Transgenic expression of BACH1 in vivo. (A) Northern blot analysis of BACH1 mRNA using total RNA extracted from the spleens of the wild-type and transgenic mouse littermates shows transgene expression in transgenic mouse lines 701, 705, and 706. (B) Northern blot analysis of BACH1, GATA-1, and p45 mRNAs using total RNA derived from various tissues in the transgenic (Tg) and wild-type littermates. Note that BACH1 mRNA was detected only in the spleens of the transgenic mice, whereas the levels of GATA-1 and p45 expression in the BACH1 transgenic mice were consistent with those of their wild-type littermates. (C) Immunoblotting analysis of BACH1 using tissue extracts from the transgenic mice (705, 706) and their wild-type littermates. Extracts were prepared from the spleens, livers, and QT6 cells that were transfected with BACH1 expression vector. Tissue extracts (20 μg) and 2 μg QT6 protein were separated by 7% SDS-PAGE. After electrophoresis, samples were incubated with anti-Bach1 antisera (A1-5).17 Bound antibody was detected by a peroxidase-conjugated secondary antibody to rabbit immunoglobulin. After stripping off the A1-5 antibody, subsequent immunoblot analysis was performed using an anti–β-tubulin antibody as a control for the amount of protein loaded in each lane.
