to which mRNA expression levels determine protein levels, it is critical to measure the error in protein and mRNA abundance data and to consider all genes, not only those whose protein expression is readily detected. This is a fact that previous proteome-widely surveys ignored. We took two independent approaches to re-estimate the percentage that mRNA levels explain in the variance of protein abundances. While the percentages estimated from the two approaches vary on different sets of genes, all suggest that previous protein-wide surveys have significantly underestimated the importance of transcription. In the third and final part, I will introduce a modENCODE (the Model Organism ENCyclopedia Of DNA Elements) project in which we compared developmental stages, tis- sues and cells (or cell lines) of Drosophila melanogaster and Caenorhabditis elegans, two well-studied model organisms in developmental biology. To understand the similarity of gene expression patterns throughout their development time courses is an interesting and important question in comparative genomics and evolutionary biology. The availability of modENCODE RNA-Seq data for different developmental stages, tissues and cells of the two organisms enables a transcriptome-wide comparison study to address this question. We undertook a comparison of their developmental time courses and tissues/cells, seeking com- monalities in orthologous gene expression. Our approach centers on using stage/tissue/cell- associated orthologous genes to link the two organisms. For every stage/tissue/cell in each organism, its associated genes are selected as the genes capturing specific transcriptional activities: genes highly expressed in that stage/tissue/cell but lowly expressed in a few other stages/tissues/cells. We aligned a pair of D. melanogaster and C. elegans stages/tissues/cells by a hypergeometric test, where the test statistic is the number of orthologous gene pairs associated with both stages/tissues/cells. The test is against the null hypothesis that the two stages/tissues/cells have independent sets of associated genes. We first carried out the alignment approach on pairs of stages/tissues/cells within D. melanogaster and C. elegans respectively, and the alignment results are consistent with previous findings, supporting the validity of this approach. When comparing fly with worm, we unexpectedly observed two parallel collinear alignment patterns between their developmental timecourses and several interesting alignments between their tissues and cells. Our results are the first findings regarding a comprehensive comparison between D. melanogaster and C. elegans time courses, tissues and cells.
| Author | : Silvio Bicciato |
| Publisher | : Humana |
| Total Pages | : 0 |
| Release | : 2022-09-04 |
| Genre | : Science |
| ISBN | : 9781071613924 |
This volume details a comprehensive set of methods and tools for Hi-C data processing, analysis, and interpretation. Chapters cover applications of Hi-C to address a variety of biological problems, with a specific focus on state-of-the-art computational procedures adopted for the data analysis. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Hi-C Data Analysis: Methods and Protocols aims to help computational and molecular biologists working in the field of chromatin 3D architecture and transcription regulation.
