Nanopore Sequencing: An Introduction

Nanopore Sequencing: An Introduction

PDF Nanopore Sequencing: An Introduction Download

  • Author: Branton Daniel
  • Publisher: World Scientific
  • ISBN: 9813270624
  • Category : Science
  • Languages : en
  • Pages : 216

This is an introductory text and laboratory manual to be used primarily in undergraduate courses. It is also useful for graduate students and research scientists who require an introduction to the theory and methods of nanopore sequencing. The book has clear explanations of the principles of this emerging technology, together with instructional material written by experts that describes how to use a MinION nanopore instrument for sequencing in research or the classroom.At Harvard University the book serves as a textbook and lab manual for a university laboratory course designed to intensify the intellectual experience of incoming undergraduates while exploring biology as a field of concentration. Nanopore sequencing is an ideal topic as a path to encourage students about the range of courses they will take in Biology by pre-emptively addressing the complaint about having to take a course in Physics or Maths while majoring in Biology. The book addresses this complaint by concretely demonstrating the range of topics — from electricity to biochemistry, protein structure, molecular engineering, and informatics — that a student will have to master in subsequent courses if he or she is to become a scientist who truly understands what his or her biology instrument is measuring when investigating biological phenomena.


Nanopore Sequencing

Nanopore Sequencing

PDF Nanopore Sequencing Download

  • Author: Kazuharu Arakawa
  • Publisher: Humana
  • ISBN: 9781071629987
  • Category : Science
  • Languages : en
  • Pages : 0

This volume provides comprehensive dry and wet experiments, methods, and applications on nanopore sequencing. Chapters guide readers through bioinformatic procedures, genome sequencing, analysis of repetitive regions, structural variations, rapid and on-site microbial identification, epidemiology, and transcriptome analysis. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and methods, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols. Authoritative and cutting-edge, Nanopore Sequencing: Methods and Protocols aims to be comprehensive guide for researchers.


Engineered Nanopores for Bioanalytical Applications

Engineered Nanopores for Bioanalytical Applications

PDF Engineered Nanopores for Bioanalytical Applications Download

  • Author: Joshua B. Edel
  • Publisher: William Andrew
  • ISBN: 143773474X
  • Category : Science
  • Languages : en
  • Pages : 189

Engineered Nanopores for Bioanalytical Applications is the first book to focus primarily on practical analytical applications of nanopore development. These nanoscale analytical techniques have exciting potential because they can be used in applications such as DNA sequencing, DNA fragment sizing, DNA/protein binding, and protein/protein binding. This book provides a solid professional reference on nanopores for readers in academia, industry and engineering and biomedical fields. In addition, the book describes the instrumentation, fabrication, and experimental methods necessary to carry out nanopore-based experiments for developing new devices. Includes application case studies for detection, identification and analysis of biomolecules and related functional nanomaterials Introduces the techniques of manufacturing solid state materials with functional nanopores Explains the use of nanopores in DNA sequencing and the wider range of applications from environmental monitoring to forensics


Improving the Accuracy and Application of Nanopore DNA Sequencing

Improving the Accuracy and Application of Nanopore DNA Sequencing

PDF Improving the Accuracy and Application of Nanopore DNA Sequencing Download

  • Author: Matthew T. Noakes
  • Publisher:
  • ISBN:
  • Category :
  • Languages : en
  • Pages : 215

