Zebrafish: Genetics, Genomics, and Transcriptomics

Zebrafish: Genetics, Genomics, and Transcriptomics
 
 
Academic Press
  • 4. Auflage
  • |
  • erschienen am 16. Juli 2016
  • |
  • 604 Seiten
 
E-Book | ePUB mit Adobe DRM | Systemvoraussetzungen
E-Book | PDF mit Adobe DRM | Systemvoraussetzungen
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978-0-12-803488-0 (ISBN)
 

The Zebrafish: Genetics, Genomics, and Transcriptomics, Fourth Edition, is the latest volume in the Methods in Cell Biology series that looks at methods for the analysis of genetics, genomics, and transcriptomics of Zebrafish.

Chapters cover such topics as gene-trap mutagenesis, genetic Screens for mutations, gene editing in zebrafish, homologous gene targeting, genome-wide RNA tomography, and developmental epigenetics and the zebrafish interactome.


  • Covers sections on model systems and functional studies, imaging-based approaches, and emerging studies
  • Presents chapters written by experts in the field
  • Contains cutting-edge material on the topic
0091-679X
  • Englisch
  • San Diego
  • |
  • USA
Elsevier Science
  • 10,42 MB
978-0-12-803488-0 (9780128034880)
0128034882 (0128034882)
weitere Ausgaben werden ermittelt
  • Front Cover
  • The Zebrafish: Genetics, Genomics, and Transcriptomics
  • Series Editors
  • The Zebrafish: Genetics, Genomics, and Transcriptomics
  • Copyright
  • Dedication
  • Contents
  • Contributors
  • Preface
  • 01 - Forward and Reverse Genetics
  • 1 - Multiplex conditional mutagenesis in zebrafish using the CRISPR/Cas system
  • INTRODUCTION
  • 1. METHODS
  • 1.1 Assembly of U6-Based sgRNA Transgenic Constructs
  • 1.2 Construction of Cas9 Expression Vectors
  • 1.3 Screening and Evaluation of Stable sgRNA or Cas9 Transgenic Fish
  • 2. DISCUSSION
  • SUMMARY
  • Acknowledgments
  • REFERENCES
  • 2 - Tol2-mediated transgenesis, gene trapping, enhancer trapping, and Gal4-UAS system
  • INTRODUCTION
  • 1. TRANSGENESIS BY USING THE TOL2 TRANSPOSABLE ELEMENT
  • 1.1 Rationale
  • 1.2 Methods
  • 1.2.1 Synthesis of transposase mRNA in vitro
  • 1.2.2 Preparation of a Tol2-donor plasmid
  • 1.2.3 Microinjection
  • 1.2.4 Excision assay
  • 1.2.5 Identification of transgenic fish
  • 1.3 Materials
  • 1.4 Discussion
  • 2. GENE TRAP, ENHANCER TRAP METHODS FOR THE GAL4FF-UAS METHODS
  • 2.1 Rationale
  • 2.2 Methods
  • 2.2.1 Gene trap and enhancer trap screens
  • 2.2.2 Analysis of Tol2 insertions by Southern blot hybridization
  • 2.2.3 Identification of Tol2 integration sites by inverse PCR
  • 2.2.4 Search for useful gene trap and enhancer trap fish using the zTrap database
  • 2.3 Materials
  • 2.4 Discussion
  • 3. TARGETED GENE EXPRESSION WITH THE GAL4-UAS SYSTEM
  • 3.1 Rationale
  • 3.2 Methods
  • 3.2.1 Inhibition of neuronal activities via the Gal4-UAS system
  • 3.2.2 Visualization of neuronal activities via the Gal4-UAS system
  • 3.2.3 Visualization of in vivo microtubule structures via the Gal4-UAS system
  • 3.2.4 Inhibition of the Wnt-signaling pathway via the Gal4-UAS system
  • 3.3 Materials
  • 3.4 Discussion
  • Acknowledgments
  • REFERENCES
  • 3 - Genetic screens for mutations affecting adult traits and parental-effect genes
  • INTRODUCTION
  • EXTENDING THE RANGE OF FORWARD GENETICS ANALYSIS IN ZEBRAFISH SCREENS
