Fig. 1. Targeting of the LRE and RARE in the mouse Cdx1 promoter. (A) Schematic of the wild-type (WT) Cdx1 locus, targeting vector, targeted allele and Cre-recombined allele. Probe A (5' external probe) was used to screen for targeted ES cells, probe B (5' internal probe) was used to confirm the predicted targeting event and probe C (3' internal probe) was used to confirm excision of the floxed neomycin selection cassette. E, EcoRI; H, HindIII; K, KpnI; S, SacI; E1, Exon 1; ^*, mutated RARE; ++, mutated LRE. (B) Southern blot analysis of DNA from wild-type (left), heterozygous targeted (middle) and heterozygous Cre-recombined (right) offspring using probe B and the indicated restriction endonucleases (see A). Concomitant integration of mutated LRE and LRE+RARE within the targeted allele was determined by the introduction of novel restriction sites: for the RARE mutation, a SacI restriction site was observed by Southern blot analysis (last lane of middle and right panels); for the LRE mutation, novel SfuI restriction sites were assessed by restriction of a PCR product spanning the LRE (lower left panel; see also Materials and methods).