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. 2011 Nov 24;1(2):120–127. doi: 10.1242/bio.2011048

Fig. 1. Generation of WNK3 knockout mice.

Fig. 1.

(A) Targeting strategy for Wnk3 gene interruption. The diagram shows the targeting construct, the wild-type WNK3 locus, and the targeted locus before and after Cre recombination. Three loxPs were inserted to flank exon 2 and the LacZ-Neo-selective marker. Exon 2 was deleted by mating the flox mice with CAG promoter Cre recombinase mice. (B) Verification of homologous recombination by PCR of genomic DNA derived from tails of mice. The primer set is indicated by dotted arrows in A. The 227-bp band and 339-bp band represent the wild-type allele and flox allele, respectively. (C) Genotyping PCR after Cre recombination, using a primer set indicated by solid arrows in A. A 516-bp PCR product was specific to the mutant allele. (D) Immunoblot of brain, kidney and testis homogenates probed with anti-WNK3 antibody. Absence of WNK3 protein in WNK3 knockout mouse was confirmed by immunoblotting in brain and testis. WNK3 was not detected by immunoblotting, even in wild-type mouse kidney, due to the low level of WNK3 protein expression in the kidney. (E) RT-PCR of brain, kidney and testis of wild-type and WNK3 knockout mice.