  • APPLICATION OF GENETIC SCREENS BEYOND THE EMBRYONIC LETHAL RANGE
  • Post-embryonic development: juvenile and adults stages
  • Prior to the midblastula transition: maternal- and paternal-effect genes
  • 1. STRATEGIES FOR ADULT TRAIT AND PARENTAL-EFFECT SCREENS
  • 1.1 F3 Screen for Adult Traits and F4 for Parental-Effect Traits Based on Natural Crosses That Integrates a Mapping Strategy
  • 1.1.1 An F3-extended family approach with integrated mapping
  • 1.1.2 Identification of maternal-zygotic, male sterile, and paternal-effect mutations
  • 1.1.3 Assessment of the F4 natural crosses screen for maternal-effect mutations
  • 1.2 F2 Adult or F3 Parental-Effect Trait Screens Based on EP-Induced Gynogenesis
  • 1.2.1 Choice of gynogenetic method
  • 1.2.2 Mutagenesis dose
  • 1.2.3 Assessment of EP-based screens for maternal-effect mutations
  • 1.3 Screening
  • 1.3.1 Screening for adult traits
  • 1.3.2 Screening embryos for parental-effects
  • 2. SELECTION OF LINES FOR GENETIC SCREENS
  • 2.1 Selection for Lethal/Sterile-Free Background Lines
  • 2.1.1 Continuous inbreeding
  • 2.1.2 Whole genome homozygosity through gynogenesis
  • 2.2 Specific Requirements for Lines in Gynogenesis-Based Screens
  • 2.2.1 Selection of lines that produce a high yield of gynogenotes
  • 2.2.2 Selection for favorable sex ratios under gynogenetic conditions
  • 2.3 A Hybrid/Inbred Approach
  • 3. RECOVERY AND MAINTENANCE OF ADULT AND PARENTAL-EFFECT MUTATIONS
  • 3.1 General Methods for Recovering Mutations
  • 3.1.1 Recovery through known heterozygous carriers
  • 3.1.2 Recovery through siblings of homozygous mutant individuals
  • 3.1.3 Recovery through rare survivors
  • 3.2 Maintenance of Adult and Parental-Effect Mutations
  • 3.2.1 Maintenance of recessive lethal adult or parental-effect mapped mutations
  • 3.2.2 Maintenance of recessive lethal adult or parental-effect unmapped mutations
  • 4. MAPPING ADULT AND PARENTAL-EFFECT MUTATIONS, AND CLONING THE MUTANT GENE
  • 4.1 Mapping and Mutant Gene Identification Using Next Generation Sequencing Technology
  • 4.2 Mapping Concomitant With F3 Adult/F4 Parental-Effect Genetic Screens
  • 4.3 Mapping After Identification and Recovery of Mutations
  • 4.4 Efficient Fine Mapping of Maternal-Effect Mutations
  • 5. SOLUTIONS, MATERIALS, AND PROTOCOLS
  • 5.1 Solutions
  • 5.2 Other Materials
  • 5.3 Protocols
  • 5.3.1 Sperm collection (adapted from D. Ransom)
  • 5.3.2 UV-inactivation of sperm
  • 5.3.3 Stripping of eggs
  • 5.3.4 In vitro fertilization
  • 5.3.5 Heat shock
  • 5.3.6 Early pressure
  • 5.3.7 Testosterone treatment
  • CONCLUSIONS
  • Acknowledgments
  • REFERENCES
  • 4 - Targeted candidate gene screens using CRISPR/Cas9 technology
  • INTRODUCTION
  • 1. GENERAL CONSIDERATIONS BEFORE STARTING A CRISPR SCREEN
  • 2. CONSIDERATIONS FOR THE CAS9 ENZYME
  • 3. CAS9 SYNTHESIS PROTOCOL
  • 4. CONSIDERATIONS FOR CHOOSING SGRNA TARGET SITES
  • 5. DESIGNING AN SGRNA
  • 6. SGRNA DESIGN AND SYNTHESIS
  • 7. SGRNA SYNTHESIS PROTOCOL
  • 8. IDENTIFYING INJECTION CONDITIONS TO USE FOR THE SCREEN
  • 9. ASSAYING THE RATE OF MUTAGENESIS IN F0-INJECTED EMBRYOS
  • 10. CONFIRMATION OF PHENOTYPES
  • 11. SUMMARY
  • REFERENCES
  • 5 - TALEN- and CRISPR-enhanced DNA homologous recombination for gene editing in zebrafish
  • INTRODUCTION
  • 1. MATERIALS
