Generation of transgenic mice expressing the Ikaros isoform IK5.

(A) Representation of the construct used to generate IK5 (bi-5) transgenic mice. The construct contains the lck proximal promoter and the Eμ immunoglobulin heavy-chain enhancer to direct expression of IK5. The IK5 cDNA lacks exons 4 to 6, which eliminates 3 of 4 possible DNA-binding zinc fingers, yet retains all of the C-terminal Zn⁺⁺ fingers. The HgX minigene has been shown to enhance the expression of transgenes and was part of the original p1026 promoter construct.9 (B) Northern blot of the 2 different bi-5 lines, 747 and 5516, using a probe containing exon 7 of Ikaros. Both endogenous Ikaros and transgene mRNA are marked in the figure. Cells used include thymocytes (T), splenocytes (Sp), and total bone marrow (M). The blot is slightly overexposed to better visualize marrow and spleen expression of the transgene. The HgX minigene, although not capable of being translated, contains both introns and exons; therefore, the transgene mRNA is expressed as multiple, overlapping bands. The lower panel shows the blot after stripping and reprobing for elongation factor-1α (EF) to account for loading differences. (C) Western blot using antisera against the C-terminal portion of Ikaros, which recognizes all Ikaros isoforms. Samples include protein isolated from the thymus (Thy) and liver (Liv) of bi-5 transgenic line 747 and a littermate control (wt). (D) Cell equivalent Western blot using 100 000 sorted cells per lane. Samples include DN and DP thymocytes and splenic T cells (Sp T) from a representative bi-5 mouse (Bi-5 Tg) and littermate control (littermate).