A Large-Scale, Gene-Driven Mutagenesis Approach for the Functional Analysis of the Mouse Genome

A major challenge of the postgenomic era is the functional characterization of every single gene within the mammalian genome. In an effort to address this challenge, we assembled a collection of mutations in mouse embryonic stem (ES) cells, which is the largest publicly accessible collection of such...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2003-08, Vol.100 (17), p.9918-9922
Main Authors: Hansen, Jens, Floss, Thomas, Van Sloun, Petra, Füchtbauer, Ernst-Martin, Vauti, Franz, Arnold, Hans-Hennig, Schnütgen, Frank, Wurst, Wolfgang, von Melchner, Harald, Ruiz, Patricia
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Biological Sciences
Cell Line
Complementary DNA
Disease models
DNA - genetics
Exons
Genes
Genetic mutation
Genetic Vectors
Genomes
Genomics - methods
Humans
Mice - genetics
Mice, Inbred C57BL
Mice, Transgenic
Mutagenesis, Insertional - methods
Mutation
Phenotype
Phenotypes
Rodents
Sequence Tagged Sites
Splicing
Stem Cells
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Summary:A major challenge of the postgenomic era is the functional characterization of every single gene within the mammalian genome. In an effort to address this challenge, we assembled a collection of mutations in mouse embryonic stem (ES) cells, which is the largest publicly accessible collection of such mutations to date. Using four different gene-trap vectors, we generated 5,142 sequences adjacent to the gene-trap integration sites (gene-trap sequence tags; http://genetrap.de) from >11,000 ES cell clones. Although most of the gene-trap vector insertions occurred randomly throughout the genome, we found both vector-independent and vector-specific integration "hot spots." Because >50% of the hot spots were vector-specific, we conclude that the most effective way to saturate the mouse genome with gene-trap insertions is by using a combination of gene-trap vectors. When a random sample of gene-trap integrations was passaged to the germ line, 59% (17 of 29) produced an observable phenotype in transgenic mice, a frequency similar to that achieved by conventional gene targeting. Thus, gene trapping allows a large-scale and cost-effective production of ES cell clones with mutations distributed throughout the genome, a resource likely to accelerate genome annotation and the in vivo modeling of human disease.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1633296100