  • 1.1 Reagents
  • 1.2 Zebrafish and Microinjection
  • 2. METHODS
  • 2.1 In Vitro Synthesis of Cas9 mRNA
  • 2.2 Preparation of CRISPR gRNAs
  • 2.3 Microinjection of Cas9 mRNA/gRNA Into Zebrafish Embryos
  • 2.4 Evaluation of CRISPR Efficiency in Injected Founder Embryos
  • 2.5 Preparation of the Homologous Donor Template
  • 2.6 Microinjection of Cas9 mRNA/gRNA and Donor Template
  • 2.7 Detection of HR Events and Screening of Positive Founder Fish
  • SUMMARY
  • Acknowledgments
  • REFERENCES
  • 6 - Precise genome editing by homologous recombination
  • INTRODUCTION
  • 1. OVERVIEW OF CONTEMPORARY APPROACHES TO GENOME EDITING
  • 2. SUMMARY OF WORKFLOW
  • 3. FACTORS TO CONSIDER FOR GENOME EDITING VIA HOMOLOGOUS RECOMBINATION
  • 3.1 Choice of Nuclease Target Site
  • 3.2 Induction of Targeted DSBs
  • 3.3 Design of Donor Sequences
  • 3.4 Configuration of Donor Molecules
  • 4. METHOD OF APPROACH
  • 4.1 Design and Preparation of Programmed Nucleases
  • 4.1.1 Design of nucleases
  • 4.1.2 Target sequence confirmation and selection of a breeding population
  • 4.1.3 Synthesis of programmable nucleases
  • 4.1.4 Determination of nuclease cleavage activity
  • 4.2 Design and Preparation of Donor Molecules Using pKHR Vectors
  • 4.2.1 Preparation of homologous arms and novel donor sequences
  • 4.2.2 Generating a loxP site within a homology arm
  • 4.2.3 Assembling donor sequences into pKHR vectors
  • 4.2.4 Sequence verification of donor plasmids
  • 4.3 Microinjection of Programmed Nuclease and I-SceI-Digested Donor DNA
  • 4.4 Detection of Recombination Events
  • 4.5 Isolation of Founder Fish That Transmit Precisely Edited Genomes
  • 4.5.1 Identification of F0 founders that transmit donor DNA sequences to progeny
  • 4.5.2 Identification of founder fish carrying precisely edited genomes
  • CONCLUSIONS
  • Acknowledgments
  • REFERENCES
  • 7 - The zebrafish genome editing toolkit
  • INTRODUCTION
  • 1. REAGENTS
  • 1.1 TALENs
  • 1.2 CRISPR-Cas9
  • 1.3 Other Reagents
  • 2. DNA REPAIR MECHANISMS-THE RISE OF MUTATIONS
  • 2.1 Nonhomologous End Joining
  • 2.2 Homology-Directed Repair
  • 2.3 Microhomology-Mediated End Joining
  • 3. MICROINJECTION
  • 3.1 Injection Setup
  • 3.2 Preparation of Reagents
  • 3.3 Injection
  • 3.4 Quality Control
  • 4. SCREENING FOR F0 MUTANTS
  • 4.1 Knock-ins
  • 4.2 Knock-outs
  • 5. SELECTING FOR DESIRABLE F1 MUTANTS
  • 5.1 Outcross F0 Fish
  • 5.2 Screen for Germline Transmissibility
  • 5.3 F2 Generation and Onward
  • CONCLUSION
  • REFERENCES
  • 8 - Clonal analysis of gene loss of function and tissue-specific gene deletion in zebrafish via CRISPR/Cas9 technology
  • INTRODUCTION
  • TISSUE-SPECIFIC KNOCKOUT IN ANIMAL MODELS
  • CLONAL ANALYSIS OF GENE LOSS OF FUNCTION
  • STATE OF THE ART IN ZEBRAFISH TISSUE-SPECIFIC GENE INACTIVATION
  • CLONAL ANALYSIS OF GENE INACTIVATION IN ZEBRAFISH
  • 1. METHODS FOR TISSUE-SPECIFIC GENE INACTIVATION AND CLONAL ANALYSIS OF MUTANT CELLS
  • 1.1 Rationale for the Design of a Vector System for CRISPR/Cas9-Induced Conditional Gene Disruption via Gal4/UAS
  • 1.2 Experimental Workflow
  • 1.2.1 Overview of the method
  • 1.2.2 Technical procedure
  • 1.3 Rationale for the Design of a Vector System for Clonal Analysis of Mutant Cells by Combining the Gal4/UAS With the Cre/loxP ...
