Fig. 2. Generation of a floxed Fgfr2 allele. (A) Schematic representation of the Fgfr2 genomic locus, the targeting vector and the Fgfr2^flox allele following homologous recombination. The loxP sites are indicated by gray arrowheads. The length of diagnostic HindIII-EcoRV and EcoRV restriction fragments are indicated by solid lines. The probes (5' and 3') used for DNA bolt analysis and the primers (F1, F2 and F3) used for PCR genotyping are indicated by filled boxes and arrowheads, respectively. The allele, generated by CRE-mediated recombination to delete all sequences between the two loxP sites, is shown at the bottom. (B) Southern blot analysis of EcoRV-digested genomic DNA from embryonic stem cells hybridized with the 3' probe. (C) Southern blot analysis of HindIII-EcoRV-digested genomic DNA from embryonic stem cells hybridized with the 5' probe. (D) PCR analysis of tail DNA (genotype is shown on the left). Primer F1 and F2 distinguish the wild-type (142 bp) and Fgfr2^flox (207 bp) alleles (bottom panel). Primer F1 and F3 produce a 471 bp fragment from the Fgfr2 allele (middle panel). The top panel shows PCR analysis of the Dermo1^cre allele with primer D1 and D2 using the same DNA samples. A 370 bp fragment is produced from the Dermo1^cre allele (lanes 1, 3 and 5). Since D2 is localized in the 5' end of the cre sequence, no PCR product is amplified from the wild-type allele (lanes 2 and 4). Note that the 471 bp Fgfr2 PCR fragment is amplified from Fgfr2^+/flox; Dermo-1^cre/+ tail DNA (lane 3). This is due to co-expression and deletion of the Fgfr2^flox allele by Dermo1-CRE in tail tissue. B, BamHI; C, ClaI; E, EcoRI; H, HindIII; Rv, EcoRV; Xb, XbaI.

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