DNA contains the code of life, forming the molecular basis for all of life's diversity. The past several decades have witnessed remarkable progress in our ability to read and understand life's code through DNA sequencing. While fast and cheap DNA sequencing technologies are revolutionizing both science and healthcare, a new generation of technologies capable of single-molecule sequencing1 promise to further revolutionize the field of DNA sequencing by addressing many of limitations of the previous methods. Nanopore DNA sequencing is one such emerging single-molecule sequencing technology, capable of long reads and direct detection of epigenetically-relevant modified bases. The basic nanopore sequencing devices consists of two wells filled with a conductive electrolyte solution separated by an impermeable membrane containing a single nanometer-size hole, or nanopore. A voltage applied across the membrane drives an ionic current through the nanopore. DNA is negatively charged in solution and so will by drawn through the pore by the voltage, blocking some of the ionic current. As the different nucleotides along the DNA block the ionic current to different extents, the series of current fluctuations in the recorded time series can be used to decode the sequence of the DNA molecule moving through the pore. DNA motion through the pore is controlled using a DNA-processing motor enzyme, which steps the DNA through in discrete steps slow enough to allow resolution of the sequence-dependent fluctuations in the ionic current. Commercial nanopore sequencing devices have recently become available, making good on the decades-long promise of this technology. However, despite considerable early success and fanfare accompanying these first nanopore sequencers, technology development is not complete. Particularly, the single-read de novo sequencing accuracy must be improved for this technology to reach its full potential2. In order to fully realize its promise, we must both improve the accuracy of nanopore sequencing and devise better methods of handling error-prone sequencing data. In this dissertation, I discuss my work in the Gundlach nanopore lab at the University of Washington towards the goals of improved nanopore sequencing accuracy and improved application of existing error-prone sequencing data. In chapter 1, I introduce the broad field of DNA sequencing. I cover the history of scientific interest in DNA and DNA sequencing and provide motivation for DNA sequencing as a worthwhile pursuit both for its scientific and medical merits. I also discuss previous and existing DNA sequencing technologies, as well as the limitations of these technologies that motivate the development of new methods such as nanopore sequencing. In chapter 2 I describe and introduce nanopore sequencing. I summarize the development of nanopore sequencing technology, how various challenges were overcome, and how currently available nanopore sequencing devices work, setting the stage for understanding the primary error modes limiting the sequencing accuracy of this technologies. In chapter 3, in I present my work on improving nanopore sequencing accuracy using a new method of DNA control for enzyme-actuated nanopore DNA sequencing. This new method, in which we use a time-varying voltage to control DNA motion through the pore in addition to a DNA-processing enzyme, is able to mitigate two of the primary error modes in nanopore sequencing and dramatically improve sequencing accuracy. I discuss the motivation behind this new method, outline how we were able to realize nanopore sequencing using this method, and demonstrate the improved sequencing accuracy it affords. In chapter 4, I shift the discussion over to my work on improving the application of nanopore sequencing data. Specifically, I introduce a method of aligning nanopore data that enables highly sensitive and specific sequence alignment and species identification even for low accuracy reads. I go over the motivation for this method, and present our findings of its improved performance over alternative methods. Finally, I conclude in chapter 5 where I discuss the implications of the demonstrated advances in the accuracy and application of nanopore sequencing, as well as look out towards further progress that can be made in both arenas.


Nanopores

Nanopores

PDF Nanopores Download

  • Author: Samir M. Iqbal
  • Publisher: Springer Science & Business Media
  • ISBN: 1441982523
  • Category : Technology & Engineering
  • Languages : en
  • Pages : 377

Nanopores are nanometer scale holes formed naturally by proteins or cells, and can be used for a variety of applications, including sequencing DNA and detecting anthrax. They can be integrated into artificially constructed encapsulated cells of silicon wafers while allowing small molecules like oxygen, glucose and insulin to pass, while keeping out large system molecules. "Nanopores: Sensing and Fundamental Biological Interactions" examines the emerging research directions surrounding nanopores such as genome sequencing and early disease detection using biomarker identification. Covering the applications of nanopores in genetics, proteomics, drug discovery, early disease detection and detection of emerging environmental threats, it is a must-have book for biomedicalengineers and research scientists.