  • 1.4 Experimental Workflow
  • 1.5 Strategies for the Detection of Gene Loss of Function
  • 1.5.1 Detection of protein loss in Cas9-expressing cells
  • 1.5.1.1 Protocol for IHC on whole mount embryos
  • 1.5.1.2 Protocol for IHC on cryosections
  • 1.5.2 Molecular assessment of the mutagenesis efficiency via fluorescence-activated cell sorting and genome locus sequencing
  • 1.5.2.1 Protocol for FACS
  • 2. DISCUSSION
  • REFERENCES
  • 9 - Tissue-specific gene targeting using CRISPR/Cas9
  • INTRODUCTION
  • 1. RATIONALE
  • 1.1 Need for Tissue-Specific Tuning of Gene Expression
  • 1.2 Limitations of Existing Gene-Silencing Methods
  • 2. METHODS
  • 2.1 Identification of CRISPR Target Sites
  • 2.2 Construction of Vectors
  • 2.3 Injection, Evaluation of Mosaicism, and Phenotype Assessment
  • 3. DISCUSSION
  • 3.1 Technical Caveats and Troubleshooting
  • 3.2 Phenotypes in F0 Animals Versus Stable Lines
  • 3.3 Developments and Alternatives
  • Acknowledgments
  • REFERENCES
  • 02 - Transgenesis and Functional Genomics
  • 10 - Transcriptional regulation using the Q system in transgenic zebrafish
  • 1. BACKGROUND
  • 2. COMPONENTS OF THE Q TRANSCRIPTIONAL REGULATORY SYSTEM
  • 2.1 QF and QF Derivatives
  • 2.2 Q Upstream Activating Sequence
  • 2.3 QS and Quinic Acid
  • 3. GENE/ENHANCER TRAPPING USING THE Q SYSTEM
  • 4. FUTURE PROSPECTS
  • Acknowledgments
  • REFERENCES
  • 11 - Contemporary zebrafish transgenesis with Tol2 and application for Cre/lox recombination experiments
  • 1. INTRODUCTION
  • 1.1 Transgenesis
  • 1.2 The Cre/lox System and Inducible CreERT2
  • 1.3 Cre/lox and Zebrafish Transgenesis: Current Status
  • 2. TRANSGENE DESIGN, CLONING, AND TRANSGENESIS
  • 2.1 Basic Considerations of Transgene Design
  • 2.2 Transgenesis for Single Insertions
  • 2.3 Reagent Considerations for Transgenesis
  • 3. TOL2 TRANSGENE GENETICS
  • 3.1 "Every Fish is Unique": Basic Genetic Considerations for Tol2 Transgenesis
  • 3.2 Transgene Quality Assessment
  • 3.3 Steps and Challenges Generating Novel creERT2 and lox Lines
  • 4. CREERT2-CONTROLLED LOX RECOMBINATION USING 4-OHT
  • 4.1 4-OHT Chemistry and Handling
  • 4.2 The "Golden Rule" for Zebrafish Lineage Tracing: Paternal Cre, Maternal lox Switch
  • 4.3 4-OHT-Mediated CreERT2 Induction
  • 4.4 Induction Time Point
  • 5. BEYOND TRADITIONAL CRE/LOX LINEAGE TRACING
  • 5.1 Single-Cell Induction
  • 5.2 Multicolor Switching for Clonal Analysis
  • 5.3 Spatiotemporally Controlled Signaling Perturbations
  • 6. DISCUSSION
  • Acknowledgments
  • REFERENCES
  • 12 - Method for somatic cell nuclear transfer in zebrafish
  • INTRODUCTION
  • 1. MATERIALS
  • 2. METHOD
  • 2.1 Fish Breeding
  • 2.2 Preparation of Reagents
  • 2.3 Set up the Micromanipulator
  • 2.4 Egg Collection
  • 2.5 Donor Cell Preparation
  • 2.6 Micromanipulation
  • 2.7 Activation of Reconstructed Eggs
  • 2.8 In Vitro Fertilization at the End of the Somatic Cell Nuclear Transfer Procedure
  • 2.9 Confirmation That a "Clone" Is a Real Clone
  • Acknowledgments
  • REFERENCES
