Figure 2

pRosa26-DEST derived constructs are activated by expression of Cre. (a) Screening scheme of pRosa26-DEST targeted embryonic stem (ES) clones via Southern blot. Note that the size of the detected fragment after activation of the construct with Cre depends on the exact position of an EcoRV site in the cDNA in the construct; if no EcoRV is present the 11 kb band found in wildtype cells will become longer. (b) Southern blot confirmation of correctly targeted and activated S33Y β-catenin and lacZ ES clones. (c) Western blot analysis of S33Y β-catenin and lacZ ES cells before and after activation of the construct using transient expression of Cre. (d) β-catenin activity measured as the ratio between TopFlash and FopFlash signals in S33Y β-catenin and lacZ ES cells. (e) Demonstration of specificity of the human and murine β-catenin TaqMan assays using absolute dilution curves of both amplification products. (f) Absolute quantification of β-catenin expression from endogenous locus (mouse β-catenin) and Rosa26 locus (human β-catenin).