Targetted correction of a mutant HPRT gene in mouse embryonic stem cells

Abstract

Two recent developments suggest a route to predetermined alterations in mammalian germlines. These are, first, the characterization of mouse embryonic stem (ES) cells that can still enter the germline after genetic manipulation in culture and second, the demonstration that homologous recombination between a native target chromosomal gene and exogenous DAN can be used in culture to modify specifically the target locus. We here use gene targetting functionally to correct the mutant hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene in the ES cell line which has previously been isolated and used to produce an HPRT-deficient mouse. This modification of a chosen gene in pluripotent ES cells demonstrates the feasibility of this route to manipulating mammalian genomes in predetermined ways.

The European dimension for the mouse genome mutagenesis program

“…Owing to the development of mouse embryonic stem (ES) cells 1,2 and a variety of mutagenic technologies [3][4][5][6][7][8][9][10][11][12] , it is now possible to plan a systematic assault on the mouse genome to document function. The most straightforward way to start this program is to create and characterize null alleles in the germ line.…”

Prix Nobel de Médecine 2007

“…Tirant profit de leurs travaux sur les cellules immortalisées, Mario Capecchi et Oliver Smithies montrèrent qu'il était possible de cibler le gène hprt des cellules ES, pour créer ou, au contraire pour corriger, des mutations nulles dans celui-ci [11][12][13]. Le gène Hprt a été choisi car des mutations aussi bien perte que gain de fonction pouvaient être directement sélectionnées en culture cellulaire.…”

Construction of transgenic swine with induced expression of Cre recombinase

“…Gene disruption is a powerful technique for studying gene function in vivo l (Doetschman et al, 1987). A common technique for ablating genes is the use of the Cre-loxP recombination system, 'Cre' being an abbreviation for the phrase 'causes recombination'.…”