  • 13 - Experimental approaches to studying the nature and impact of splicing variation in zebrafish
  • 1. OVERVIEW OF RNA SPLICING
  • 1.1 Spliceosome: Composition and Function
  • 1.2 Regulation of Splice Site Selection
  • 1.3 Intron Retention
  • 1.4 Splicing Dysfunction and Disease
  • 2. EMPLOYING ZEBRAFISH MODELS FOR STUDYING SPLICING VARIATION
  • 2.1 Zebrafish Genomics and Reverse Genetics
  • 2.2 Biological Insights from Zebrafish SF mutants
  • 2.3 Zebrafish Models of Human Diseases due to SF Dysfunction
  • 2.3.1 Retinitis pigmentosa
  • 2.3.2 Spinal muscular atrophy
  • 2.3.3 Other diseases
  • 3. TOOLS FOR GLOBAL SPLICEOME ANALYSIS IN ZEBRAFISH
  • 3.1 RNAseq
  • 3.2 RT-PCR Validation
  • 3.3 In Vivo Minigene Assay
  • 4. BIOCHEMICAL ANALYSIS OF SPLICEOSOME COMPONENT ASSEMBLY
  • 4.1 Glycerol Gradient Sedimentation
  • 4.2 Northern Analysis
  • 5. ASSESSING THE BIOLOGICAL IMPACT OF SPLICING VARIATION OR DYSFUNCTION IN ZEBRAFISH
  • 5.1 Zebrafish as a Model to Develop Therapies Directed at Disease Correction by Targeting Splicing
  • 5.2 Using Zebrafish to Search for Drugs That Modulate Splicing
  • 6. SUMMARY
  • Acknowledgments
  • REFERENCES
  • 14 - PICCORO: A technique for manipulating the activity of transcription factors with blue light
  • INTRODUCTION
  • 1. MECHANISM OF PICCORO
  • 2. METHOD FOR CONSTRUCTING A PICCORO PLASMID
  • 2.1 Construction of a Chimeric Transcription Factor
  • 2.2 Evaluating the Appropriate Amount of mRNA for Transformation
  • 2.3 Injecting PixE-geneXR mRNA into Eggs of Tg(EF1a:PixD)
  • CONCLUSION
  • REFERENCES
  • 03 - Transcriptomics
  • 15 - Tomo-seq: a method to obtain genome-wide expression data with spatial resolution
  • INTRODUCTION
  • 1. TISSUE EXTRACTION AND EMBEDDING
  • 2. CRYOSECTIONING AND TRIZOL EXTRACTION
  • 3. CDNA SYNTHESIS
  • 4. IN VITRO TRANSCRIPTION
  • 5. LIBRARY PREPARATION
  • 6. DATA ANALYSIS
  • SUMMARY AND CONCLUSIONS
  • REFERENCES
  • 16 - Cell type-specific transcriptomic analysis by thiouracil tagging in zebrafish
  • INTRODUCTION
  • PART 1. ESTABLISHING AND CHARACTERIZING A CELL TYPE-RESTRICTED UPRT TRANSGENIC LINE OF FISH
  • 1.1 Creating a Cell Type-Specific UPRT Transgenic
  • 1.2 Using RNA Dot Blots to Characterize UPRT Transgenics and Optimize 4TU Treatment Conditions
  • 1.3 Dot Blot Materials
  • 1.4 Dot Blot Procedure (Estimated Time ~2Days)
  • PART 2. EXPERIMENTAL DESIGN AND 4TU TREATMENTS
  • 2.1 Experimental Design: Type I Versus Type II Experiments
  • 2.2 Biological Replicates
  • 2.3 Developmental Staging
  • 2.4 Number of Larvae Required for TU-Tagging
  • PART 3. RNA MANIPULATIONS, BIOTINYLATION, AND PURIFICATION
  • 3.1 Removing Ribosomal RNA
  • 3.2 RNA Fragmentation
  • 3.3 Biotinylation and Purification of TU-Tagged RNA
  • PART 4: RNA SEQUENCING, DATA ANALYSIS, AND VALIDATION
  • 4.1 Library Preparation and RNA Sequencing
  • 4.2 Data Analysis
  • 4.3 Validation of the RNA-seq Results
  • CONCLUSIONS
  • REFERENCES
  • 17 - RT-qPCR gene expression analysis in zebrafish: preanalytical precautions and use of expressed repetitive eleme ...
