Figure 1.
Generation of ROCK1-knockout allele. A, Schematic of structures of the WT ROCK1 allele, targeting vector, targeting allele, and ROCK1-knockout allele. The targeting vector containing loxP sites flanking exon1b of ROCK1 was injected into embryonic stem cells. The loxP-flanked exon 1b and pGK-Neo gene in correctly targeted clones were deleted by transfection with Cre. Shown are positions of neomycin resistance (neo), loxP sites (loxP); restriction enzyme sites for BglII (Bg), BamHI (B), StudI (S), and PestI (P); and 5′ and 3′ genomic probes. B, Southern blots showing the presence of WT (+) and deleted (−) ROCK1 allele. Genomic DNA was isolated from tails of WT (+/+), ROCK1+/− (+/−), and ROCK1−/− (−/−) mice, digested with BamHI, and subjected to Southern blots. C, Western blots showing the protein levels of ROCK1 and ROCK2. Proteins were extracted from hearts of WT (+/+), ROCK1+/− (+/−), and ROCK1−/− (−/−) mice and analyzed by Western blots using anti-ROCK1, anti-ROCK2, and anti-actin antibodies.