FIG. 1.

Generation of Nck1- and Nck2-deficient mouse strains. (A) Schematic representation of the gene-targeting strategies used to mutate the Nck genes. The restriction maps of the Nck1 and Nck2 loci are indicated (B, BamHI; E, EcoRI; X, XbaI). The Nck1^IRES-lacZ targeting vector included an IRES-lacZ reporter gene cassette. The Nck2^tau-lacZ targeting vector comprised a tau-lacZ reporter gene fused in frame with the first coding exon. The protein region employed to raise the 3x3 polyclonal antibody used for Western blot analysis is indicated. (B) Genotypic analysis of the mutant mouse strains by PCR. Primer combinations are indicated in the Methods. The WT and mutant bands were, respectively, of 210 and 280 bp for Nck1, 250 and 610 bp for Nck2^ko, and 170 and 280 bp for Nck2^tau-lacZ mutant mouse strains. (C) Expression of the Nck genes was assessed by Western blotting of protein extracts of MEFs with the indicated genotypes. Western blots of protein extracts of mutant MEFs infected with IRES-EGFP and myc-tagged Nck1-IRES-EGFP retroviral vectors are also shown.