  • INTRODUCTION
  • 1. SAMPLE COLLECTION, RNA EXTRACTION AND QUALITY CONTROL, AND REVERSE TRANSCRIPTION
  • 1.1 Method A: Sample Collection Zebrafish Embryos and Tissues
  • 1.1.1 Reagents
  • 1.1.2 Protocol
  • 1.2 Method B: Tissue Homogenization Using the TissueLyser
  • 1.2.1 Reagents/devices
  • 1.2.2 Protocol
  • 1.3 Method C: Tissue Homogenization Using Cryotome
  • 1.3.1 Reagents
  • 1.3.2 Protocol
  • 2. ASSAY DESIGN AND QUALITY CONTROL
  • 2.1 Method D: Assessment of PCR Efficiency Using Standard Curve Analysis
  • 2.1.1 Reagents
  • 2.1.2 Protocol
  • 2.1.3 Notes
  • 3. REFERENCE TARGET VALIDATION
  • 4. EXPRESSED REPEAT RT-QPCR NORMALIZATION IN ZEBRAFISH
  • REFERENCES
  • 04 - Epigenetics
  • 18 - Genome-wide DNA methylation profiling in zebrafish
  • INTRODUCTION
  • METHODS FOR STUDYING DNA METHYLATION IN ZEBRAFISH
  • SIGNIFICANCE-DNAME IN HUMAN DEVELOPMENT AND DISEASE
  • WRITERS, ERASERS, AND READERS OF DNAME IN ZEBRAFISH
  • DNAME REPROGRAMMING DURING VERTEBRATE DEVELOPMENT-A COMPARISON OF MAMMALS AND ZEBRAFISH
  • 1. RATIONALE
  • 2. METHODS
  • 2.1 Extraction and Fragmentation of Genomic DNA
  • 2.2 Methylated DNA Immunoprecipitation and Sequencing
  • 2.3 Whole Genome Shotgun Bisulfite Sequencing
  • 2.4 Reduced Representation Bisulfite Sequencing
  • DISCUSSION AND CONCLUSIONS
  • REFERENCES
  • 19 - Histone modifications in zebrafish development
  • INTRODUCTION
  • ROLES FOR CHROMATIN MODIFICATIONS IN THE EPIGENETIC REGULATION OF GENE EXPRESSION
  • TYPES OF HISTONE MODIFICATIONS: PRODUCTION, RECOGNITION, AND REMOVAL
  • THE IMPACTS OF HISTONE MODIFICATIONS ON GENE TRANSCRIPTION
  • 1. RECENT PROGRESS IN UNDERSTANDING THE ROLES OF HISTONE MODIFICATIONS IN ZEBRAFISH DEVELOPMENT
  • 1.1 Chromatin Modifications at the Promoters of Zygotically Active Genes
  • 1.2 Interplay Between Histone Modifications and Chromatin Remodeling Machinery
  • 1.3 Association of Covalently Modified Histones With Developmentally Regulated Enhancer Sequences
  • 2. CHROMATIN IMMUNOPRECIPITATION FROM ZEBRAFISH EMBRYOS
  • 2.1 Rationale
  • 2.2 Chromatin Immunoprecipitation Method
  • 2.2.1 Dechorionation of embryos and cross-linking of chromatin with EGS
  • 2.2.2 Cross-linking of chromatin with formaldehyde
  • 2.2.3 Deyolking of embryos
  • 2.2.4 Chromatin preparation from embryos
  • 2.2.5 Sonication
  • 2.2.6 Sonication test
  • 2.2.7 Incubation of chromatin extract with antibodies
  • 2.2.8 Chromatin-antibody complex washes
  • 2.2.9 Purification of immunoprecipitated DNA
  • 2.2.10 Buffers and solutions
  • 3. HIGH-THROUGHPUT WHOLE-GENOME DNA SEQUENCING AND INTEGRATIVE ANALYSIS OF EPIGENOMIC DATA
  • CONCLUSIONS AND PROSPECTS
  • Acknowledgments
  • REFERENCES
  • 20 - Chromatin immunoprecipitation and an open chromatin assay in zebrafish erythrocytes
  • INTRODUCTION
  • 1. RATIONALE
  • 2. ADULT ZEBRAFISH EXSANGUINATION
  • 3. CHROMATIN IMMUNOPRECIPITATION AND CHIP-SEQ LIBRARY CONSTRUCTION (LEE, JOHNSTONE, & YOUNG, 2006)
  • 3.1 Cross-Linking
  • 3.2 Sonication
  • 3.3 Pretreatment of Beads and Antibodies
  • 3.4 Chromatin Immunoprecipitation
  • 3.5 Chromatin Immunoprecipitation Washes
  • 3.6 Elution and Reversal of Cross-Linking
  • 3.7 DNA Recovery and Quality Control
  • 3.8 ChIP-seq Library Preparation
  • 4. ATAC-SEQ ANALYSIS OF OPEN CHROMATIN REGIONS
  • 4.1 Cell Treatment and Nucleus Isolation
  • 4.2 Transposition of Open Chromatin Regions
  • 4.3 DNA Isolation and Library Preparation
  • 4.4 Library Amplification and Indexing
  • 4.5 Library Quality Control and Quantification
  • 4.6 Sequencing Considerations
  • 5. CHIP-SEQ AND ATAC-SEQ DATA ANALYSIS
  • 6. RESULTS
  • SUMMARY AND CONCLUSION
  • Acknowledgments
  • REFERENCES
  • 21 - Assay for transposase-accessible chromatin and circularized chromosome conformation capture, two methods to ex ...
