Figure 1. Efficacy of Recombination and Shh Removal Using a Hoxb6CreERT Deleter Line.
(A) Cre+;LacZ reporter-positive embryos show substantial recombination at 8 hr and essentially complete recombination by 12 hr after tamoxifen (Tam) injection.
(B) Tam-induced deletion of Shh prior to the normal onset of expression in hindlimb reproduces a null mutant phenotype.
(C) Shh (exon2-deleted) and Ptc1 RNA are undetectable in limb buds at 18 hr after Tam-induced Shh removal. RNAs analyzed at both 18 hr and at 24 hr after each of the Tam injection times gave the same results. Examples shown were collected at 18 hr, except E9.5- and E10-treated Shh, and E10-treated Ptc1, which were at 24 hr. Ptc1 transcripts were occasionally weakly detected in forelimb bud after Tam at E10.5 (arrow). Controls used were Shh+/flox;Cre+ and Shh+/Δ;Cre− sibling embryos; both showed similar Shh and Ptc1 expression.
(D) Blue timelines summarize normal duration of Shh activity (Ptc1 detection; see also Buscher et al., 1997; Platt et al., 1997; Lewis et al., 2001) and duration after Tam-activated Cre removal of Shh, based on a detailed time course of Shh and Ptc1 expression (data from Figure S1) assayed at 3–5 hr intervals after different Tam injection times (marked by arrowheads).