Nesprin 1α2 KO mice died perinatally. (A) Construct used for targeting the nesprin 1 gene, with exon 1α2 flanked by two LoxP sites (arrowheads). DTA, diphtheria toxin A; black rectangles, flippase recombination target sites; Neo, neomycin cassette. (B) Southern blot confirmation of the WT allele at 9.2 kb and the presence of the mutant allele (MU) at the predicted size of 7 kb. (C) Western blot of skeletal muscle lysates from WT (+/+) and nesprin 1α2 KO (−/−) mice using a nesprin 1 antibody. An arrowhead indicates the predicted molecular weight of nesprin 1α2. Note the absence of the band at the predicted molecular weight of nesprin 1α2. GAPDH served as a loading control. (D) Semiquantitative RT-PCR of WT and nesprin 1α2 KO mRNA isolated from skeletal muscle. Note that similarly to WT and as expected, other nesprin 1 isoforms were present in nesprin 1α2 KO mice; however, the nesprin 1α2 isoform was absent. (E) E18.5 embryos were extracted from pregnant females and imaged after 5 min. Note that WT embryos were able to breathe and turned pink shortly after delivery, whereas nesprin 1α2 KO embryos turned cyanotic (blue). Bars, 5 mm.