Skip to main content
. 2004 Jan 27;101(6):1725–1730. doi: 10.1073/pnas.0308217100

Fig. 1.

Fig. 1.

Targeted conditional deletion of Pten in murine skin, thymus, and prostate. (A) Southern-blot genotypic analysis for deletion of Pten in keratinocytes and thymus. Genomic DNA extracted from cultured keratinocytes and thymus tissue was digested with HindIII and hybridized with a probe situated upstream of exon 4 to show bands for wild-type (wt, 2-kb), nonrecombined (loxP, 6-kb), and recombined (Δ4-5, 2.3-kb) Pten alleles. DNA from keratinocytes and thymi is shown in Left and Right, respectively. Genotypes for each lane are as follows: 1 and 5, PtenloxP/loxP;2, Ptenwt/loxP;3,8,and9, PtenloxP/loxP; MMTV-cre; and 4 and 7, Ptenwt/loxP; MMTV-cre mice. Lane 9 is DNA from a thymic lymphoma. (B) Western blot analysis of cultured keratinocytes. Pten expression is lost in PtenloxP/loxP; MMTV-cre keratinocytes (Mut.) compared with Ptenwt/loxP; MMTV-cre control (Con.) keratinocytes. Weak expression of Pten in mutant samples indicates incomplete Cre-mediated recombination within the keratinocyte population. Phospho-PKB levels are elevated in PtenloxP/loxP; MMTV-cre keratinocytes compared with Ptenwt/loxP; MMTV-cre control keratinocytes under basal (10% FCS), serum-starved (0.5% FCS), and serum-stimulated conditions (Stim., starved and stimulated with 10% FCS). (C) PCR analysis for Cre-mediated recombination of the Pten locus (Δ4-5, Top), genotyping for Pten-loxP (Middle), and MMTV-cre (Bottom). Genomic DNA was isolated from the prostate of mice for PCR analysis, and products were run on an agarose gel. For Δ4-5 PCR, primers situated upstream of the 5′ loxP site and downstream of the 3′ loxP site amplified an 849-bp product only upon Cre-mediated deletion of Pten4-5. The Pten-loxP and wt alleles yielded products of 335 and 228 bp, respectively. Genotypes for each lane are as follows: 1, Ptenwt/loxP; 2, PtenloxP/loxP; 3–4, Ptenwt/loxP; MMTV-cre; and 5–8, PtenloxP/loxP; MMTV-cre mice. Positive control in lane 9 corresponds to PCR products from genomic DNA of Ptenwt/loxP; Gfap-cre brain tissue (16). Numbers on the left indicate size of DNA marker in base pairs.