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. 1997 Dec 1;25(23):4803–4807. doi: 10.1093/nar/25.23.4803

A gene trap approach to isolate mammalian genes involved in the cellular response to genotoxic stress.

P Menichini 1, S Viaggi 1, E Gallerani 1, G Fronza 1, L Ottaggio 1, A Comes 1, J W Ellwart 1, A Abbondandolo 1
PMCID: PMC147125  PMID: 9365260

Abstract

Treatment of cells with DNA damaging agents leads to induction of a variety of genes involved in different cellular processes. We have applied a lacZ-based gene trap strategy to search for new mammalian genes induced by genotoxic stress. A population of 32 x 10(3) neo r clones stably transfected with a gene trap vector was obtained, stained with fluorescein di-beta-d-galactopyranoside and analyzed by flow activated cell sorting and replica plating. This strategy allowed isolation of 30 neo r 'putative inducible' cell lines expressing lacZ only after a DNA damaging treatment. For three clones the site of integration and the degree of inducibility after UV treatment were determined by Southern blot and beta-galactosidase measurement respectively. One cell line (clone VI) showed a single integration site and a reproducible 3-fold induction of beta-galactosidase activity following UV irradiation. Fused transcripts were isolated from induced cells and a portion of the trapped gene was amplified by rapid amplification of cDNA ends. Sequence analysis and comparison with available gene and protein databanks revealed that the gene was novel.

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Selected References

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