  • INTRODUCTION
  • 1. 4C-SEQ PROTOCOL
  • 1.1 Cell Isolation
  • 1.1.1 Day 1
  • 1.1.1.1 Zebrafish embryos
  • 1.1.1.2 Medaka embryos
  • 1.2 Chromatin Cross-linking
  • 1.3 Cell Lysis
  • 1.4 Digestion and Ligation of the Cross-linked DNA
  • 1.4.1 Day 2
  • 1.4.2 Day 3
  • 1.5 Reverse Cross-linking and Precipitation
  • 1.5.1 Day 4
  • 1.6 Second Round of Digestion and Ligation
  • 1.6.1 Day 5
  • 1.7 Sample Purification
  • 1.7.1 Day 6
  • 1.8 Preparation of 4C Libraries for Illumina Sequencing
  • 2. ATAC-SEQ PROTOCOL
  • 2.1 Cell Lysis
  • 2.1.1 Zebrafish
  • 2.1.2 Medaka
  • 2.2 Tagmentation
  • 2.3 Amplification and Sequencing of Tagmentation Libraries
  • Acknowledgments
  • REFERENCES
  • 22 - Epigenetic regulation of hematopoietic stem cell development
  • INTRODUCTION
  • 1. MECHANISMS OF EPIGENETIC REGULATION
  • 1.1 5-Methylcytosine
  • 1.2 5-Hydroxymethylcytosine
  • 1.3 Histone Modification
  • 2. HEMATOPOIETIC STEM CELL DEVELOPMENT
  • 3. APPROACHES
  • 3.1 Unbiased Forward Genetic Approaches
  • 3.2 Reverse Genetic Approaches
  • 3.3 Candidate Screens
  • 4. DISEASE MODELS
  • 5. CURRENT CHALLENGES AND FUTURE DIRECTIONS
  • 5.1 Tools for Unbiased Screening of Epigenetic States In Vivo
  • 5.2 Tools for Tissue-Specific Mutation
  • CONCLUSIONS
  • REFERENCES
  • 05 - Zebrafish Infrastructure
  • 23 - A scientist's guide for submitting data to ZFIN
  • INTRODUCTION
  • 1. WHY LOAD DATA INTO ZFIN?
  • 2. THE STRUCTURE OF THE ZFIN DATABASE
  • 3. THE DATA SUBMISSION PROCESS
  • 4. DATA SUBMISSIONS
  • 4.1 Mutant and Transgenic Line Submission
  • 4.1.1 Genotypes
  • 4.1.2 Genomic feature
  • 4.1.3 Feature zygosity
  • 4.1.4 Feature maternal and paternal zygosity
  • 4.1.5 Genetic background
  • 4.1.6 Affected gene symbol
  • 4.1.7 Affected gene accession
  • 4.1.8 Transgene type
  • 4.1.9 Mutation type
  • 4.1.10 Mutagen
  • 4.1.11 Subject
  • 4.1.12 Construct
  • 4.1.13 Laboratory of origin
  • 4.1.14 Sequence accession
  • 4.1.15 Link to alternate resource
  • 4.1.16 Citations
  • 4.1.17 Note
  • 4.1.18 Sperm samples
  • 4.2 Transgenic Constructs
  • 4.2.1 Construct name
  • 4.2.2 Promoter gene symbol
  • 4.2.3 Promoter gene accession
  • 4.2.4 Coding sequence gene symbol
  • 4.2.5 Coding sequence gene accession
  • 4.2.6 Engineered region name
  • 4.2.7 Construct sequence accession
  • 4.2.8 Construct map image name
  • 4.2.9 Link to alternate construct resource
  • 4.2.10 Citation
  • 4.2.11 Construct note
  • 4.3 Morpholinos, TALENs, and CRISPRs
  • 4.3.1 MO/TALEN/CRISPR name
  • 4.3.2 Target sequence 1
  • 4.3.3 Target sequence 2
  • 4.3.4 Target gene symbol
  • 4.3.5 Target gene accession
  • 4.3.6 Link to alternate resource
  • 4.3.7 Citations
  • 4.4 Expression Data
  • 4.4.1 Expressed gene symbol
  • 4.4.2 Expressed gene accession
  • 4.4.3 Genotype
  • 4.4.4 Morpholinos, TALENs, and CRISPRs
  • 4.4.5 Anatomical structure
  • 4.4.6 Developmental stage
  • 4.4.7 Experimental conditions
  • 4.4.8 Experimental conditions note
  • 4.4.9 Citations
  • 4.4.10 Assay type
  • 4.4.11 Antibody name
  • 4.4.12 Probe GenBank accession number
  • 4.4.13 Images and movies
  • 4.5 Phenotype Data
  • 4.5.1 Genotype
  • 4.5.2 Morpholinos, TALENS, and CRISPRs
  • 4.5.3 Developmental stage
  • 4.5.4 Experimental condition
  • 4.5.5 Phenotype entity
  • 4.5.6 Phenotype quality
  • 4.5.7 Tag
  • 4.5.8 Media file name
  • 4.5.9 Citations
  • 4.6 Genome Browser Tracks
  • 4.6.1 Track files
  • 4.6.2 Track configuration and description
  • 4.6.3 Citation
  • 4.6.4 Track maintenance
  • 4.7 Disease Models
  • 4.7.1 Genotype
  • 4.7.2 Experimental condition
  • 4.7.3 Morpholino, TALEN, CRISPR
  • 4.7.4 Disease term ID
  • 4.7.5 Citation
  • 5. WHAT HAPPENS TO YOUR DATA AFTER SUBMISSION TO ZFIN?
