Generating the ZASC1−/−mouse model. A.) Schematic representation of the ZASC1 genomic locus (top) and ZASC1 locus after inserting loxp sites via cassettes containing puromycin and neomycin resistance genes. White boxes represent exons. Black line represents intronic or other sequence. Black arrow indicates translational start site. eGFP with a splice acceptor sequence was inserted downstream of the Neomycin targeting in an effort to express eGFP from recombined ZASC1 gene. B.) Southern blot of Sph1-digested DNA showing homologous recombination at the ZASC1 locus in genomic DNA isolated from 129 J ES cells; the 8,213 bp band represents the ZASC1 genomic locus and the 5,041 bp band represents the floxed ZASC1 locus. C.) ZASC1 gene copy number measured by qPCR, normalized to GAPDH gene copy number, in animals with different ZASC1 genotypes. D.) ZASC1 RNA expression in whole blood cells from each ZASC1 genotype measured by RTPCR. Lower band in ZASC1−/− lane represents primer-dimer.