Fig. 2. Effective elimination of E-cadherin and ß-catenin sequences encompassed by loxP sites in oocytes, and characteristics of the floxed allele of ß-catenin. (A) Genotypes of some pups from females producing oocytes lacking E-cadherin. The PCR product characterizing the floxed-deleted E-cadherin allele inherited from the mother is indicated. The internal control is an unrelated wild-type product used as an indicator of successful PCR reactions. Genotypes of the control DNA are indicated at the top of each lane. (B) Genotypes of some pups from females producing oocytes lacking ß-catenin. The upper, floxed-deleted ß-catenin allele inherited from the female is indicated, as well as the lower wild-type ß-catenin allele inherited from the male. The middle product representing the floxed allele is clearly absent in the DNA of the pups analyzed. Genotypes of control DNA are indicated at top of each lane. (C) RT-PCR using primers specific for the 3' sequences of ß-catenin encoding the C-terminal part of the protein. A PCR replicon is detectable in all the embryos ostensibly lacking maternal ß-catenin (i.e. O, Z and ^e2; ß^F/ß^F; cre/Ø; top panel), as well as in the control embryos (ß^F/ß^F; Ø/Ø; bottom panel). Mitochondrial ATP synthase (mt-Atp6) primers were used as a control in both cases. O, ovulated oocyte; Z, zygote; ^e2, early two-cell embryo; ^l2, late two-cell embryo; ^l4, late four-cell embryo; 5/7, five- to seven-cell embryo; 8, eight-cell embryo; M, morula; B, blastocyst. (D) Western blot analysis using the polyclonal ß-catenin antibody recognizing the C-terminal part of the protein. ß^F/+ indicates extract obtained from a control animal containing one floxed ß-catenin allele and one wild-type allele. ß^F-del/+ indicates extract from a heterozygous animal containing a deleted-floxed allele and a wild-type allele. ß-catenin, wild-type ß-catenin; ^–N-ß-catenin, 52 kDa protein recognized by the polyclonal ß-catenin antibody. (E) Schematic representation of the interaction of ß-catenin with different binding partners (not to scale). The N-terminal and C-terminal are depicted by rectangular black boxes with an N or C, respectively. The 12 armadillo repeats are depicted by square, numbered black boxes. The regions where specific proteins interact with ß-catenin are depicted by brackets, with the name of the protein indicated. The large gray box indicates the part of ß-catenin absent in the floxed ß-catenin (^–N-ß-catenin or truncated) allele. Ebi, Ebi; ß-Trcp, ß-transducin repeat containing protein; Gsk3ß, glycogen synthase kinase 3ß; CKI, casein kinase I; APC, adenomatous polyposis coli; BRG1, SMARCA4; XSox3/17, Xenopus Sox3 or 17; ICAT, CATNBIP1, catenin beta interacting protein 1; CBP/p300, CREBBP, CREB binding protein/E1A binding protein p300; SDCCAG33, serologically defined colon cancer antigen 33.