Introduction to Genomics

Introduction to Genomics

PDF Introduction to Genomics Download

  • Author: Arthur Lesk
  • Publisher: Oxford University Press, USA
  • ISBN: 0199564353
  • Category : Medical
  • Languages : en
  • Pages : 421

This book covers the latest techniques that enable us to study the genome in detail, the book explores what the genome tells us about life at the level of the molecule, the cell, and the organism


Biological Sequence Analysis

Biological Sequence Analysis

PDF Biological Sequence Analysis Download

  • Author: Richard Durbin
  • Publisher: Cambridge University Press
  • ISBN: 113945739X
  • Category : Science
  • Languages : en
  • Pages : 372

Probabilistic models are becoming increasingly important in analysing the huge amount of data being produced by large-scale DNA-sequencing efforts such as the Human Genome Project. For example, hidden Markov models are used for analysing biological sequences, linguistic-grammar-based probabilistic models for identifying RNA secondary structure, and probabilistic evolutionary models for inferring phylogenies of sequences from different organisms. This book gives a unified, up-to-date and self-contained account, with a Bayesian slant, of such methods, and more generally to probabilistic methods of sequence analysis. Written by an interdisciplinary team of authors, it aims to be accessible to molecular biologists, computer scientists, and mathematicians with no formal knowledge of the other fields, and at the same time present the state-of-the-art in this new and highly important field.


Nanopore Sequencing

Nanopore Sequencing

PDF Nanopore Sequencing Download

  • Author: Kazuharu Arakawa
  • Publisher: Springer Nature
  • ISBN: 1071629964
  • Category : Science
  • Languages : en
  • Pages : 318

This volume provides comprehensive dry and wet experiments, methods, and applications on nanopore sequencing. Chapters guide readers through bioinformatic procedures, genome sequencing, analysis of repetitive regions, structural variations, rapid and on-site microbial identification, epidemiology, and transcriptome analysis. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and methods, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols. Authoritative and cutting-edge, Nanopore Sequencing: Methods and Protocols aims to be comprehensive guide for researchers.


Genomics in the Cloud

Genomics in the Cloud

PDF Genomics in the Cloud Download

  • Author: Geraldine A. Van der Auwera
  • Publisher: O'Reilly Media
  • ISBN: 1491975164
  • Category : Computers
  • Languages : en
  • Pages : 496

Data in the genomics field is booming. In just a few years, organizations such as the National Institutes of Health (NIH) will host 50+ petabytes—or over 50 million gigabytes—of genomic data, and they’re turning to cloud infrastructure to make that data available to the research community. How do you adapt analysis tools and protocols to access and analyze that volume of data in the cloud? With this practical book, researchers will learn how to work with genomics algorithms using open source tools including the Genome Analysis Toolkit (GATK), Docker, WDL, and Terra. Geraldine Van der Auwera, longtime custodian of the GATK user community, and Brian O’Connor of the UC Santa Cruz Genomics Institute, guide you through the process. You’ll learn by working with real data and genomics algorithms from the field. This book covers: Essential genomics and computing technology background Basic cloud computing operations Getting started with GATK, plus three major GATK Best Practices pipelines Automating analysis with scripted workflows using WDL and Cromwell Scaling up workflow execution in the cloud, including parallelization and cost optimization Interactive analysis in the cloud using Jupyter notebooks Secure collaboration and computational reproducibility using Terra


Introduction to Single Cell Omics

Introduction to Single Cell Omics

PDF Introduction to Single Cell Omics Download

  • Author: Xinghua Pan
  • Publisher: Frontiers Media SA
  • ISBN: 2889459209
  • Category :
  • Languages : en
  • Pages : 129

