Figure 2.

The targeted deletion in the Alox12b-gene generates a null mutation. (A) Detection of wild-type and mutant 12R-LOX mRNA in neonatal skin by RT-PCR. Primer combination ol230/ol709 was used to amplify a 346-bp product of the wild type and a 142-bp product of the deleted allele. (B) Loss of 12R-LOX protein in neonatal skin of 12R-LOX^−/− mice examined by Western blot analysis using a polyclonal antiserum raised against a 12R-LOX–specific peptide (top). As a control, the cell lysate of HEK 293 transfectants exogenously expressing m12R-LOX was used. An antibody recognizing actin was used as a control for equal loading (bottom). (C) In wild-type mice, immunostaining using a 12R-LOX–specific mAb shows prominent staining of the plasma membranes of the keratinocytes in the granular layers subjacent to the stratum corneum. In 12R-LOX^−/− mice, these structures appear negative for such a staining. Nuclei are counterstained with Hoechst 33258. sc, stratum corneum; sg, stratum granulosum; ss, stratum spinosum; sb, stratum basale. Bars, 20 μm.