Fig. 1. Gli1 gene targeting strategy and ES cell screening. (A, top) The Gli1 genomic locus and targeting construct. White boxes indicate Gli1 exons. Black boxes indicate Gli1 zinc-finger domains 1-5. B, BamHI; RV, EcoRV; H, HindIII; Hs, Hsp921; Xb, XbaI. (A, middle) Targeted allele with neo cassette. The N-terminal and zinc-finger domains are replaced by nuclear lacZ and a neo cassette. The neo cassette is in the opposite orientation relative to Gli1 transcription. (A, bottom) Targeted allele without neo allele. Cre recombinase was used to remove the loxP-neo-loxP cassette. (B) ES cell Southern blot hybridization using 5' and 3' external probes indicated in A following EcoRV and XbaI digestion, respectively. The 5' probe identifies a 9 kb mutant fragment and a >20 kb wild-type fragment, whereas the 3' probe identifies a 4.5 kb mutant fragment and a 9 kb wild-type fragment.