Fig. 2. Allelic series of morphology and expression levels in three lines of TRE-Klf4 mice that were crossed with K5-tTA mice. (A) Gross morphology and skeletal preparations of wild-type and the three lines of K5-Klf4 DTs. (B) Northern blot analysis of mRNAs isolated from E16.5 embryonic skin from the line indicated and hybridized with a Klf4 cDNA probe. The endogenous Klf4 is 3.5 kb and the ectopic Klf4 is at 2.5 kb in the K5-Klf4 DTs. Lines 6 and 10 show a faint band at 3 kb that may be an alternatively spliced form of the endogenous Klf4 gene. G3PDH probe is a control and signals are quantified using Phosphorimager. (C) Western blot analysis of proteins isolated from E16.5 embryonic skin from the line indicated and hybridized with anti-KLF4 polyclonal antibody. The endogenous KLF4 protein is 60 kDa and the ectopic KLF4 protein has a slightly higher molecular mass because of the N-terminal tag. Hybridization with p84 antibody, that recognizes an 84 kDa nuclear matrix protein, is a control.