Fig. 1. Generation of mice deficient for PTEN in a tissue specific fashion: (A) organisation of the PTEN wild-type locus; (B) targeting construct used for electroporation into ES cells; (C) targeted locus containing the neomycin resistance gene flanked by LoxP sites; (D) ES cells lacking the neo cassette were generated by transient cre expression, resulting in a Pten^LoxP allele; (E) tissue-specific Cre-mediated recombination results in loss of exon 5. (F) Southern blot analysis of targeted clones after homologous recombination (BglII digest, probe 1; exon 4) shows a 6.6 kb wild-type band and a 5 kb band of the recombined allele in lane 1. (G) Probing with probe 2 (exon 5) of NcoI/EcoRI-digested genomic DNA resulted in a 6 kb wild-type and a 2 kb recombined band in lanes 1+2. (H) Southern blot analysis (probe 2, NcoI/EcoRI-digested genomic DNA) after transient transfection with a Cre-expressing plasmid shows a 6 kb wild-type band and a 2 kb floxed exon 5 band. (lane 3). (I) Loss of the neo cassette was confirmed after re-probing the blot with a neo probe (lane 3).