Single-cell omics is a progressing frontier that stems from the sequencing of the human genome and the development of omics technologies, particularly genomics, transcriptomics, epigenomics and proteomics, but the sensitivity is now improved to single-cell level. The new generation of methodologies, especially the next generation sequencing (NGS) technology, plays a leading role in genomics related fields; however, the conventional techniques of omics require number of cells to be large, usually on the order of millions of cells, which is hardly accessible in some cases. More importantly, harnessing the power of omics technologies and applying those at the single-cell level are crucial since every cell is specific and unique, and almost every cell population in every systems, derived in either vivo or in vitro, is heterogeneous. Deciphering the heterogeneity of the cell population hence becomes critical for recognizing the mechanism and significance of the system. However, without an extensive examination of individual cells, a massive analysis of cell population would only give an average output of the cells, but neglect the differences among cells. Single-cell omics seeks to study a number of individual cells in parallel for their different dimensions of molecular profile on genome-wide scale, providing unprecedented resolution for the interpretation of both the structure and function of an organ, tissue or other system, as well as the interaction (and communication) and dynamics of single cells or subpopulations of cells and their lineages. Importantly single-cell omics enables the identification of a minor subpopulation of cells that may play a critical role in biological process over a dominant subpolulation such as a cancer and a developing organ. It provides an ultra-sensitive tool for us to clarify specific molecular mechanisms and pathways and reveal the nature of cell heterogeneity. Besides, it also empowers the clinical investigation of patients when facing a very low quantity of cell available for analysis, such as noninvasive cancer screening with circulating tumor cells (CTC), noninvasive prenatal diagnostics (NIPD) and preimplantation genetic test (PGT) for in vitro fertilization. Single-cell omics greatly promotes the understanding of life at a more fundamental level, bring vast applications in medicine. Accordingly, single-cell omics is also called as single-cell analysis or single-cell biology. Within only a couple of years, single-cell omics, especially transcriptomic sequencing (scRNA-seq), whole genome and exome sequencing (scWGS, scWES), has become robust and broadly accessible. Besides the existing technologies, recently, multiplexing barcode design and combinatorial indexing technology, in combination with microfluidic platform exampled by Drop-seq, or even being independent of microfluidic platform but using a regular PCR-plate, enable us a greater capacity of single cell analysis, switching from one single cell to thousands of single cells in a single test. The unique molecular identifiers (UMIs) allow the amplification bias among the original molecules to be corrected faithfully, resulting in a reliable quantitative measurement of omics in single cells. Of late, a variety of single-cell epigenomics analyses are becoming sophisticated, particularly single cell chromatin accessibility (scATAC-seq) and CpG methylation profiling (scBS-seq, scRRBS-seq). High resolution single molecular Fluorescence in situ hybridization (smFISH) and its revolutionary versions (ex. seqFISH, MERFISH, and so on), in addition to the spatial transcriptome sequencing, make the native relationship of the individual cells of a tissue to be in 3D or 4D format visually and quantitatively clarified. On the other hand, CRISPR/cas9 editing-based In vivo lineage tracing methods enable dynamic profile of a whole developmental process to be accurately displayed. Multi-omics analysis facilitates the study of multi-dimensional regulation and relationship of different elements of the central dogma in a single cell, as well as permitting a clear dissection of the complicated omics heterogeneity of a system. Last but not the least, the technology, biological noise, sequence dropout, and batch effect bring a huge challenge to the bioinformatics of single cell omics. While significant progress in the data analysis has been made since then, revolutionary theory and algorithm logics for single cell omics are expected. Indeed, single-cell analysis exert considerable impacts on the fields of biological studies, particularly cancers, neuron and neural system, stem cells, embryo development and immune system; other than that, it also tremendously motivates pharmaceutic RD, clinical diagnosis and monitoring, as well as precision medicine. This book hereby summarizes the recent developments and general considerations of single-cell analysis, with a detailed presentation on selected technologies and applications. Starting with the experimental design on single-cell omics, the book then emphasizes the consideration on heterogeneity of cancer and other systems. It also gives an introduction of the basic methods and key facts for bioinformatics analysis. Secondary, this book provides a summary of two types of popular technologies, the fundamental tools on single-cell isolation, and the developments of single cell multi-omics, followed by descriptions of FISH technologies, though other popular technologies are not covered here due to the fact that they are intensively described here and there recently. Finally, the book illustrates an elastomer-based integrated fluidic circuit that allows a connection between single cell functional studies combining stimulation, response, imaging and measurement, and corresponding single cell sequencing. This is a model system for single cell functional genomics. In addition, it reports a pipeline for single-cell proteomics with an analysis of the early development of Xenopus embryo, a single-cell qRT-PCR application that defined the subpopulations related to cell cycling, and a new method for synergistic assembly of single cell genome with sequencing of amplification product by phi29 DNA polymerase. Due to the tremendous progresses of single-cell omics in recent years, the topics covered here are incomplete, but each individual topic is excellently addressed, significantly interesting and beneficial to scientists working in or affiliated with this field.