Figure 1.

Targeted disruption of the mouse hausp gene. (a) Diagram of the hausp genomic region containing exon 14 is shown with restriction fragments of BamHI and XbaI indicated. Targeting vector contains IRES-lac Z and a neo cassette was inserted into exon 14, flanking with 1.7 kb 5′-homology and 3.5kb 3′-homology (thicker lines). The diagram of the targeted mutant allele is shown, with altered restriction fragments of BamHI and XbaI after correct gene targeting. (b) Genotyping by Southern blot using genomic DNA digested by BamHI and XbaI, respectively. Hybridization with a 5′-external probe detected an 11.7kb wild-type band and an additional 7.6-kb mutant band in heterozygote mouse. Hybridization using 3′-external probe detected an 11.6 wild-type band and a 5.6-kb mutant band. (c) Genotyping by PCR showed a 202-bp band for wild-type allele and a 433-bp band for mutant allele, simultaneously. (d) The Hausp protein is absent in hausp knockout embryo. Protein extracts prepared from wild-type embryo and hausp knockout embryo at day E8.5, were analyzed by western blot using an anti-Hausp polyclonal antibody and an anti-β-actin monoclonal antibody. The Hausp protein was shown absent in hausp knockout embryo, whereas the total proteins, indicated by β-actin level, were comparable. B, BamHI; X, XbaI.