Microinjection And Transgenesis

The establishment of microinjection protocols about 20 years ago for cultured cells and shortly thereafter for the generation of transgenic mice by microinjection of DNA into fertilized mouse eggs greatly influ enced many fields of biology. Not only have the data generated using these approaches contributed to a large extent to our present under standing of gene regulation and cellular function of higher eukaryotic cells, but current knowledge and future developments in this area will certainly have a great impact on basic and applied research for many years to come. This laboratory manual describes the current state of the art in this research area and focuses primarily on both the experimental strategies with an extensive bibliography and the detailed procedures. A large number of studies are presently being performed and a great variety of new experimental designs are rapidly being developed. The book con tains protocols on injection of somatic cells as well as on injection of embryos, the use of similar equipment being a common feature. In the articles dedicated to somatic cells, full descriptions of the manual and automatic injection systems are given as well as the methods for the analysis of injected cells by video-microscopy, electron microscopy or in situ hybridizations. In addition, comprehensive protocols are given for injection experiments with very different purposes, such as to study sig nal transduction or microtubule dynamics.

Transgenic animal technologies and the ability to introduce functional genes into animals have revolutionized our ability to address complex biomedical and biological questions. This well-illustrated handbook covers the technical aspects of gene transfer — from molecular methods to whole animal considerations — for important laboratory and domestic animal species. It describes methodologies as employed by leading laboratories and is a key resource for researchers, as well as a tool for training technicians and students. This second edition incorporates updates on a variety of genetic engineering technologies ranging from microinjection and ES cell transfer to nuclear transfer in a broad range of animal modeling systems. Contains a comprehensive collection of transgenic animal and gene transfer methods Discusses background and introduction to techniques and animal systems Teaches practical step-by-step protocols Fully revised with updates to reflect state-of-the-art technology and associated changes to date

This detailed book explores how microinjection will be used in the foreseeable future, not only for generating animal models for biomedical research but also for changing economically or ecologically important species that can broadly impact our society in general. The opening half of the book focuses on methods for generating mouse models, as they are still the most popular in genome engineering research, while the second half examines gene-editing in a variety of other species, opened up by the developments in ZFN, TALEN, and CRISPR techniques. Written for the highly successful Methods in Molecular Biology series, 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 practical, Microinjection: Methods and Protocols serves as an ideal guide for researchers looking to take advantage of the breakthrough technologies in gene-editing and embryo micromanipulations.

This laboratory manual, published in cooperation with the International Society for Transgenic Technology (ISTT), provides almost all current methods that can be applied to the creation and analysis of genetically modified animals. The chapters have been contributed by leading scientists who are actively using the technology in their laboratories. Based on their first-hand experience the authors also provide helpful notes and troubleshooting sections. Topics range from standard techniques, such as pronuclear microinjection of DNA, to more sophisticated and modern methods, such as the derivation and establishment of embryonic stem (ES) cell lines, with defined inhibitors in cell culture medium. In addition, related topics with relevance to the field are addressed, including global web-based resources, legal issues, colony management, shipment of mice and embryos, and the three R’s: refinement, reduction and replacement.

Assists policymakers in evaluating the appropriate scientific methods for detecting unintended changes in food and assessing the potential for adverse health effects from genetically modified products. In this book, the committee recommended that greater scrutiny should be given to foods containing new compounds or unusual amounts of naturally occurring substances, regardless of the method used to create them. The book offers a framework to guide federal agencies in selecting the route of safety assessment. It identifies and recommends several pre- and post-market approaches to guide the assessment of unintended compositional changes that could result from genetically modified foods and research avenues to fill the knowledge gaps.

The threespine stickleback is a small fish of temperate coastal and fresh waters that exhibits extraordinary phenotypic diversity. Benefiting from its amenability to observation in the field and manipulation in the laboratory, Niko Tinbergen pioneered the threespine stickleback's use in behavioral studies and established it as a model system in ethology. This up-to-date volume incorporates reviews from active researchers who use studies of the fish to address a broad variety of evolutionary issues, including optimal foraging, armor variation, speciation, and the endocrine basis for correlated behavioral characters. The work demonstrates the value of viewing the biology of a single organism simultaneously from multiple perspectives. Students and researchers in ecology, evolution, animal behavior, and vertebrate zoology will find much of interest in this useful book.

This manual describes the methodology and applications of microinjection, a powerful technique for transposing material into living cells through needle injection. Information is presented on the techniques principles, the required equipment, preparation of receiving cells and injected material, and the analysis of results. Detailed protocols for a wide range of applications in different cellular systems are included. A large part of the book is also dedicated to the Xenopus laevis oocyte system.