  • 5.1 Gene Identification
  • 5.2 Morpholino, TALEN, CRISPR Identification and Target Validation
  • 5.3 Anatomy Term and Stage Validation
  • 5.4 EQ Syntax Validation
  • 5.5 Antibody Identification
  • 6. DATA SUBMISSION TEMPLATES
  • FUNDING INFORMATION
  • REFERENCES
  • 24 - New frontiers for zebrafish management
  • INTRODUCTION
  • 1. PHYSICOCHEMICAL ENVIRONMENT
  • 2. NUTRITION AND FEEDING
  • 3. BREEDING
  • 4. GENETIC MANAGEMENT
  • 5. LARVICULTURE
  • 6. HEALTH MANAGEMENT
  • 7. HOUSING/AQUACULTURE
  • 8. WELFARE
  • 9. EDUCATION AND TRAINING
  • 10. COMMERCIAL PROVISION OF MODEL INFRASTRUCTURAL SERVICES
  • SUMMARY
  • REFERENCES
  • 25 - Aquaculture, husbandry, and shipping at the Zebrafish International Resource Center
  • INTRODUCTION
  • 1. METHODS
  • 1.1 ZIRC's Aquatic Infrastructure
  • 1.1.1 Overview
  • 1.1.2 Water source and volume
  • 1.1.3 Conductivity and buffering
  • 1.1.4 Water quality and composition
  • 1.1.5 Filtration systems
  • 1.1.6 Ultraviolet water sterilization
  • 1.1.7 System monitoring
  • 1.2 Tanks, Rack System, and Fish Maintenance
  • 1.2.1 ZIRC tanks and rack systems
  • 1.2.2 Tank cleaning
  • 1.2.3 Fish facility cleaning
  • 1.2.4 Breeding fish and collecting embryos
  • 1.3 Raising Larvae
  • 1.3.1 Tank system, water, and cleaning
  • 1.3.2 Nursery schedule
  • 1.4 Live Food Production
  • 1.4.1 Live food types
  • 1.4.2 Juvenile and adult feeding schedules
  • 1.5 Line Management Tools
  • 1.5.1 Record keeping
  • 1.5.2 Cryopreservation
  • 1.6 Shipping Fish
  • 1.6.1 Packaging of embryos
  • 1.6.2 Packaging of adults
  • 1.6.3 General packaging and labeling
  • 1.6.4 Shipping documentation
  • 1.6.5 Zebrafish import to the United States
  • 2. MATERIALS
  • Acknowledgments
  • REFERENCES
  • 26 - Health monitoring and disease prevention at the Zebrafish International Resource Center
  • INTRODUCTION
  • 1. METHODS
  • 1.1 Assessing Colony Health
  • 1.1.1 Moribund fish
  • 1.1.2 Sentinel fish
  • 1.1.3 Wild-type lines
  • 1.1.4 Additional sampling
  • 1.1.5 Diagnostic tests and results
  • 1.2 Preventing Pathogen Spread Within the Colony
  • 1.2.1 UV sterilizers
  • 1.2.2 Husbandry
  • 1.2.3 Tank cleaning
  • 1.2.4 Surface and equipment disinfection
  • 1.2.5 Database tracking and record keeping
  • 1.3 Preventing Entry of New Pathogens
  • 1.3.1 Request of health history from submitting laboratory
  • 1.3.2 Quarantine room
  • 1.3.3 Lines arriving as cryopreserved sperm
  • 1.3.4 People
  • 2. MATERIALS
  • 2.1 Assessing Colony Health
  • 2.2 Preventing Pathogen Spread Within the Colony
  • 2.3 Preventing Entry of New Pathogens
  • REFERENCES
  • Volumes in Series
  • Index
  • A
  • B
  • C
  • D
  • E
  • F
  • G
  • H
  • I
  • L
  • M
  • N
  • O
  • P
  • Q
  • R
  • S
  • T
  • U
  • W
  • X
  • Z
  • Back Cover

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