Figure 1.

 Targeted disruption of the caspase-2 gene. (A) The structure of the 3′ end of the mouse caspase-2 gene is depicted. The last four exons are shown as boxes, and the nucleotides are numbered starting from the ATG translation initiation codon. The enzymatic active site pentapeptide domain QACRG is indicated. The differential splicing events that produce two messages encoding the short or long form of caspase-2 are indicated. The position of the probe used for the genomic Southern blot analysis is shown. (B) RT–PCR analysis of spleen mRNA for caspase-2-deficient and wild-type mice. Sequences amplified are indicated at the top. A 300-bp fragment from actin mRNA was amplified as a control, and the caspase-2 cDNA sequence was amplified from nucleotide 901 to 1079 within the active site or from nucleotide 181 to 598 upstream of the active site. The positive control is a fragment amplified from a cloned caspase-2 cDNA; the negative control is a PCR reaction performed without the addition of template cDNA. The sizes of the amplified fragments are indicated at right. (C,D) Immunoblot analysis of tissues from wild-type and mutant mice. Immunoblot analysis of adult (C) (strain 72) and E11.5 (D) (strain 511) mouse tissues using a rabbit polyclonal antibody raised against the full-length recombinant caspase-2 protein. This antibody cross-reacts with two other unrelated proteins (44 and 33 kD).