Generation of Mll5^tm1Apa mice. (A) Schematic representation of the genomic structure of the portion of Mll5 locus between PvuII restriction enzyme sites (top), the targeting vector (middle), and the portion of the targeted allele between PvuII sites (bottom). Exons are represented by solid boxes and LoxP sequences as triangular flags. The positions of primers for genotyping analysis (arrowheads labeled hefF, hetR, and Asc403) and probes used for Southern blots (double-headed arrows) are indicated. The Neo gene under the controls of the MC promoter is flanked by LoxP sequences. The IRES-LacZ cassette was designed to be driven off the endogenous Mll5 promoter. (B) Southern blot of genomic DNA from Mll5^tm1Apa homozygous (−/−), Mll5^tm1Apa heterozygous (+/−), and wild-type (+/+) mice cut with PvuII and hybridized with the external probes as indicated in A. (C) Mice were genotyped by PCR with the primers (indicated in A) and a representative gel is shown. Amplification with primers hetF and hetR (wild-type allele) yielded a 191-bp product, whereas amplification with primers Asc403 and hetR (recombinant allele) yielded a 442-bp product. (D) Schematic representation of the murine Mll5 protein depicting the domains of the mouse Mll5 protein and the truncation (labeled KO, if the protein were to be translated) of knockout protein. The epitope of the antibody 31 959 is also indicated. (E) Western blot of thymus and spleen from −/− and +/+ animals, blotted with anti-Mll5 polyclonal antibody (top panel; 31 959 epitope indicated in D) or anti-ACTIN (Santa Cruz Biotechnology, Santa Cruz, CA) antibody (bottom panel).