Independent integration of two inducible HNF4α proteins. A: Western blot analysis was performed on the 19-2 parent cell line containing the myc tagged HNF4α2 coding region in the FRT site and on the four ΦC31 mediated descendent cell lines 19-2-2, 19-2-6, 19-2-4, and 19-2-5. The anti-GFP antibody recognizes ECFP. B: Western blot analysis of the cell lines 19-2-2, 19-2-6, 19-2-4, and 19-2-5 using an anti-HNF4α or anti-myc antibody. The cells were cultured in six well plates, incubated with 1 μg/ml doxycycline and/or 1 μM Shld1 as indicated and protein extracts prepared after 24 h. As the HNF4α transgene introduced into the FRT docking site has a myc-tag, it can be detected specifically with the anti-myc antibody. The anti-HNF4α antibody detects the myc-tagged as well as the DD-tagged HNF4α proteins that differ slightly in size (compare e.g. lanes 2 and 3).