Gene Knockout Protocols

1. Auflage. Softcover version of original hardcover Auflage 2001
ISBN: 978-1-61737-080-9
Verlag: Humana Press

Buch, Englisch, 431 Seiten, Previously published in hardcover, Format (B × H): 152 mm x 229 mm, Gewicht: 639 g

Reihe: Methods in Molecular Biology

As the major task of sequencing the human genome is near completion and full complement of human genes are catalogued, attention will be focused on the ultimate goal: to understand the normal biological functions of these genes, and how alterations lead to disease states. In this task there is a severe limitation in working with human material, but the mouse has been adopted as the favored animal model because of the available genetic resources and the highly conserved gene conservation linkage organization. In just of ten years since the first gene-targeting experiments were p- formed in embryonic stem (ES) cells and mutations transmitted through the mouse germline, more than a thousand mouse strains have been created. These achievements have been made possible by pioneering work that showed that ES cells derived from preimplantation mouse embryos could be cultured for prolonged periods without differentiation in culture, and that homologous rec- bination between targeting constructs and endogenous DNA occurred at a f- quency sufficient for recombinants to be isolated. In the next few years the mouse genome will be systematically altered, and the techniques for achi- ing manipulations are constantly being streamlined and improved.

Zielgruppe

Research

Autoren/Hrsg.

Kola, Ismail

Tymms, Martin J.

Fachgebiete

• Medizin | Veterinärmedizin Medizin | Public Health | Pharmazie | Zahnmedizin Vorklinische Medizin: Grundlagenfächer Humangenetik

Inhaltsverzeichnis

Overview.- Isolation and Maintenance of Primate Embryonic Stem Cells.- Gene Targeting in ES Cells.- Manipulating Mouse Embryonic Stem Cells.- Gene Targeting in a Centralized Facility.- The LoxP/CRE System and Genome Modification.- Creation and Use of a Cre Recombinase Transgenic Database.- Choice of Mouse Strains for Gene Targeting.- Isolation, Microinjection, and Transfer of Mouse Blastocysts.- Aggregation Chimeras.- How to Study Pathologic Phenotypes of Knockout Mice.- Analysis of Hematopoietic Phenotypes in Knockout Mouse Models.- Isolation of Embryonic Fibroblasts and Their Use in the In Vitro Characterization of Gene Function.- Influence of Genetic Background on Knockout Mouse Phenotypes.- Lineage Allocation During Early Embryogenesis.- Generation of Double-Knockout Embryonic Stem Cells.- In Vitro Differentiation of Embryonic Stem Cells and Analysis of Cellular Phenotypes.- Embryonic Stem Cells in the Study of Hematopoiesis.- Interferon-Inducible ES Cell Expression Systems.- Transgenic Studies in the Mouse.- In Vivo Libraries of Large Insert Transgenic Mice for Genetic Mapping.- Epigenetic Effects on Transgene Expression.- Positional-Candidate Cloning of Genes from Mouse Mutants.- Genetically Engineered Mice.- Embryo Cryopreservation for Transgenic Mouse Lines.