The protein product of the Mcl-1 mini-transgene is prominently expressed in hematolymphoid tissues. (A) The Mcl-1 mini-gene construct used in the generation of transgenic mice (pSS17.5 plasmid) contains all exons (▪, including the 3′-untranslated region [▧] and introns (□) of human Mcl-1. It also contains Mcl-1 5′- and 3′-genomic flanking regions (▨). Fragments 1, 2, and 3 are genomic subclones used in making the transgene construct. Restriction sites are indicated as follows: S, SalI; X,XhoI; B, BamHI; and Xb, XbaI. The BS3.0 genomic subclone shown was used as probe to identify transgenic mice by Southern blotting. (B) Expression of the Mcl-1 transgene was assayed in a variety of tissues by Western blotting. ML-1 cells incubated with 5 × 10^-10 mol/L TPA for 3 hours served as a positive control. Each lane represents 50 μg of protein, except lane 13 which represents 100 μg of protein. (C) Expression of the human Mcl-1 transgene in the spleen and lymph node (L.N.) of a transgenic mouse (Tr.) was assayed in parallel with human lymph node. ML-1 cells incubated with 5 × 10^-10 mol/L TPA for 3 hours served as a positive control. Each lane represents 100 μg of protein, except lane 4 which represents 50 μg of protein. (D) Expression of the Mcl-1 transgene was assayed in enriched T- and B-cell populations from transgenic mouse spleen. Unfractionated spleen from a transgenic (Tr.) and a nontransgenic (Non-Tr.) mouse, as well as ML-1 cells incubated in the presence or absence of 5 × 10^-10 mol/L TPA for 3 hours served as controls. Each lane represents 50 μg of protein. (E) Expression of the Mcl-1 transgene was assayed in cell lines derived from cells from the spleen or bone marrow of transgenic mice. Lanes 1 through 3 (mast cell lines) represent 1 × 10⁶ cells and lane 4 represents 5 × 10⁵ cells. Lanes 5 and 6 (monocytic cell lines) and lane 7 represent 50 μg protein.