Figure 2.

Knockout mice do not express protein from the deleted locus. (A) Total RNA from the brain and testes (n = 4/genotype) were assessed for expression of parkin and Pacrg. Expression of parkin and pacrg was not detected in the double parkin-Pacrg knockout (dKO), and expression of Pacrg was not detected in the single Pacrg knockout (sKO) but the full length transcripts were detected in wildtype (WT) tissues. Amplification of mRNA from the unrelated gene encoding RAN binding protein 9 (RanBP9) was performed in parallel to confirm RNA integrity. (B) Whole protein lysates from the brain and testes (n ≥ 3/genotype) were used to investigate steady-state parkin and PACRG levels in WT, dKO and sKO mice. Western blot analysis using an anti-parkin antibody detected parkin in WT and sKO tissues but not in dKO tissues, while an anti-PACRG antibody detected PACRG only in the WT tissue. The membranes were reprobed with an anti-β-Actin antibody to confirm protein integrity and equivalent loading. *Non-specific band.