Targeted disruption and expression analysis of the PCFT gene. (A) Targeting strategy is shown, including relevant restriction sites used for confirmation of Southern hybridization analysis. Homologous recombination between the targeting vector and the PCFT gene replaced exons 1 to 3 with the selection cassette. The 5′ internal and 3′ external probes used for Southern hybridization analysis were designed inside and outside the target vector arms of homology, respectively. (B) Southern hybridization demonstrating proper targeting events in ESC clones. Clone 1D1 was selected for blastocyst injection; Lex2 indicates untransfected ESC DNA. (C) Expression of the PCFT gene was detected in ESCs and in all adult tissues by reverse-transcribed PCR, except bone and adipose.