Fig. 1. Targeted disruption of the Cdx4 gene. (A) The gene-targeting vector inserts an antisense floxed PGK-hygro^r cassette into an EcoRV site upstream of exon 2 and an antisense flipped PGK-neo^r cassette, including a single loxP site into an EcoRI site downstream of exon 3, resulting in a conditional allele of Cdx4 (Cdx4^c). Cre-mediated deletion of the homeodomain in exon 2 and exon 3 generates the Cdx4-null allele (Cdx4^-). LoxP and FRT sites are represented by closed and open red triangles, respectively. HIII, HindIII; KI, KpnI; RI, EcoRI; RV, EcoRV; SI, SpeI. (B) Southern blot analysis of male ES cell clones with a unique 5' genomic probe reveals a 17 kb KpnI fragment generated by the targeted conditional allele. A 3' internal probe reveals correct 3' targeting by identification of a 17 kb KpnI fragment. (C) Histological evaluation of the digestive tract of adult Cdx4-null and control (wild-type) mice. Ki67 immunohistochemistry shows normal proliferation in the intestinal and colonic crypts (brown cells). Unlike previous findings in Cdx2^+/- mice, no intestinal polyp-like lesions were observed.