The Zebrafish: Genetics, Genomics and Informatics

Contributors

Preface

Chapter 1: Generating Conditional Mutations in Zebrafish Using Gene-trap Mutagenesis

I. Introduction

II. Rationale for Conditional Mutations

III. Rationale for Gene-trap Mutagenesis to Generate Conditional Mutations

IV. Rationale for Vector Design

V. Methods

VI. Discussion

VII. Summary

Chapter 2: Tol2-mediated Transgenesis, Gene Trapping, Enhancer Trapping, and the Gal4-UAS System

I. Introduction

II. Transgenesis by Using the Tol2 Transposable Element

III. Tol2-mediated BAC Transgenesis

IV. Gene Trapping and Enhancer Trapping with the Tol2 Transposon System

V. Gene Trapping and Enhancer Trapping with the Gal4FF-UAS System

Chapter 3: Engineering Zinc Finger Nucleases for Targeted Mutagenesis of Zebrafish

I. Introduction

II. Methods and Resources for Engineering Zinc Finger Nucleases

III. Choice of Method to Engineer Zinc Finger Nucleases

Chapter 4: Retroviral-mediated Insertional Mutagenesis in Zebrafish

I. Introduction

II. Mutagenesis

III. Cloning the Mutated Genes

IV. Retroviral-mediated Insertional Mutagenesis as a Reverse Genetic Tool

V. Future Directions

Chapter 5: Genetic Screens for Mutations Affecting Adult Traits and Parental-effect Genes

I. Introduction

II. Strategies for Adult Trait and Parental-effect Screens

III. Selection of Lines for Genetic Screens

IV. Recovery and Maintenance of Adult and Parental-effect Mutations

V. Mapping Adult and Parental-effect Mutations

VI. Solutions, Materials, and Protocols

VII. Conclusions

Chapter 6: High-Throughput Target-Selected Gene Inactivation in Zebrafish

I. Introduction

II. Mutagenesis and Library Creation

III. PCR and Re-sequencing of Exons

IV. Analysis of Re-sequencing Data and Retrieval of Mutations

Chapter 7: Genetic Suppressor Screens in Haploids

I. Introduction and Rationale

II. Methods – an Example of Suppressor Screen in Moonshine Mutants

III. Conclusions and Prospective

Chapter 8: Transgenic Zebrafish Using Transposable Elements

I. Introduction

II. Requesting and Assembling Tol2 System

III. Microinjection

IV. Quality Control

V. Select Injected Fish to Raise as Founder Generation F0

VI. Production of F1 Generation

VII. Select the Best Substrain(s) Based on Observation of F2 Generation

VIII. Discussion

Chapter 9: Spatiotemporal Control of Embryonic Gene Expression Using Caged Morpholinos

I. Introduction

II. Design and Synthesis of cMOs

III. Synthesis of cFD

IV. Microinjection of Caged Reagents

V. Global Photoactivation of Caged Reagents

VI. Titration of cMO Dose for Optimum Dynamic Range

VII. Localized Photoactivation of Caged Reagents

VIII. Conclusion

Chapter 10: Advanced Zebrafish Transgenesis with Tol2 and Application for Cre/lox Recombination Experiments

I. Introduction

II. Transgene Design, Cloning, and Tol2 Transgenesis

III. Tol2 Transgene Genetics

IV. CreERT2-Controlled lox Recombination Using 4-OHT

V. Discussion

Conflict of interest statement

Chapter 11: Use of Phage PhiC31 Integrase as a Tool for Zebrafish Genome Manipulation

I. Introduction

II. Rationale

III. Materials and Methods

IV. Discussion

V. Summary

Chapter 12: Method for Somatic Cell Nuclear Transfer in Zebrafish

I. Methods

II. Materials

Chapter 13: Single Nucleotide Polymorphism (SNP) Panels for Rapid Positional Cloning in Zebrafish

I. Introduction

II. Extraction of Zebrafish Genomic DNA

III. Affymetrics Molecular Inversion Probe Panel

IV. Suggested Mapping Strategy

V. General Considerations

Chapter 14: Molecular Cytogenetic Methodologies and a BAC Probe Panel Resource for Genomic Analyses in the Zebrafish

I. Introduction

II. Cytogenetic Methods

III. Application of Cytogenetic Methods in Zebrafish Studies

IV. Methods for Array CGH

V. Application of Array CGH in Zebrafish Studies

VI. Summary

Chapter 15: Conserved Synteny and the Zebrafish Genome

I. Introduction

II. Rationale: Gene Duplication and Orthology Assignments

III. Methods

IV. Using the Synteny Database: A Case Study of Nerve Growth Factor Receptors

V. Zebrafish, Conserved Syntenies, and the Teleost Genome Duplication

VI. Summary

Chapter 16: The Zon Laboratory Guide to Positional Cloning in Zebrafish

I. Introduction

II. Mapping Strains

III. Families and Genetic Markers

IV. Crosses for Line Maintenance and Mapping

V. Preparation of the DNA

VI. Mapping Genes

VII. General Flow of Information from the Zebrafish Genome Sequence Assemblies

VIII. Synteny between Human, Zebrafish, Fugu, and Tetraodon Genomes

IX. Proving a Candidate Gene is Responsible for the Mutant Phenotype

X. Morpholinos

XI. Future Technologies

Chapter 17: Data Extraction, Transformation, and Dissemination through ZFIN

I. Introduction

II. Extracting Data and Data Flow

III. Ensuring Accurate and Rapid Data Dissemination

IV. Submitting Data Directly to ZFIN

V. Conclusions

Chapter 18: DNA Methylation Profiling in Zebrafish

I. Introduction

II. Rationale

III. Methods

IV. Discussion

Chapter 19: Chromatin Immunoprecipitation in Adult Zebrafish Red Cells

I. Introduction

II. Adult Zebrafish Exsanguination

III. Chromatin Immunoprecipitation (Lee et al., 2006)

IV. ChIP-seq Sample Preparation (Guenther et al., 2008)

V. ChIP-seq Data Analysis

VI. Summary and Conclusions

VII. Online tools

VIII. Conflicts of Interest

Chapter 20: Discerning Different In vivo Roles of MicroRNAs by Experimental Approaches in Zebrafish

I. Introduction

II. Using Zebrafish to Study the Function of miRNAs

III. Summary

Chapter 21: Sequencing-based Expression Profiling in Zebrafish

I. Introduction

II. Digital Gene Expression

III. Potential Issues in DGE

Chapter 22: Chromatin Modification in Zebrafish Development

I. Chromatin Modifications

II. Chromatin Modifications in Development

III. Functions of Chromatin Modifications in Tissue Regeneration

IV. Outlook

Chapter 23: Advances in Zebrafish Husbandry and Management

I. Introduction

II. Breeding

III. Larviculture

IV. Aquaculture Systems

V. Nutrition

VI. Health Management

VII. Genetic Management

VIII. Regulatory Compliance

IX. Education and Training

X. Summary

Chapter 24: Aquaculture and Husbandry at the Zebrafish International Resource Center

I. Introduction

II. Methods

III. Materials

Index

Volumes in Series