FIG. 1.
Generation and biochemical characterization of bigenic IκBα-SR mice. (A) Transgenic mice encoding an N-terminal, HA-tagged IκBα-(SS32/36AA) superrepressor under the control of tetracycline response element (Tet-O) were crossed with transgenic mice expressing the tetracycline transactivator (tTA) under the control of the prion promoter-enhancer (Prp/tTA) to generate bigenic mice that express IκBα-SR in the absence of doxycycline. (B) IκBα-SR+ bigenic mice from three independently derived bigenic lines (FVBN/DBA), exhibit grossly normal hippocampal morphology as assessed by Nissl staining and calbindin immunostaining. (C) Doxycycline regulation of IκBα-SR transgene expression. HA-IκBα-SR expression in bigenic lines (lanes 1 to 6) in the absence and presence of doxycycline versus control Prp/tTA (asterisk denotes bigenic lines selected for further analysis) (lane 7) and pTet-O-HA-IκBα-SR (lane 8) singly transgenic lines. Bigenic litters were divided at 1 month of age and maintained for 28 to 31 days on a normal or doxycycline-supplemented diet (asterisk denotes transgenic lines selected for further analysis). (D) Evaluation of transgene expression in various tissues in IκBα-SR+ mice. (E) IκBα-SR transgene expression in distinct regions of the brain in an IκBα-SR+ mouse. (F) Levels of IκBα-SR protein expressed in the hippocampal region were similar to the levels of endogenous IκBα protein in both IκBα-SR− and control mice. (G) Detection of IκBα-SR transgene expression using indirect immunofluorescence with anti-HA antibodies (green fluorescence). Glial and neuronal cells were identified using with anti-GFAP or anti-MAP2 antibodies, respectively, with Alexa568-conjugated secondary antibodies (red fluorescence). Certain sections were counterstained with DAPI (blue fluorescence) to identify nuclear regions. All mice designated IκBα-SR− are on a diet supplemented with doxycycline to suppress transgene expression.