Targeted insertion of Cre recombinase into the TNAP gene: Excision in primordial germ cells

Cell type-specific genomic alterations afforded by the Cre/loxP system are a powerful approach to study the function of genes in particular cell lineages and for which generalized null mutations result in early lethal phenotypes (Lobe and Nagy, 1998). Here we report the creation of a primarily PGC (primordial germ cell) specific Cre recombinase transgenic line, designated TNAP-Cre. In this strain, the Cre recombinase has been knocked into the locus of the TNAP (Tissue Non-Specific Alkaline Phosphatase) gene. The structure of the TNAP-Cre targeting construct is depicted in Figure 1a. After electroporation into R1 ES (Nagy et al., 1993a) cells, about 30% of the clones screened by Southern analysis had undergone homologous recombination (Fig. 1b). Two clones were used to derive germ-line chimaeras (Nagy et al., 1993b). Both lines gave germline transmission and the two transgenic lines did not differ from each other in any aspects studied. Our heterozygous TNAP-Cre mice on a hybrid background did not show any abnormality, in agreement with previous report of the null mutation of the TNAP gene (MacGregor et al., 1995). To test the in vivo specificity of excision achieved by the TNAP-Cre mouse lines, we crossed them with the double-reporter line, Z/AP (Lobe et al., 1999).