Fig. 1. Targeted inactivation of the Hnf1ß gene in the liver. (A) Strategy used to generate the conditional allele, featuring the map of the murine locus, the recombinant allele Hnf1ß^3lox and the Cre-mediated alleles Hnf1ß^flox and Hnf1ß^del. The DNA fragment used as probe for the genomic analysis is indicated as well as the primers used for genotyping. The first exon is featured by a gray box, the selection genes by white boxes, the loxP sites by triangles, and the promoter by an arrow. Restriction sites: Bm BamHI; H, HindIII; P, PmlI; R, EcoRI; X, XbaI. (B,C) Cell specificity of Cre-mediated recombination in newborn AlfpCre; R26R mice. (B) ß-galactosidase staining on vibratome section of liver, showing the specific activation of the lacZ transgene in the hepatic parenchyma (hp) and in the bile ducts (bd) but not in the wall of the portal vein (pv) and of the hepatic arterioles (ha). (C) Thin section through a gallbladder after ß-galactosidase staining. Arrows indicate the staining of fields of the inner epithelium. lu, lumen. (D) Southern blot analysis of Cre-mediated recombination on liver DNA. Liver DNA from mice of either mutant (Hnf1ß^lacZ/floxAlfpCre) or control genotypes (Hnf1ß^flox/+, Hnf1ß^flox/+AlfpCre or Hnf1ß^flox/lacZ) were digested by the restriction enzyme HindIII and probed with the KH probe indicated on Fig. 1A. Bands characteristics for the different alleles are indicated on the right. Del* indicates a product of partial digestion specifically observed for the Hnf1ß^del allele. A rate of 80% conversion of the Hnf1ß^flox allele into the Hnf1ß^del form was estimated, by semi-quantitative PCR, for the Cre-positive animals. As hepatocytes represent about 70% of liver cells and biliary tree cells approximately 10%, we concluded that Hnf1ß inactivation was achieved in a majority of hepatic cells. (E) Northern blot analysis of 1-month-old mutant and control liver RNA, showing the complete loss of HNF1ß expression after Cre recombination. Total RNA was hybridized with HNF1ß cDNA and after stripping probed for HPRT expression to check for loading. Scale bars: 100 µm.