GENE TRAP STRATEGIES FOR THE IDENTIFICATION OF DEVELOPMENTALLY IMPORTANT MURINE GENES

The availability of mouse embryonic stem (ES) cells,which have the, re markable ability to colonize the germline of an host embryo has opened the opportunity to transfer genetic modifications, selected in cultur ed cells, to the living animal. In this paper, is described a strategy of gene identification which uses:es particular constructs called tra pping vectors basically made;with a reporter gene, generally beta-gala ctosidase devoid of regulatory sequences, and with a cassette containi ng a selection marker. After transfer into ES cells, the reporter gene may be expressed only when integrated in the correct orientation into a transcription unit.beta-galactosidase expression can then be monito red not only in ES cells but also in chimeric mice, thereby uncovering the pattern of expression in vivo of the ''trapped'' endogenous gene. Thus the trapping strategy has, several virtues: (1) the reporter gen e reveals the expression profile of the gene in which the construct ha s been integrated; (2) the construct, which interrupts the transcripti on unit, behaves as a mutagen; (3) the synthesis of a fusion transcrip t made of endogenous and reporter gene sequences; facilitates the clon ing of the cDNA of the trapped gene. Due to these virtues, this strate gy has become a privileged way to identify genes, whose expressions de velopmentally regulated and therefore likely candidates to be importan t in development. More generally, this strategy could serve to tag any endogenous gene, and therefore provide a way of creating whole genome EST libraries.