Gene Targeting Vector Design for Embryonic Stem Cell Modifications

Protocol
Part of the Springer Protocols Handbooks book series (SPH)

Abstract

The use of genetically engineered mice to understand gene function is widespread. Changes to the mouse genome can be introduced with gene targeting vectors or with transgenes. Targeting vectors are usually used to ablate gene expression while transgenes are designed to express proteins that are normally absent from the organism. For example, gene targeting in mouse embryonic stem cells can be used to generate a mutant mouse model that fails to express a physiologically important protein. Transgenes that express the missing protein or a substitute for the missing protein can be used to assess possible gene therapies for the mutant mouse. Both gene targeting and transgene approaches can be used to study regulatory elements that control gene function. Putative control elements can be added to or removed from the chromosome with targeting vectors. Transgenes carrying long DNA sequences that include different combinations of potential control elements can be introduced into the genome to assess their effects on gene expression. The exploration of how genes interact to control development, homeostasis, and pathophysiological conditions can be dissected by introducing carefully designed genetic constructs into model organisms.

Abbreviations

BAC

Bacterial artificial chromosome

ES cell

Embryonic stem cell

IKMC

International Mouse Knockout Consortium

Kb

Kilo base pairs of DNA

PGKneo

DNA cassette featuring neomycin phosphotransferase II expression controlled by the phosphoglycerol kinase 1 promoter/enhancer

ZFN

Zinc finger nuclease

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Copyright information

© Springer-Verlag Berlin Heidelberg 2011

Authors and Affiliations

  1. 1.Transgenic Animal Model CoreUniversity of Michigan Medical SchoolAnn ArborUSA
  2. 2.Division of Molecular Medicine and Genetics, Department of Internal MedicineUniversity of Michigan Medical SchoolAnn ArborUSA

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