Mutagenesis of Diploid Mammalian Genes by Gene Entrapment.
From: Department of Microbiology and Immunology, Room AA4206, Medical Center North, Vanderbilt University, School of Medicine 1161 21st Avenue South, AA4210, Nashville, TN 37232-2363, USA.
Nucleic acids research
- Publish Date: 2006
- ISSN: 1362-4962
- Volume: 34
- Issue: 20
- Pages: e139
- Medium: Internet
- Language: English
- Citation (JAMA): Lin Qing, Donahue Sarah L, Moore-Jarrett Tracy, et al. Mutagenesis of Diploid Mammalian Genes by Gene Entrapment.. Nucleic Acids Res. 2006;34:e139
Abstract
The present study describes a genome-wide method for biallelic mutagenesis in mammalian cells. Novel poly(A) gene trap vectors, which contain features for direct cloning vector-cell fusion transcripts and for post-entrapment genome engineering, were used to generate a library of 979 mutant ES cells. The entrapment mutations generally disrupted gene expression and were readily transmitted through the germline, establishing the library as a resource for constructing mutant mice. Cells homozygous for most entrapment loci could be isolated by selecting for enhanced expression of an inserted neomycin-resistance gene that resulted from losses of heterozygosity (LOH). The frequencies of LOH measured at 37 sites in the genome ranged from 1.3 x 10(-5) to 1.2 x 10(-4) per cell and increased with increasing distance from the centromere, implicating mitotic recombination in the process. The ease and efficiency of obtaining homozygous mutations will (i) facilitate genetic studies of gene function in cultured cells, (ii) permit genome-wide studies of recombination events that result in LOH and mediate a type of chromosomal instability important in carcinogenesis, and (iii) provide new strategies for phenotype-driven mutagenesis screens in mammalian cells.
Mesh Headings (Keywords): Animals, Base Sequence, Cell Line, Diploidy, Embryonic Stem Cells, Gene Targeting, Genetic Vectors, Genomics, Loss of Heterozygosity, Mice, Molecular Sequence Data, Mutagenesis, Sequence Tagged Sites
Check for Full Text / PubMed Unique Identifier (PMID): 17062627
This abstract is part of PubMed, a service of the U.S. National Library of Medicine. PubMed includes more than 17 million citations from MEDLINE and other life science journals for biomedical articles. See Copyright and Disclaimers.
Linked medical terms appearing on this page are added by Healia to help readers find more information and are not part of the original PubMed document.
The data herein was last updated on July 8th, 2008 and may not reflect the most current and accurate data available from NLM.