Gene trapping

Gene trapping is a high-throughput approach that is used to introduce insertional mutations across the mammalian genome. It is performed with gene trap vectors whose principal element is a gene trapping cassette consisting of a promoterless reporter gene and/or selectable genetic marker flanked by an upstream 3’ splice site (splice acceptor; SA) and a downstream transcriptional termination sequence (polyadenylation sequence; polyA). When inserted into an intron of an expressed gene, the gene trap cassette is transcribed from the endogenous promoter of that gene in the form of a fusion transcript in which the exon(s) upstream of the insertion site is spliced in frame to the reporter/selectable marker gene. Since transcription is terminated prematurely at the inserted polyadenylation site, the processed fusion transcript encodes a truncated and nonfunctional version of the cellular protein and the reporter/selectable marker. Thus, gene traps simultaneously inactivate and report the expression of the trapped gene at the insertion site, and provide a DNA tag (gene trap sequence tag, GTST) for the rapid identification of the disrupted gene. An international public consortium International Gene Trap Consortium is centralizing the data and cell lines can be requested from them.

References

Gossler, A., A. L. Joyner, et al. (1989). "Mouse embryonic stem cells and reporter constructs to detect developmentally regulated genes." Science 244(4903): 463-5.
von Melchner, H. and H. E. Ruley (1989). "Identification of cellular promoters by using a retrovirus promoter trap." J Virol 63(8): 3227-33.
Friedrich, G. and P. Soriano (1991) "Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice." Genes Dev 5:1413-1523.
Zambrowicz, B. P., G. A. Friedrich, et al. (1998). "Disruption and sequence identification of 2,000 genes in mouse embryonic stem cells." Nature 392(6676): 608-11.
Wiles, M. V., F. Vauti, et al. (2000). "Establishment of a gene-trap sequence tag library to generate mutant mice from embryonic stem cells." Nat Genet 24(1): 13-4.
Gene Trap Mutagenesis past, present & beyond; Stanford WL, Cohn JB, Cordes SP (2001)Nature Rev. Genet., 2, 756–768
Skarnes, W. C., H. von Melchner, et al. (2004). "A public gene trap resource for mouse functional genomics." Nat Genet 36(6): 543-4.
Hansen, J., T. Floss, et al. (2003). "A large-scale, gene-driven mutagenesis approach for the functional analysis of the mouse genome." Proc Natl Acad Sci U S A 100(17): 9918-22.
Zambrowicz, B. P., A. Abuin, et al. (2003). "Wnk1 kinase deficiency lowers blood pressure in mice: A gene-trap screen to identify potential targets for therapeutic intervention." Proc Natl Acad Sci U S A 100(24): 14109-14.
Schnutgen, F., S. De-Zolt, et al. (2005). "Genomewide production of multipurpose alleles for the functional analysis of the mouse genome." Proc Natl Acad Sci U S A 102(20): 7221-
De-Zolt, S., F. Schnutgen, et al. (2006). "High-throughput trapping of secretory pathway genes in mouse embryonic stem cells." Nucleic Acids Res 34(3): e25.
Patricia S. Springer (2000). "Gene Traps: Tools for Plant Development and Genomics." The Plant Cell, Vol. 12:1007-1020.

This article is issued from Wikipedia. The text is licensed under Creative Commons - Attribution - Sharealike. Additional terms may apply for the media files.